Synergistic Effect of Transient Receptor Potential Antagonist and Amiloride against Maitotoxin Induced Calcium Increase and Cytotoxicity in Human Neuronal Stem Cells.

PubMed

Boente-Juncal, Andrea; Vale, Carmen; Alfonso, Amparo; Botana, Luis M

2018-05-16

Maitotoxins (MTX) are among the most potent marine toxins identified to date causing cell death trough massive calcium influx. However, the exact mechanism for the MTX-induced calcium entry and cytotoxicity is still unknown. In this work, the effect of MTX-1 on the cytosolic free calcium concentration and cellular viability of human neuronal stem cells was evaluated. MTX elicited a concentration-dependent decrease in cell viability which was already evident after 1 h of treatment with 0.25 nM MTX; however, at a concentration of 0.1 nM, the toxin did not cause cell death even after 14 days of exposure. Moreover, the toxin caused a concentration dependent rise in the cytosolic calcium concentration which was maximal at toxin concentrations of 1 nM and dependent on the presence of extracellular calcium on the bathing solution. Several pharmacological approaches were employed to evaluate the role of canonical transient potential receptor channels (TRPC) on the MTX effects. The results presented here lead to the identification of the TRPC4 channels as contributors to the MTX effects in human neuronal cells. Both, the calcium increase and the cytotoxicity of MTX were either fully (for the calcium increase) or partially (in the case of cytotoxicity) reverted by the blockade of canonical TRPC4 receptors with the selective antagonist ML204. Furthermore, the sodium proton exchanger blocker amiloride also partially inhibited the calcium rise and the cell death elicited by MTX while the combination of amiloride and ML204 fully prevented both the cytotoxicity and the calcium rise elicited by the toxin.

xCT expression reduces the early cell cycle requirement for calcium signaling

PubMed Central

Lastro, Michele; Kourtidis, Antonis; Farley, Kate; Conklin, Douglas S.

2009-01-01

Calcium has long been recognized as an important regulator of cell cycle transitions although the mechanisms are largely unknown. A functional genomic screen has identified genes involved in the regulation of early cell cycle progression by calcium. These genes when overexpressed confer the ability to bypass the G1/S arrest induced by Ca2+- channel antagonists in mouse fibroblasts. Overexpression of the cystine-glutamate exchanger, xCT, had the greatest ability to evade calcium antagonist-induced cell cycle arrest. xCT carries out the rate limiting step of glutathione synthesis in many cell types and is responsible for the uptake of cystine in most human cancer cell lines. Functional analysis indicates that the cystine uptake activity of xCT overcomes the G1/S arrest induced by Ca2+- channel antagonists by bypassing the requirement for calcium signaling. Since cells overexpressing xCT were found to have increased levels and activity of the AP-1 transcription factor in G1, redox stimulation of AP-1 activity accounts for the observed growth of these cells in the presence of calcium channel antagonists. These results suggest that reduced calcium signaling impairs AP-1 activation and that xCT expression may directly affect cell proliferation. PMID:18054200

P2Y1 receptor antagonists mitigate oxygen and glucose deprivation‑induced astrocyte injury.

PubMed

Guo, Hui; Liu, Zhong-Qiang; Zhou, Hui; Wang, Zhi-Ling; Tao, Yu-Hong; Tong, Yu

2018-01-01

The aim of the present study was to elucidate the effects of blocking the calcium signaling pathway of astrocytes (ASs) on oxygen and glucose deprivation (OGD)‑induced AS injury. The association between the changes in the concentrations of AS‑derived transmitter ATP and glutamic acid, and the changes in calcium signaling under the challenge of OGD were investigated. The cortical ASs of Sprague Dawley rats were cultured to establish the OGD models of ASs. The extracellular concentrations of ATP and glutamic acid in the normal group and the OGD group were detected, and the intracellular concentration of calcium ions (Ca2+) was detected. The effects of 2'‑deoxy‑N6‑methyl adenosine 3', 5'‑diphosphate diammonium salt (MRS2179), a P2Y1 receptor antagonist, on the release of calcium and glutamic acid of ASs under the condition of OGD were observed. The OGD challenge induced the release of glutamic acid and ATP by ASs in a time‑dependent manner, whereas elevation in the concentration of glutamic acid lagged behind that of the ATP and Ca2+. The concentration of Ca2+ inside ASs peaked 16 h after OGD, following which the concentration of Ca2+ was decreased. The effects of elevated release of glutamic acid by ASs when challenged by OGD may be blocked by MRS2179, a P2Y1 receptor antagonist. Furthermore, MRS2179 may significantly mitigate OGD‑induced AS injury and increase cell survival. The ASs of rats cultured in vitro expressed P2Y1 receptors, which may inhibit excessive elevation in the concentration of intracellular Ca2+. Avoidance of intracellular calcium overload and the excessive release of glutamic acid may be an important reason why MRS2179 mitigates OGD‑induced AS injury.

Differential intracellular calcium influx, nitric oxide production, ICAM-1 and IL8 expression in primary bovine endothelial cells exposed to nonesterified fatty acids.

PubMed

Loaiza, Anitsi; Carretta, María D; Taubert, Anja; Hermosilla, Carlos; Hidalgo, María A; Burgos, Rafael A

2016-02-25

Nonesterified fatty acids (NEFAs) are involved in proinflammatory processes in cattle, including in the increased expression of adhesion molecules in endothelial cells. However, the mechanisms underlying these effects are still unknown. The aim of this study was to assess the effects of NEFAs on the intracellular calcium (Ca(2+) i) influx, nitric oxide production, and ICAM-1 and IL-8 expression in primary bovine umbilical vein endothelial cells (BUVECs). Myristic (MA), palmitic (PA), stearic (SA), oleic (OA) and linoleic acid (LA) rapidly increased Ca(2+) i. The calcium response to all tested NEFAs showed an extracellular calcium dependence and only the LA response was significantly inhibited until the intracellular calcium was chelated. The EC50 values for MA and LA were 125 μM and 37 μM, respectively, and the MA and LA effects were dependent on calcium release from the endoplasmic reticulum stores and on the L-type calcium channels. Only the calcium response to MA was significantly reduced by GW1100, a selective G-protein-coupled free fatty acid receptor (GPR40) antagonist. We also detected a functional FFAR1/GPR40 protein in BUVECs by using western blotting and the FFAR1/GPR40 agonist TAK-875. Only LA increased the cellular nitric oxide levels in a calcium-dependent manner. LA stimulation but not MA stimulation increased ICAM-1 and IL-8-expression in BUVECs. This effect was inhibited by GW1100, an antagonist of FFAR1/GPR40, but not by U-73122, a phospholipase C inhibitor. These findings strongly suggest that each individual NEFA stimulates endothelial cells in a different way, with clearly different effects on intracellular calcium mobilization, NO production, and IL-8 and ICAM-1 expression in primary BUVECs. These findings not only extend our understanding of NEFA-mediated diseases in ruminants, but also provide new insight into the different molecular mechanisms involved during endothelial cell activation by NEFAs.

Metabotropic and ionotropic glutamate receptors regulate calcium channel currents in salamander retinal ganglion cells

PubMed Central

Shen, Wen; Slaughter, Malcolm M

1998-01-01

Glutamate suppressed high-voltage-activated barium currents (IBa,HVA) in tiger salamander retinal ganglion cells. Both ionotropic (iGluR) and metabotropic (mGluR) receptors contributed to this calcium channel inhibition. Trans-ACPD (1-aminocyclopentane-trans-1S,3R-dicarboxylic acid), a broad-spectrum metabotropic glutamate receptor agonist, suppressed a dihydropyridine-sensitive barium current. Kainate, an ionotropic glutamate receptor agonist, reduced an ω-conotoxin GVIA-sensitive current. The relative effectiveness of selective agonists indicated that the predominant metabotropic receptor was the L-2-amino-4-phosphonobutyrate (l-AP4)-sensitive, group III receptor. This receptor reversed the action of forskolin, but this was not responsible for calcium channel suppression. l-AP4 raised internal calcium concentration. Antagonists of phospholipase C, inositol trisphosphate (IP3) receptors and ryanodine receptors inhibited the action of metabotropic agonists, indicating that group III receptor transduction was linked to this pathway. The action of kainate was partially suppressed by BAPTA, by calmodulin antagonists and by blockers of calmodulin-dependent phosphatase. Suppression by kainate of the calcium channel current was more rapid when calcium was the charge carrier, instead of barium. The results indicate that calcium influx through kainate-sensitive glutamate receptors can activate calmodulin, which stimulates phosphatases that may directly suppress voltage-sensitive calcium channels. Thus, ionotropic and metabotropic glutamate receptors inhibit distinct calcium channels. They could act synergistically, since both increase internal calcium. These pathways provide negative feedback that can reduce calcium influx when ganglion cells are depolarized. PMID:9660896

The new generation dihydropyridine type calcium blockers, bearing 4-phenyl oxypropanolamine, display alpha-/beta-adrenoceptor antagonist and long-acting antihypertensive activities.

PubMed

Liang, Jhy-Chong; Yeh, Jwu-Lai; Wang, Chia-Sui; Liou, Shwu-Fen; Tsai, Chieh-Ho; Chen, Ing-Jun

2002-03-01

A new series of dihydropyridine derivatives, bearing oxypropanolamine moiety on phenyl ring at the 4-position of the dihydropyridine base, were prepared. Oxypropanolamine was synthesized by replacing the phenolic OH of vanillin or other compounds, having a phenyl aldehyde group, with epichlorohydrin, followed by cleavaging the obtained epoxide compounds with tert-butylamine, n-butylamine or 2-methoxy-1-oxyethylamino benzene (guaiacoxyethylamine), respectively. Obtained various oxypropanolamine compounds, still remaining a phenyl aldehyde moiety, were then performed by Hantzsch condensation reaction with methylacetoacetate or ethylacetoacetate, respectively, to give our new series of dihydropyridine linked with the 4-phenyl ring. These compounds were evaluated for inotropic, chronotropic, and aorta contractility that associated with calcium channel and adrenoceptor antagonist activities. Dihydropyridine derivatives that with oxypropanolamine side chain on their 4-phenyl ring associated alpha-/beta-adrenoceptor blocking activities created a new family of calcium entry and the third generation beta-adrenoceptor blockers. Optimizing this research to obtain more potent alpha-/beta-adrenoceptor blocking and long-acting antihypertensive oxypropanolamine on the 4-phenyl ring of dihydropyridine series compounds was thus accomplished and classified as third generation dihydropyridine type calcium channel blockers, in comparison with previous short-acting type nifedipine and long-acting type amlodipine. We concluded that compounds 1a, 1b and 1g showed not only markedly high calcium-antagonistic activity but also the highest antihypertensive effect; compounds 1b, 1c, 1f, 1g, 1i and 1j induced sustained antihypertensive effects are major and attributed to their calcium entry and alpha-adrenoceptor blocking activities in the blood vessel due to their introduction of 2-methoxy, 1-oxyethylamino benzene moiety in the side chain on the 4-phenyl ring of dihydropyridine. Bradycardiac effects of all the compounds 1a-1j resulted from calcium entry and beta-adrenoceptor blocking, which attenuate the sympathetic activation-associated reflex tachycardia in the heart. We selected compound 1b as candidate compound for further pharmacological and pre-clinical evaluation studies.

SALT EFFECTS ON SWARMERS OF DUNALIELLA VIRIDIS TEOD

PubMed Central

Baas-Becking, L. G. M.

1931-01-01

1. Dunaliella viridis Teodoresco thrives equally well in solutions of NaCl 1 to 4 mol and pH 6 to 9. 2. The organism is sensitive to calcium and magnesium, especially in acid medium. 3. Calcium and magnesium are antagonistic. In a molar solution of NaCl the antagonistic relation Mg:Ca is 4 to 5. In a 4 molar solution of NaCl the proportion becomes many times as great (20:1). 4. Although the strains used in this investigation did not occur in sea water concentrates, the increase in the antagonistic ratio Mg:Ca in which they can live closely paralleled the changes in this ratio which take place when sea water evaporates. 5. The other organisms which occurred in the cultures each show a specific relation to Ca and Mg. 6. The size of the cells of Dunaliella does not decrease with increasing NaCl content. PMID:19872621

Differential regulation of smooth muscle contraction in rabbit internal anal sphincter by substance P and bombesin.

PubMed

Bitar, K N; Hillemeier, C; Biancani, P

1990-01-01

Substance P and bombesin induce contraction of isolated IAS smooth muscle cells by different intracellular mechanisms. The cells contracted in a dose dependent manner to both peptides. The kinetics of contraction were different. Substance P induced contraction peaked at 30 seconds and declined in a time dependent manner while bombesin induced contraction peaked at 30 seconds and was maintained for up to 8 minutes. The absence of extracellular calcium in the medium (0 calcium and 2 mM EGTA) had no affect on substance P induced contraction while it blocked bombesin induced contraction. Substance P induced contraction was blocked by the calmodulin antagonist W7 (10(-9)M) and was not affected by the PKC antagonist H7 (10(-6)M). Bombesin induced contraction was blocked by the PKC antagonist H7 and was not affected by the calmodulin antagonist W7. Our data indicate that substance P induces a transient contraction utilizing intracellular calcium and a calmodulin dependent pathway, while bombesin induces a sustained contraction utilizing calcium from extracellular sources and a calmodulin independent pathway.

[Effects of the monosaccharide derivative 8RN-DAGal on the putative P-type calcium channel expressed in Xenopus oocytes].

PubMed

Fournier, F; Charpentier, G; Lahyani, A; Bruner, J; Czternasty, G; Marlot, D; Ronco, G; Villa, P; Brule, G

1993-01-01

P-type calcium channels are expressed in Xenopus oocytes after injection of rat cerebellar mRNA. The FTX and omega-Aga-IVa toxins extracted from Agelenopsis aperta venom are known to inhibit the activity of this channel. The present results demonstrate that 8RN-DAGal is also a antagonist of P-type calcium channels. The inhibition of the current, obtained with Ba2+, as charge carrier, is voltage dependent.

Calcium-regulated in vivo protein phosphorylation in Zea mays L. root tips

NASA Technical Reports Server (NTRS)

Raghothama, K. G.; Reddy, A. S.; Friedmann, M.; Poovaiah, B. W.

1987-01-01

Calcium dependent protein phosphorylation was studied in corn (Zea mays L.) root tips. Prior to in vivo protein phosphorylation experiments, the effect of calcium, ethyleneglycol-bis-(beta-aminoethyl ether)-N-N' -tetraacetic acid (EGTA) and calcium ionophore (A-23187) on phosphorus uptake was studied. Calcium increased phosphorus uptake, whereas EGTA and A-23187 decreased it. Consequently, phosphorus concentration in the media was adjusted so as to attain similar uptake in different treatments. Phosphoproteins were analyzed by two-dimensional gel electrophoresis. Distinct changes in phosphorylation were observed following altered calcium levels. Calcium depletion in root tips with EGTA and A-23187 decreased protein phosphorylation. However, replenishment of calcium following EGTA and ionophore pretreatment enhanced phosphorylation of proteins. Preloading of the root tips with 32P in the presence of EGTA and A-23187 followed by a ten minute calcium treatment, resulted in increased phosphorylation indicating the involvement of calcium, calcium and calmodulin-dependent kinases. Calmodulin antagonist W-7 was effective in inhibiting calcium-promoted phosphorylation. These studies suggest a physiological role for calcium-dependent phosphorylation in calcium-mediated processes in plants.

Effects of isradipine and other calcium antagonists on arteriovenous-shunt flow in anesthetized rabbits and cats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hof, R.P.

The effects of vasodilators on arteriovenous (AV)-shunt flow was investigated in anesthetized cats and rabbits, using the tracer microsphere method. In cats, the calcium antagonist isradipine reduced AV-shunt flow; verapamil showed a similar tendency and nicardipine was without effect. Dihydralazine strongly increased, but nitroglycerin and dipyridamole decreased AV-shunt flow. In rabbits, the effects of isradipine and verapamil were similar to those seen in cats. Sodium nitroprusside had no effect, whereas prazosin, minoxidil, and the potassium-channel activator cromakalim increased AV-shunt flow. The contrasting effects of drugs sharing the same mechanism of action suggest that target-tissue selectivity is more important than themore » mechanism of action. An increase of AV-shunt flow is unlikely to be beneficial but could be associated with a number of undesirable side effects. It might negatively affect migraine sufferers and, if AV-shunt dilatation shows no tolerance development, it represents an unnecessary hemodynamic burden for the heart.« less

Rational use of calcium-channel antagonists in Raynaud's phenomenon.

PubMed

Sturgill, M G; Seibold, J R

1998-11-01

Raynaud's phenomenon (RP) is a peripheral circulatory disorder characterized by sudden episodes of digital artery spasm, often precipitated by cold temperature or emotional stress. Although the cause of RP is not fully known, it appears to involve inappropriate adrenergic response to cold stimuli. Treatment of RP is conservative in most patients, but in patients with severe disease includes the use of agents that promote digital vasodilation. The calcium-channel antagonists, particularly the dihydropyridine derivative nifedipine, are the most thoroughly studied drug class for the treatment of RP. Approximately two thirds of patients respond favorably, with significant reductions in the frequency and severity of vasospastic attacks. Nifedipine use is often limited by the appearance of adverse vasodilatory effects such as headache or peripheral edema. The newer second-generation dihydropyridines such as amlodipine, isradipine, nicardipine, and felodipine also appear to be effective in patients with RP and may be associated with fewer adverse effects.

Isoflurane-Induced Caspase-3 Activation Is Dependent on Cytosolic Calcium and Can Be Attenuated by Memantine

PubMed Central

Zhang, Guohua; Dong, Yuanlin; Zhang, Bin; Ichinose, Fumito; Wu, Xu; Culley, Deborah J.; Crosby, Gregory

2008-01-01

Increasing evidence indicates that caspase activation and apoptosis are associated with a variety of neurodegenerative disorders, including Alzheimer's disease. We reported that anesthetic isoflurane can induce apoptosis, alter processing of the amyloid precursor protein (APP), and increase amyloid-β protein (Aβ) generation. However, the mechanism by which isoflurane induces apoptosis is primarily unknown. We therefore set out to assess effects of extracellular calcium concentration on isoflurane-induced caspase-3 activation in H4 human neuroglioma cells stably transfected to express human full-length APP (H4-APP cells). In addition, we tested effects of RNA interference (RNAi) silencing of IP3 receptor, NMDA receptor, and endoplasmic reticulum (ER) calcium pump, sacro-/ER calcium ATPase (SERCA1). Finally, we examined the effects of the NMDA receptor partial antagonist, memantine, in H4-APP cells and brain tissue of naive mice. EDTA (10 mm), BAPTA (10 μm), and RNAi silencing of IP3 receptor, NMDA receptor, or SERCA1 attenuated capase-3 activation. Memantine (4 μm) inhibited isoflurane-induced elevations in cytosolic calcium levels and attenuated isoflurane-induced caspase-3 activation, apoptosis, and cell viability. Memantine (20 mg/kg, i.p.) reduced isoflurane-induced caspase-3 activation in brain tissue of naive mice. These results suggest that disruption of calcium homeostasis underlies isoflurane-induced caspase activation and apoptosis. We also show for the first time that the NMDA receptor partial antagonist, memantine, can prevent isoflurane-induced caspase-3 activation and apoptosis in vivo and in vitro. These findings, indicating that isoflurane-induced caspase activation and apoptosis are dependent on cytosolic calcium levels, should facilitate the provision of safer anesthesia care, especially for Alzheimer's disease and elderly patients. PMID:18434534

Mechanism of inhibition of net ion transport across frog corneal epithelium by calcium channel antagonists.

PubMed

Huff, J W; Reinach, P S

1985-01-01

In the isolated bullfrog cornea, three calcium channel antagonists had dose-dependent inhibitory effects on the Cl-originated short-circuit current (SCC). Their order of decreasing potency was bepridil, verapamil and diltiazem. One millimolar diltiazem inhibited the SCC by 98% and subsequent incubation with the calcium ionophore A23187 had no restorative effect. Increasing the bathing solution Ca concentration from 0.05 to 15 mM, however, decreased diltiazem's inhibitory efficacy. This antagonist depolarized the intracellular potential difference Vsc from -54 to -18 mV (tear:reference) and the voltage divider ratio FRo decreased from 0.58 to 0.30, suggesting an increase in basolateral membrane electrical resistance. Additional indication of a basolateral membrane effect by the drug was that preincubation with 10(-5) M amphotericin B in Cl-free Ringer's did not eliminate the inhibitory effect of the drug on the Na- and K-elicited SCC. In the absence of amphotericin B in Cl-free Ringer's (SCC = 0), 1 X 10(-3) M diltiazem depolarized the Vsc from -78 to -9 mV suggesting that the increase in basolateral membrane resistance was due to K channel blockade. Diltiazem (1 X 10(-3) M) significantly decreased cyclic AMP content; however, isoproterenol in the presence of the drug increased cyclic AMP fourfold without having any restorative effect on the inhibited SCC. Therefore, the inhibition of the Cl-originated SCC resulting from an increase in basolateral membrane K resistance is not caused by a decline in cyclic AMP content.(ABSTRACT TRUNCATED AT 250 WORDS)

[Systematic review on the short-term efficacy and safety of nicorandil for stable angina pectoris in comparison with those of β-blockers, nitrates and calcium antagonists].

PubMed

Hanai, Yuki; Mita, Mitsuo; Hishinuma, Shigeru; Shoji, Masaru

2010-11-01

Nicorandil significantly reducted the incidence of major coronary events in patients with stable angina in a long-term trial, although there are few reports on its short-term efficacy in the treatment and prevention of angina symptoms. We performed a meta-analysis of the short-term efficacy of nicorandil compared with antianginal drugs for stable angina. We selected 20 reports (vs. β-blockers, n=6; vs. nitrates, n=6; vs. calcium antagonists, n=8) of prospective controlled trials from MEDLINE, the Cochrane Library, and Japana Centra Revuo Medicina. The trials were short in duration (median 5 weeks). We combined the results using odds ratios (OR) for discrete data and weighted mean differences (WMD) for continuous data. Compared with antianginal drugs, nicorandil did not show significant reduction of angina episodes per week (vs. β-blockers, -1.50 [95% confidence interval (CI): -4.09, 1.09]; vs. nitrates, 0.22 [95% CI: -1.22, 1.65]; vs. calcium antagonists, -0.23 [95% CI: -1.37, 0.90]). Furthermore, there were no significant differences in time to ischemia (total exercise duration, time to 1-mm ST depression, time to onset of pain). Although the total numbers of adverse events with each antianginal drug were similar, heart rate and blood pressure were significantly decreased by calcium antagonists but not changed by nicorandil (8.09 [95% CI: 3.20, 12.98] and 8.64 [95% CI: 3.28, 13.99], respectively). Thus this study suggests that short-term therapy with nicorandil is as effective as standard therapy and that nicorandil can also be used as a first-line agent in patients with stable angina.

Angiotensin II modulates mouse skeletal muscle resting conductance to chloride and potassium ions and calcium homeostasis via the AT1 receptor and NADPH oxidase

PubMed Central

Cozzoli, Anna; Liantonio, Antonella; Conte, Elena; Cannone, Maria; Massari, Ada Maria; Giustino, Arcangela; Scaramuzzi, Antonia; Pierno, Sabata; Mantuano, Paola; Capogrosso, Roberta Francesca; Camerino, Giulia Maria

2014-01-01

Angiotensin II (ANG II) plays a role in muscle wasting and remodeling; however, little evidence shows its direct effects on specific muscle functions. We presently investigated the acute in vitro effects of ANG II on resting ionic conductance and calcium homeostasis of mouse extensor digitorum longus (EDL) muscle fibers, based on previous findings that in vivo inhibition of ANG II counteracts the impairment of macroscopic ClC-1 chloride channel conductance (gCl) in the mdx mouse model of muscular dystrophy. By means of intracellular microelectrode recordings we found that ANG II reduced gCl in the nanomolar range and in a concentration-dependent manner (EC50 = 0.06 μM) meanwhile increasing potassium conductance (gK). Both effects were inhibited by the ANG II receptors type 1 (AT1)-receptor antagonist losartan and the protein kinase C inhibitor chelerythrine; no antagonism was observed with the AT2 antagonist PD123,319. The scavenger of reactive oxygen species (ROS) N-acetyl cysteine and the NADPH-oxidase (NOX) inhibitor apocynin also antagonized ANG II effects on resting ionic conductances; the ANG II-dependent gK increase was blocked by iberiotoxin, an inhibitor of calcium-activated potassium channels. ANG II also lowered the threshold for myofiber and muscle contraction. Both ANG II and the AT1 agonist L162,313 increased the intracellular calcium transients, measured by fura-2, with a two-step pattern. These latter effects were not observed in the presence of losartan and of the phospholipase C inhibitor U73122 and the in absence of extracellular calcium, disclosing a Gq-mediated calcium entry mechanism. The data show for the first time that the AT1-mediated ANG II pathway, also involving NOX and ROS, directly modulates ion channels and calcium homeostasis in adult myofibers. PMID:25080489

Angiotensin II modulates mouse skeletal muscle resting conductance to chloride and potassium ions and calcium homeostasis via the AT1 receptor and NADPH oxidase.

PubMed

Cozzoli, Anna; Liantonio, Antonella; Conte, Elena; Cannone, Maria; Massari, Ada Maria; Giustino, Arcangela; Scaramuzzi, Antonia; Pierno, Sabata; Mantuano, Paola; Capogrosso, Roberta Francesca; Camerino, Giulia Maria; De Luca, Annamaria

2014-10-01

Angiotensin II (ANG II) plays a role in muscle wasting and remodeling; however, little evidence shows its direct effects on specific muscle functions. We presently investigated the acute in vitro effects of ANG II on resting ionic conductance and calcium homeostasis of mouse extensor digitorum longus (EDL) muscle fibers, based on previous findings that in vivo inhibition of ANG II counteracts the impairment of macroscopic ClC-1 chloride channel conductance (gCl) in the mdx mouse model of muscular dystrophy. By means of intracellular microelectrode recordings we found that ANG II reduced gCl in the nanomolar range and in a concentration-dependent manner (EC50 = 0.06 μM) meanwhile increasing potassium conductance (gK). Both effects were inhibited by the ANG II receptors type 1 (AT1)-receptor antagonist losartan and the protein kinase C inhibitor chelerythrine; no antagonism was observed with the AT2 antagonist PD123,319. The scavenger of reactive oxygen species (ROS) N-acetyl cysteine and the NADPH-oxidase (NOX) inhibitor apocynin also antagonized ANG II effects on resting ionic conductances; the ANG II-dependent gK increase was blocked by iberiotoxin, an inhibitor of calcium-activated potassium channels. ANG II also lowered the threshold for myofiber and muscle contraction. Both ANG II and the AT1 agonist L162,313 increased the intracellular calcium transients, measured by fura-2, with a two-step pattern. These latter effects were not observed in the presence of losartan and of the phospholipase C inhibitor U73122 and the in absence of extracellular calcium, disclosing a Gq-mediated calcium entry mechanism. The data show for the first time that the AT1-mediated ANG II pathway, also involving NOX and ROS, directly modulates ion channels and calcium homeostasis in adult myofibers. Copyright © 2014 the American Physiological Society.

Electrophysiological characterization of activation state-dependent Ca(v)2 channel antagonist TROX-1 in spinal nerve injured rats.

PubMed

Patel, R; Rutten, K; Valdor, M; Schiene, K; Wigge, S; Schunk, S; Damann, N; Christoph, T; Dickenson, A H

2015-06-25

Prialt, a synthetic version of Ca(v)2.2 antagonist ω-conotoxin MVIIA derived from Conus magus, is the first clinically approved voltage-gated calcium channel blocker for refractory chronic pain. However, due to the narrow therapeutic window and considerable side effects associated with systemic dosing, Prialt is only administered intrathecally. N-triazole oxindole (TROX-1) is a novel use-dependent and activation state-selective small-molecule inhibitor of Ca(v)2.1, 2.2 and 2.3 calcium channels designed to overcome the limitations of Prialt. We have examined the neurophysiological and behavioral effects of blocking calcium channels with TROX-1. In vitro, TROX-1, in contrast to state-independent antagonist Prialt, preferentially inhibits Ca(v)2.2 currents in rat dorsal root ganglia (DRG) neurons under depolarized conditions. In vivo electrophysiology was performed to record from deep dorsal horn lamina V/VI wide dynamic range neurons in non-sentient spinal nerve-ligated (SNL) and sham-operated rats. In SNL rats, spinal neurons exhibited reduced responses to innocuous and noxious punctate mechanical stimulation of the receptive field following subcutaneous administration of TROX-1, an effect that was absent in sham-operated animals. No effect was observed on neuronal responses evoked by dynamic brushing, heat or cold stimulation in SNL or sham rats. The wind-up response of spinal neurons following repeated electrical stimulation of the receptive field was also unaffected. Spinally applied TROX-1 dose dependently inhibited mechanically evoked neuronal responses in SNL but not sham-operated rats, consistent with behavioral observations. This study confirms the pathological state-dependent actions of TROX-1 through a likely spinal mechanism and reveals a modality selective change in calcium channel function following nerve injury. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Agonist activation of cytosolic Ca2+ in subfornical organ cells projecting to the supraoptic nucleus

NASA Technical Reports Server (NTRS)

Johnson, R. F.; Beltz, T. G.; Sharma, R. V.; Xu, Z.; Bhatty, R. A.; Johnson, A. K.

2001-01-01

The subfornical organ (SFO) is sensitive to both ANG II and ACh, and local application of these agents produces dipsogenic responses and vasopressin release. The present study examined the effects of cholinergic drugs, ANG II, and increased extracellular osmolarity on dissociated, cultured cells of the SFO that were retrogradely labeled from the supraoptic nucleus. The effects were measured as changes in cytosolic calcium in fura 2-loaded cells by using a calcium imaging system. Both ACh and carbachol increased intracellular ionic calcium concentration ([Ca2+]i). However, in contrast to the effects of muscarinic receptor agonists on SFO neurons, manipulation of the extracellular osmolality produced no effects, and application of ANG II produced only moderate effects on [Ca2+]i in a few retrogradely labeled cells. The cholinergic effects on [Ca2+]i could be blocked with the muscarinic receptor antagonist atropine and with the more selective muscarinic receptor antagonists pirenzepine and 4-diphenylacetoxy-N-methylpiperdine methiodide (4-DAMP). In addition, the calcium in the extracellular fluid was required for the cholinergic-induced increase in [Ca2+]i. These findings indicate that ACh acts to induce a functional cellular response in SFO neurons through action on a muscarinic receptor, probably of the M1 subtype and that the increase of [Ca2+]i, at least initially, requires the entry of extracellular Ca2+. Also, consistent with a functional role of M1 receptors in the SFO are the results of immunohistochemical preparations demonstrating M1 muscarinic receptor-like protein present within this forebrain circumventricular organ.

The Marine Guanidine Alkaloid Crambescidin 816 Induces Calcium Influx and Cytotoxicity in Primary Cultures of Cortical Neurons through Glutamate Receptors.

PubMed

Mendez, Aida G; Juncal, Andrea Boente; Silva, Siguara B L; Thomas, Olivier P; Martín Vázquez, Víctor; Alfonso, Amparo; Vieytes, Mercedes R; Vale, Carmen; Botana, Luís M

2017-07-19

Crambescidin 816 is a guanidine alkaloid produced by the sponge Crambe crambe with known antitumoral activity. While the information describing the effects of this alkaloid in central neurons is scarce, Cramb816 is known to block voltage dependent calcium channels being selective for L-type channels. Moreover, Cramb816 reduced neuronal viability through an unknown mechanism. Here, we aimed to describe the toxic activity of Cramb816 in cortical neurons. Since calcium influx is considered the main mechanism responsible for neuronal cell death, the effects of Cramb816 in the cytosolic calcium concentration of cortical neurons were studied. The alkaloid decreased neuronal viability and induced a dose-dependent increase in cytosolic calcium that was also related to the presence of calcium in the extracellular media. The increase in calcium influx was age dependent, being higher in younger neurons. Moreover, this effect was prevented by glutamate receptor antagonists, which did not fully block the cytotoxic effect of Cramb816 after 24 h of treatment but completely prevented Cramb816 cytotoxicity after 10 min exposure. Therefore, the findings presented herein provide new insights into the cytotoxic effect of Cramb816 in cortical neurons.

Calmodulin-mediated activation of Akt regulates survival of c-Myc-overexpressing mouse mammary carcinoma cells.

PubMed

Deb, Tushar B; Coticchia, Christine M; Dickson, Robert B

2004-09-10

c-Myc-overexpressing mammary epithelial cells are proapoptotic; their survival is strongly promoted by epidermal growth factor (EGF). We now demonstrate that EGF-induced Akt activation and survival in transgenic mouse mammary tumor virus-c-Myc mouse mammary carcinoma cells are both calcium/calmodulin-dependent. Akt activation is abolished by the phospholipase C-gamma inhibitor U-73122, by the intracellular calcium chelator BAPTA-AM, and by the specific calmodulin antagonist W-7. These results implicate calcium/calmodulin in the activation of Akt in these cells. In addition, Akt activation by serum and insulin is also inhibited by W-7. EGF-induced and calcium/calmodulin-mediated Akt activation occurs in both tumorigenic and non-tumorigenic mouse and human mammary epithelial cells, independent of their overexpression of c-Myc. These results imply that calcium/calmodulin may be a common regulator of Akt activation, irrespective of upstream receptor activator, mammalian species, and transformation status in mammary epithelial cells. However, only c-Myc-overexpressing mouse mammary carcinoma cells (but not normal mouse mammary epithelial cells) undergo apoptosis in the presence of the calmodulin antagonist W-7, indicating the vital selective role of calmodulin for survival of these cells. Calcium/calmodulin-regulated Akt activation is mediated directly by neither calmodulin kinases nor phosphatidylinositol 3-kinase (PI-3 kinase). Pharmacological inhibitors of calmodulin kinase kinase and calmodulin kinases II and III do not inhibit EGF-induced Akt activation, and calmodulin antagonist W-7 does not inhibit phosphotyrosine-associated PI-3 kinase activation. Akt is, however, co-immunoprecipitated with calmodulin in an EGF-dependent manner, which is inhibited by calmodulin antagonist W-7. We conclude that calmodulin may serve a vital regulatory function to direct the localization of Akt to the plasma membrane for its activation by PI-3 kinase.

Role of antispasmodics in the treatment of irritable bowel syndrome

PubMed Central

Annaházi, Anita; Róka, Richárd; Rosztóczy, András; Wittmann, Tibor

2014-01-01

Irritable bowel syndrome (IBS) is a long-lasting, relapsing disorder characterized by abdominal pain/discomfort and altered bowel habits. Intestinal motility impairment and visceral hypersensitivity are the key factors among its multifactorial pathogenesis, both of which require effective treatment. Voltage-gated calcium channels mediate smooth muscle contraction and endocrine secretion and play important roles in neuronal transmission. Antispasmodics are a group of drugs that have been used in the treatment of IBS for decades. Alverine citrate, a spasmolytic, decreases the sensitivity of smooth muscle contractile proteins to calcium, and it is a selective 5-HT1A receptor antagonist. Alverine, in combination with simethicone, has been demonstrated to effectively reduce abdominal pain and discomfort in a large placebo-controlled trial. Mebeverine is a musculotropic agent that potently blocks intestinal peristalsis. Non-placebo-controlled trials have shown positive effects of mebeverine in IBS regarding symptom control; nevertheless, in recent placebo-controlled studies, mebeverine did not exhibit superiority over placebo. Otilonium bromide is poorly absorbed from the GI tract, where it acts locally as an L-type calcium channel blocker, an antimuscarinic and a tachykinin NK2 receptor antagonist. Otilonium has effectively reduced pain and improved defecation alterations in placebo-controlled trials in IBS patients. Pinaverium bromide is also an L-type calcium channel blocker that acts locally in the GI tract. Pinaverium improves motility disorders and consequently reduces stool problems in IBS patients. Phloroglucinol and trimethylphloroglucinol are non-specific antispasmodics that reduced pain in IBS patients in a placebo-controlled trial. Antispasmodics have excellent safety profiles. T-type calcium channel blockers can abolish visceral hypersensitivity in animal models, which makes them potential candidates for the development of novel therapeutic agents in the treatment of IBS. PMID:24876726

Selective dopamine receptor 4 activation mediates the hippocampal neuronal calcium response via IP3 and ryanodine receptors.

PubMed

Wang, Ya-Li; Wang, Jian-Gang; Guo, Fang-Li; Gao, Xia-Huan; Zhao, Dan-Dan; Zhang, Lin; Wang, Jian-Zhi; Lu, Cheng-Biao

2017-09-01

Intracellular calcium is a key factor in most cellular processes, including cell growth, differentiation, proliferation and neurotransmitter release. Dopamine (DA) mediates synaptic transmission by regulating the intracellular calcium content. It is not clear, however, which specific subunit of the DA receptor contributes to DA modulation of intracellular calcium content changes. Through the traditional technique of Fura-2 calcium imaging, this study demonstrated that the DA can induce transient calcium in cultured hippocampal neurons and that this response can be mimicked by a selective dopamine receptor 4 (DR4) agonist PD168077 (PD). PD-induced calcium transience can be blocked by a calcium chelator, such as BAPTA-AM, or by pre-treatment of neurons with thapsigargin, a IP 3 receptor antagonist, or a micromolar concentration of ryanodine, a ryanodine receptor (RyR) antagonist. However PD-induced calcium transience cannot be blocked by pre-treatment of neurons with a free-calcium medium or a cocktail of NMDA receptor, L-type calcium channel and alpha7 nicotinic acetylcholine receptor blockers. These results indicate that the calcium response induced by DR4 activation is mainly through activation of IP 3 receptor in internal stores, which is likely to contribute to the DA modulation of synaptic transmission and cognitive function. Copyright © 2017. Published by Elsevier B.V.

Endothelin induces two types of contractions of rat uterus: phasic contractions by way of voltage-dependent calcium channels and developing contractions through a second type of calcium channels

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kozuka, M.; Ito, T.; Hirose, S.

1989-02-28

Effects of endothelin on nonvascular smooth muscle have been examined using rat uterine horns and two modes of endothelin action have been revealed. Endothelin (0.3 nM) caused rhythmic contractions of isolated uterus in the presence of extracellular calcium. The rhythmic contractions were completely inhibited by calcium channel antagonists. These characteristics of endothelin-induced contractions were very similar to those induced by oxytocin. Binding assays using /sup 125/I-endothelin showed that endothelin and the calcium channel blockers did not compete for the binding sites. However, endothelin was unique in that it caused, in addition to rhythmic contractions, a slowly developing monophasic contraction thatmore » was insensitive to calcium channel blockers. This developing contraction became dominant at higher concentrations of endothelin and was also calcium dependent.« less

Lead-induced ER calcium release and inhibitory effects of methionine choline in cultured rat hippocampal neurons.

PubMed

Fan, Guangqin; Zhou, Fankun; Feng, Chang; Wu, Fengyun; Ye, Weiwei; Wang, Chunhong; Lin, Fen; Yan, Ji; Li, Yanshu; Chen, Ying; Bi, Yongyi

2013-02-01

Lead, a ubiquitous neurotoxicant, can result in learning and memory dysfunction. Long term potentiation in the hippocampus, a potential neural substrate for learning and memory, is thought to be linked to calcium-triggered intracellular events. In this study, laser scanning confocal microscopy was used to examine the effects of Pb(2+) on intracellular and endoplasmic reticulum free calcium concentration ([Ca(2+)](i) and [Ca(2+)](ER)) in cultured neonatal rat hippocampal neurons and their possible antagonism by methionine choline; understanding these effects would help explain the lead-induced cognitive and learning dysfunction and explore efficient safety and relief strategies. The results showed that Pb(2+) increased [Ca(2+)](i) and decreased [Ca(2+)](ER) linearly in a time- and concentration-dependant manner, and Pb(2+) addition after the applying of a ryanodine receptor (RyR) antagonist and an inositol-1,4,5-triphosphate receptor (IP(3)R) antagonist did not increase [Ca(2+)](i). The addition of 10, 20, or 40 mmol/L methionine choline simultaneously with addition of 10 μmol/L Pb(2+) decreased [Ca(2+)](i) in Ca(2+)-free culture medium by 39.0%, 66.0%, and 61.6%, respectively, in a concentration-dependant manner in a certain dose range. Our results suggest that Pb(2+) induces ER calcium release to increase the resting [Ca(2+)](i); and methionine choline inhibit this increase in [Ca(2+)](i). Copyright © 2012 Elsevier Ltd. All rights reserved.

[Control of generalized chronic periodontitis combined with calcium-antagonist-related gingival overgrowth by a complex periodontal-endodontic-prosthodontic treatment. Case report].

PubMed

Szánto, Erika; Gera, István

2011-12-01

To day a relatively high percentage of elderly population of the industrialized world suffers with different cardiovascular diseases and are on permanent antihypertensive therapy. One of the most frequently used drugs is the calcium channel blockers prescribed against high blood pressure. The most common oral side effect of these drugs is the gingival enlargement that might develop even on otherwise healthy gingiva. The incidence of chronic periodontitis in this age group is also high and the Ca antagonist medication in those individuals might substantially modify the clinical course of periodontal inflammation leading to gingival enlargement and hypertrophic pocket wall. The case presented here is a 52 years old hypertonic woman with a long history of Ca-antagonist therapy and generalized chronic periodontitis combined with gingival hyperplasia. After the change of medication the 1,5 years comprehensive periodontal endodontic and prosthodontic therapy restored patient's periodontal health and provided complex dental rehabilitation. Nevertheless, only regular periodontal supportive therapy could ensure predictable outcome and guarantee long lasting periodontal health.

Off Label Antiviral Therapeutics for Henipaviruses: New Light Through Old Windows

PubMed Central

Aljofan, Mohamad; Lo, Michael K.; Rota, Paul A.; Michalski, Wojtek P.; Mungall, Bruce A.

2010-01-01

Hendra and Nipah viruses are recently emerged zoonotic paramyxoviruses for which there is no vaccine or protective therapy available. While a number of experimental therapeutics and vaccines have recently been reported, all of these will require lengthy approval processes, limiting their usefulness in the short term. To address the urgent need for henipavirus therapeutics, a number of currently licensed pharmaceuticals have been evaluated for off label efficacy against henipavirus replication in vitro. Initially it was observed that compounds which released intracellular calcium stores induced a potent inhibition of henipaviruses replication, prompting the evaluation of known drugs with a similar effect on calcium mobilisation. Of the eight compounds randomly selected based on existing literature, seven inhibited virus replication in the micromolar range while the remaining compound also inhibited virus replication but only at millimolar concentrations. Pretreatment experiments with various calcium chelators, channel antagonists or endoplasmic reticulum release inhibitors supported a calcium mediated mechanism of action for five of these compounds. The mechanism of antiviral action for the remaining three compounds is currently unknown. Additionally, a number of other modulators of calcium flux, including calcium channel and calmodulin antagonists also exhibited potent antiviral activity in vitro providing a broad range of potential therapeutic options for the treatment of henipavirus infections. Importantly, as many of these compounds are currently licensed drugs, regulatory approval should be a much more streamlined process, with the caveat that appropriate in vivo efficacy can be demonstrated in animal models. PMID:20668647

Effects of single and repeated doses of the calcium antagonist felodipine on blood pressure, renal function, electrolytes and water balance, and renin-angiotensin-aldosterone system in hypertensive patients.

PubMed

Leonetti, G; Gradnik, R; Terzoli, L; Fruscio, M; Rupoli, L; Cuspidi, C; Sampieri, L; Zanchetti, A

1986-01-01

Doses of 10 mg b.i.d. of the new dihydropyridine calcium antagonist, felodipine, were tested for seven consecutive days in 11 hospitalized hypertensive patients. A significant reduction of both systolic and diastolic blood pressures, with patients in both the supine and upright positions, occurred immediately after the first dose and was maintained (daily average 15%) throughout the following days. An increase in heart rate was observed after the first dose (15 and 23 beats/min, in supine and upright postures), and subsequently declined to average values of 8 and 14 beats/min on the seventh day. There was a marked natriuretic response during the first and second day, during which an average negative sodium balance of 95 mmol developed; on the following days sodium output was not significantly different from control, but a negative balance averaging 135 mmol was still present on the seventh day of felodipine administration. A moderate negative potassium balance also progressively developed and reached -48 mmol on the seventh day. Glomerular filtration rate was unchanged, but renal plasma flow increased significantly during administration of felodipine. Plasma renin activity and plasma aldosterone were also increased very moderately by felodipine. Compared with previous observations by our group with higher doses of felodipine (12.5, 25, and 50 mg t.i.d.), 10 mg b.i.d. of this new calcium antagonist appear to exert a marked and prolonged blood pressure reduction, accompanied by a definite natriuretic instead of an antinatriuretic effect.

Effects of two medicinal plants Psidium guajava L. (Myrtaceae) and Diospyros mespiliformis L. (Ebenaceae) leaf extracts on rat skeletal muscle cells in primary culture.

PubMed

Belemtougri, R G; Constantin, B; Cognard, C; Raymond, G; Sawadogo, L

2006-01-01

Crude decoction, aqueous and ethanolic extracts of two medicinal plants (Psidium guajava and Diospyros mespiliformis), widely used in the central plateau of Burkina Faso to treat many diseases were evaluated for their antagonistic effects on caffeine induced calcium release from sarcoplasmic reticulum of rat skeletal muscle cells. These different extracts showed a decrease of caffeine induced calcium release in a dose dependent manner. Comparison of the results showed that Psidium guajava leaf extracts are more active than extracts of Diospyros mespiliformis and that crude decoctions show better inhibitory activity. The observed results could explain their use as antihypertensive and antidiarrhoeal agents in traditional medicine, by inhibiting intracellular calcium release.

Effects of two medicinal plants Psidium guajava L. (Myrtaceae) and Diospyros mespiliformis L. (Ebenaceae) leaf extracts on rat skeletal muscle cells in primary culture

PubMed Central

Belemtougri, R.G.; Constantin, B.; Cognard, C.; Raymond, G.; Sawadogo, L.

2006-01-01

Crude decoction, aqueous and ethanolic extracts of two medicinal plants (Psidium guajava and Diospyros mespiliformis), widely used in the central plateau of Burkina Faso to treat many diseases were evaluated for their antagonistic effects on caffeine induced calcium release from sarcoplasmic reticulum of rat skeletal muscle cells. These different extracts showed a decrease of caffeine induced calcium release in a dose dependent manner. Comparison of the results showed that Psidium guajava leaf extracts are more active than extracts of Diospyros mespiliformis and that crude decoctions show better inhibitory activity. The observed results could explaine their use as antihypertensive and antidiarrhoeal agents in traditional medicine, by inhibiting intracellular calcium release. PMID:16365927

Nuclear Membranes ETB Receptors Mediate ET-1-induced Increase of Nuclear Calcium in Human Left Ventricular Endocardial Endothelial Cells.

PubMed

Jules, Farah; Avedanian, Levon; Al-Khoury, Johny; Keita, Ramatoulaye; Normand, Alexandre; Bkaily, Ghassan; Jacques, Danielle

2015-07-01

In fetal human left ventricular endocardial endothelial cells (EECLs), both plasma membrane (PM) ET(A)R and ET(B)R were reported to mediate ET-1-induced increase of intracellular calcium [Ca](i); however, this effect was mediated by ET(A)R in right EECs (EECRs). In this study, we verified whether, as for the PM, nuclear membranes (NMs) ET-1 receptors activation in EECLs and EECRs induce an increase of nuclear calcium ([Ca](n)) and if this effect is mediated through the same receptor type as in PM. Using a plasmalemma-perforated technique and 3D confocal microscopy, our results showed that, as in PM intact cells, superfusion of nuclei of both cell types with cytosolic ET-1 induced a concentration-dependent sustained increase of [Ca](n). In EECRs, the ET(A)R antagonist prevented the effect of ET-1 on [Ca](n) without affecting EECLs. However, in both cell types, the effect of cytosolic ET-1 on [Ca](n) was prevented by the ETBR antagonist. In conclusion, both NMs' ET(A)R and ET(B)R mediated the effect of cytosolic ET-1 on [Ca](n) in EECRs. In contrast, only NMs' ET(B)R activation mediated the effect of cytosolic ET-1 in EECLs. Hence, the type of NMs' receptors mediating the effect of ET-1 on [Ca](n) are different from those of PM mediating the increase in [Ca](i).

The solution structure of omega-Aga-IVB, a P-type calcium channel antagonist from venom of the funnel web spider, Agelenopsis aperta.

PubMed

Reily, M D; Thanabal, V; Adams, M E

1995-02-01

The 48 amino acid peptides omega-Aga-IVA and omega-Aga-IVB are the first agents known to specifically block P-type calcium channels in mammalian brain, thus complementing the existing suite of pharmacological tools used for characterizing calcium channels. These peptides provide a new set of probes for studies aimed at elucidating the structural basis underlying the subtype specificity of calcium channel antagonists. We used 288 NMR-derived constraints in a protocol combining distance geometry and molecular dynamics employing the program DGII, followed by energy minimization with Discover to derive the three-dimensional structure of omega-Aga-IVB. The toxin consists of a well-defined core region, comprising seven solvent-shielded residues and a well-defined triple-stranded beta-sheet. Four loop regions have average backbone rms deviations between 0.38 and 1.31 A, two of which are well-defined type-II beta-turns. Other structural features include disordered C- and N-termini and several conserved basic amino acids that are clustered on one face of the molecule. The reported structure suggests a possible surface for interaction with the channel. This surface contains amino acids that are identical to those of another known P-type calcium channel antagonist, omega-Aga-IVA, and is rich in basic residues that may have a role in binding to the anionic sites in the extracellular regions of the calcium channel.

Cilnidipine, an L/N-type calcium channel blocker prevents acquisition and expression of ethanol-induced locomotor sensitization in mice.

PubMed

Bhutada, Pravinkumar; Mundhada, Yogita; Patil, Jayshree; Rahigude, Anand; Zambare, Krushna; Deshmukh, Prashant; Tanwar, Dhanshree; Jain, Kishor

2012-04-11

Several evidences indicated the involvement of L- and N-type calcium channels in behavioral effects of drugs of abuse, including ethanol. Calcium channels are implicated in ethanol-induced behaviors and neurochemical responses. Calcium channel antagonists block the psychostimulants induced behavioral sensitization. Recently, it is demonstrated that L-, N- and T-type calcium channel blockers attenuate the acute locomotor stimulant effects of ethanol. However, no evidence indicated the role of calcium channels in ethanol-induced psychomotor sensitization. Therefore, present study evaluated the influence of cilnidipine, an L/N-type calcium channel blocker on acquisition and expression of ethanol-induced locomotor sensitization. The results revealed that cilnidipine (0.1 and 1.0μg/mouse, i.c.v.) attenuates the expression of sensitization to locomotor stimulant effect of ethanol (2.0g/kg, i.p.), whereas pre- treatment of cilnidipine (0.1 and 1.0μg/mouse, i.c.v.) during development of sensitization blocks acquisition and attenuates expression of sensitization to locomotor stimulant effect of ethanol. Cilnidipine per se did not influence locomotor activity in tested doses. Further, cilnidipine had no influence on effect of ethanol on rotarod performance. These results support the hypothesis that neuroadaptive changes in calcium channels participate in the acquisition and the expression of ethanol-induced locomotor sensitization. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Inflammation alters AMPA-stimulated calcium responses in dorsal striatal D2 but not D1 spiny projection neurons.

PubMed

Winland, Carissa D; Welsh, Nora; Sepulveda-Rodriguez, Alberto; Vicini, Stefano; Maguire-Zeiss, Kathleen A

2017-11-01

Neuroinflammation precedes neuronal loss in striatal neurodegenerative diseases and can be exacerbated by the release of proinflammatory molecules by microglia. These molecules can affect trafficking of AMPARs. The preferential trafficking of calcium-permeable versus impermeable AMPARs can result in disruptions of [Ca 2+ ] i and alter cellular functions. In striatal neurodegenerative diseases, changes in [Ca 2+ ] i and L-type voltage-gated calcium channels (VGCCs) have been reported. Therefore, this study sought to determine whether a proinflammatory environment alters AMPA-stimulated [Ca 2+ ] i through calcium-permeable AMPARs and/or L-type VGCCs in dopamine-2- and dopamine-1-expressing striatal spiny projection neurons (D2 and D1 SPNs) in the dorsal striatum. Mice expressing the calcium indicator protein, GCaMP in D2 or D1 SPNs, were utilized for calcium imaging. Microglial activation was assessed by morphology analyses. To induce inflammation, acute mouse striatal slices were incubated with lipopolysaccharide (LPS). Here we report that LPS treatment potentiated AMPA responses only in D2 SPNs. When a nonspecific VGCC blocker was included, we observed a decrease of AMPA-stimulated calcium fluorescence in D2 but not D1 SPNs. The remaining agonist-induced [Ca 2+ ] i was mediated by calcium-permeable AMPARs because the responses were completely blocked by a selective calcium-permeable AMPAR antagonist. We used isradipine, the highly selective L-type VGCC antagonist to determine the role of L-type VGCCs in SPNs treated with LPS. Isradipine decreased AMPA-stimulated responses selectively in D2 SPNs after LPS treatment. Our findings suggest that dorsal striatal D2 SPNs are specifically targeted in proinflammatory conditions and that L-type VGCCs and calcium-permeable AMPARs are important mediators of this effect. © 2017 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gaba, D.M.; Metz, S.; Maze, M.

Transthoracic electric countershock can cause necrotic myocardial lesions in humans as well as experimental animals. The authors investigated the effect on postcountershock myocardial damage of pretreatment with prazosin, an alpha-1 antagonist; L-metoprolol, a beta-1 antagonist, and verapamil, a calcium channel-blocking agent. Twenty dogs were anesthetized with halothane and given two transthoracic countershocks of 295 delivered joules each after drug or vehicle treatment. Myocardial injury was quantitated 24 h following countershock by measuring the uptake of technetium-99m pyrophosphate in the myocardium. Elevated technetium-99m pyrophosphate uptake occurred in visible lesions in most dogs regardless of drug treatment. For each of four parametersmore » of myocardial damage there was no statistically significant difference between control animals and those treated with prazosin, metoprolol, or verapamil. These data suggest that adrenergic or calcium channel-mediated mechanisms are not involved in the pathogenesis of postcountershock myocardial damage.« less

25-Hydroxycholecalciferol as an antagonist of adverse corticosteroid effects on phosphate and calcium metabolism in man.

PubMed

Nuti, R; Vattimo, A; Turchetti, V; Righi, G

1984-10-01

The present study was performed in 30 patients who needed steroid therapy: courses of triamcinolone or DTM 8-15 given orally lasted 30 days. In 15 of these patients glucoactive corticosteroids were administered in combination with 5 micrograms/day of 25OH-vitamin D3 (25OHD3). 47Calcium oral test and 99mTc-MDP kinetics, as an index of bone turnover, were performed at the beginning of the therapy and after 30 days. At the end of treatment a significant improvement of intestinal radiocalcium transport together with a decrease in bone turnover in the group of patients treated with 25OHD3 was observed. As it concerns plasma calcium level, inorganic phosphate, the urinary excretion of calcium, phosphate and hydroxyproline no significant difference between the two groups examined were noticed. These results indicate that the adverse effects of glucoactive corticosteroids on intestinal calcium transport and bone turnover may be counteracted by the combined administration of physiological doses of 25OHD3.

Omega-conotoxin- and nifedipine-insensitive voltage-operated calcium channels mediate K(+)-induced release of pro-thyrotropin-releasing hormone-connecting peptides Ps4 and Ps5 from perifused rat hypothalamic slices.

PubMed

Valentijn, K; Tranchand Bunel, D; Vaudry, H

1992-07-01

The rat thyrotropin-releasing hormone (TRH) precursor (prepro-TRH) contains five copies of the TRH progenitor sequence linked together by intervening sequences. Recently, we have shown that the connecting peptides prepro-TRH-(160-169) (Ps4) and prepro-TRH-(178-199) (Ps5) are released from rat hypothalamic neurones in response to elevated potassium concentrations, in a calcium-dependent manner. In the present study, the role of voltage-operated calcium channels in potassium-induced release of Ps4 and Ps5 was investigated, using a perifusion system for rat hypothalamic slices. The release of Ps4 and Ps5 stimulated by potassium (70 mM) was blocked by the inorganic ions Co2+ (2.6 mM) and Ni2+ (5 mM). In contrast, the stimulatory effect of KCl was insensitive to Cd2+ (100 microM). The dihydropyridine antagonist nifedipine (10 microM) had no effect on K(+)-evoked release of Ps4 and Ps5. Furthermore, the response to KCl was not affected by nifedipine (10 microM) in combination with diltiazem (1 microM), a benzothiazepine which increases the affinity of dihydropyridine antagonists for their receptor. The dihydropyridine agonist BAY K 8644, at concentrations as high as 1 mM, did not stimulate the basal secretion of Ps4 and Ps5. In addition, BAY K 8644 had no potentiating effect on K(+)-induced release of Ps4 and Ps5. The marine cone snail toxin omega-conotoxin, a blocker of both L- and N-type calcium channels had no effect on the release of Ps4 and Ps5 stimulated by potassium. Similarly, the omega-conopeptide SNX-111, a selective blocker of N-type calcium channels, did not inhibit the stimulatory effect of potassium. The release of Ps4 and Ps5 evoked by high K+ was insensitive to the non-selective calcium channel blocker verapamil (20 microM). Amiloride (1 microM), a putative blocker of T-type calcium channels, did not affect KCl-induced secretion of the two connecting peptides. Taken together, these results indicate that two connecting peptides derived from the pro-TRH, Ps4 and Ps5, are released by K(+)-induced depolarization through activation of voltage-sensitive calcium channels. The calcium channels appear to have a pharmacological profile different from that of L- and N-type channels. Although, their insensitivity to low Cd2+ concentrations and sensitivity to Ni2+ ions would support the involvement of T-type calcium channels, the lack of effect of amiloride suggests that they belong to a yet undefined class of calcium channels.

Impact of magnesium:calcium ratio on calcification of the aortic wall.

PubMed

Villa-Bellosta, Ricardo

2017-01-01

An inverse relationship between serum magnesium concentration and vascular calcification has been reported following observational clinical studies. Moreover, several studies have been suggesting a protective effect of magnesium on the vascular calcification. However, the exact mechanism remains elusive, and investigators have speculated among a myriad of potential actions. The effect of magnesium on calcification of the aortic wall is yet to be investigated. In the present study, the effects of magnesium and calcium on the metabolism of extracellular PPi, the main endogenous inhibitor of vascular calcification, were investigated in the rat aorta. Calcium and magnesium have antagonist effects on PPi hydrolysis in the aortic wall. Km and Ki values for PPi hydrolysis in rat aortic rings were 1.1 mmol/L magnesium and 32 μmol/L calcium, respectively, but ATP hydrolysis was not affected with calcium. Calcium deposition in the rat aortic wall dramatically increased when the magnesium concentration was increased (ratio of Mg:Ca = 1:1; 1.5 mmol/L calcium and 1.5 mmol/L magnesium) respect to low magnesium concentration (ratio Mg:Ca = 1:3, 1.5 mmol/L calcium and 0.75 mmol/L magnesium). Data from observational clinical studies showing that the serum magnesium concentration is inversely correlated with vascular calcification could be reinterpreted as a compensatory regulatory mechanism that reduces both PPi hydrolysis and vascular calcification. The impact of magnesium in vascular calcification in humans could be studied in association with calcium levels, for example, as the magnesium:calcium ratio.

Identification and characterization of a selective allosteric antagonist of human P2X4 receptor channels.

PubMed

Ase, Ariel R; Honson, Nicolette S; Zaghdane, Helmi; Pfeifer, Tom A; Séguéla, Philippe

2015-04-01

P2X4 is an ATP-gated nonselective cation channel highly permeable to calcium. There is increasing evidence that this homomeric purinoceptor, which is expressed in several neuronal and immune cell types, is involved in chronic pain and inflammation. The current paucity of unambiguous pharmacological tools available to interrogate or modulate P2X4 function led us to pursue the search for selective antagonists. In the high-throughput screen of a compound library, we identified the phenylurea BX430 (1-(2,6-dibromo-4-isopropyl-phenyl)-3-(3-pyridyl)urea, molecular weight = 413), with antagonist properties on human P2X4-mediated calcium uptake. Patch-clamp electrophysiology confirmed direct inhibition of P2X4 currents by extracellular BX430, with submicromolar potency (IC50 = 0.54 µM). BX430 is highly selective, having virtually no functional impact on all other P2X subtypes, namely, P2X1-P2X3, P2X5, and P2X7, at 10-100 times its IC50. Unexpected species differences were noticed, as BX430 is a potent antagonist of zebrafish P2X4 but has no effect on rat and mouse P2X4 orthologs. The concentration-response curve for ATP on human P2X4 in the presence of BX430 shows an insurmountable blockade, indicating a noncompetitive allosteric mechanism of action. Using a fluorescent dye uptake assay, we observed that BX430 also effectively suppresses ATP-evoked and ivermectin-potentiated membrane permeabilization induced by P2X4 pore dilation. Finally, in single-cell calcium imaging, we validated its selective inhibitory effects on native P2X4 channels at the surface of human THP-1 cells that were differentiated into macrophages. In summary, this ligand provides a novel molecular probe to assess the specific role of P2X4 in inflammatory and neuropathic conditions, where ATP signaling has been shown to be dysfunctional. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Bradykinin induced a positive chronotropic effect via stimulation of T- and L-type calcium currents in heart cells.

PubMed

El-Bizri, Nesrine; Bkaily, Ghassan; Wang, Shimin; Jacques, Danielle; Regoli, Domenico; D'Orléans-Juste, Pedro; Sukarieh, Rami

2003-03-01

Using Fluo-3 calcium dye confocal microscopy and spontaneously contracting embryonic chick heart cells, bradykinin (10(-10) M) was found to induce positive chronotropic effects by increasing the frequency of the transient increase of cytosolic and nuclear free Ca2+. Pretreatment of the cells with either B1 or B2 receptor antagonists (R126 and R817, respectively) completely prevented bradykinin (BK) induced positive chronotropic effects on spontaneously contracting single heart cells. Using the whole-cell voltage clamp technique and ionic substitution to separate the different ionic current species, our results showed that BK (10(-6) M) had no effect on fast Na+ inward current and delayed outward potassium current. However, both L- and T-type Ca2+ currents were found to be increased by BK in a dose-dependent manner (10(-10)-10(-7) M). The effects of BK on T- and L-type Ca2+ currents were partially blocked by the B1 receptor antagonist [Leu8]des-Arg9-BK (R592) (10(-7) M) and completely reversed by the B2 receptor antagonist D-Arg[Hyp3,D-Phe7,Leu8]BK (R-588) (10(-7) M) or pretreatment with pertussis toxin (PTX). These results demonstrate that BK induced a positive chronotropic effect via stimulation of T- and L-type Ca2+ currents in heart cells mainly via stimulation of B2 receptor coupled to PTX-sensitive G-proteins. The increase of both types of Ca2+ current by BK in heart cells may explain the positive inotropic and chronotropic effects of this hormone.

Neurotransmitter Amines in Hemorrhagic Shock.

DTIC Science & Technology

1984-03-02

and synthesis for future studies. Crit Care Med 11:202-207, 1983. 7. Gelmer HJ: Effect of ninodipine (Bay e 9736) on post-ischaemic- cerebro - vascular...the calcium antagonist, nimodipine on cerebral blood flow and metabolism in the primate . JCereb Blood Flow and Metabol 1:349-356, 1981. 9. Edvinsson L

Chick cerebellar Purkinje cells express omega-conotoxin GVIA-sensitive rather than funnel-web spider toxin-sensitive calcium channels.

PubMed

Angulo, M C; Parra, P; Dieudonné, S

1998-03-01

Voltage-gated calcium channels form a complex family of distinct molecular entities which participate in multiple neuronal functions. In cerebellar Purkinje cells these channels contribute to the characteristic electrophysiological pattern of complex spikes, first described in birds and later in mammals. A specific calcium channel, the P-type channel, has been shown to mediate the majority of the voltage-gated calcium flux in mammalian Purkinje cells. P-type channels play an essential role in synaptic transmission of mammalian cerebellum. It is unclear whether the P-type calcium channel is present in birds. Studies in chick synaptosomal preparations show that the pharmacological profile of calcium channels is complex and suggest a minimal expression of the P-type channel in avian central nervous system. In the present work, we studied voltage-gated calcium channels in dissociated chick cerebellar Purkinje cells to examine the presence of different calcium channel types. Purkinje cells were used because, in mammals, they express predominantly P-type channels and because the morphology of these cells is thought to be phylogenetically conserved. We found that omega-conotoxin GVIA (omega-CgTx GVIA), a specific antagonist of N-type calcium channel, rather than the synthetic funnel-web spider toxin (sFTX), a P-type channel antagonist, blocks the majority of the barium current flowing through calcium channels in chick Purkinje neurons.

The effects of 3,4-methylenedioxymethamphetamine (MDMA) on nicotinic receptors: Intracellular calcium increase, calpain/caspase 3 activation, and functional upregulation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garcia-Rates, Sara; Camarasa, Jordi; Sanchez-Garcia, Ana I.

2010-05-01

Previous work by our group demonstrated that homomeric alpha7 nicotinic acetylcholine receptors (nAChR) play a role in the neurotoxicity induced by 3,4-methylenedioxymethamphetamine (MDMA), as well as the binding affinity of this drug to these receptors. Here we studied the effect of MDMA on the activation of nAChR subtypes, the consequent calcium mobilization, and calpain/caspase 3 activation because prolonged Ca{sup 2+} increase could contribute to cytotoxicity. As techniques, we used fluorimetry in Fluo-4-loaded PC12 cells and electrophysiology in Xenopus oocytes. MDMA produced a rapid and sustained increase in calcium without reaching the maximum effect induced by ACh. It also concentration-dependently inhibitedmore » the response induced by ACh, nicotine, and the specific alpha7 agonist PNU 282987 with IC{sub 50} values in the low micromolar range. Similarly, MDMA induced inward currents in Xenopus oocytes transfected with human alpha7 but not with alpha4beta2 nAChR and inhibited ACh-induced currents in both receptors in a concentration-dependent manner. The calcium response was inhibited by methyllycaconitine (MLA) and alpha-bungarotoxin but not by dihydro-beta-erythroidine. These results therefore indicate that MDMA acts as a partial agonist on alpha7 nAChRs and as an antagonist on the heteromeric subtypes. Subsequently, calcium-induced Ca{sup 2+} release from the endoplasmic reticulum and entry through voltage-operated calcium channels are also implicated as proved using specific antagonists. In addition, treatment with MDMA for 24 h significantly increased basal Ca{sup 2+} levels and induced an increase in alpha-spectrin breakdown products, which indicates that calpain and caspase 3 were activated. These effects were inhibited by pretreatment with MLA. Moreover, pretreatment with MDMA induced functional upregulation of calcium responses to specific agonists of both heteromeric and alpha7 nAChR. Sustained calcium entry and calpain activation could favor the activation of Ca{sup 2+}-dependent enzymes such as protein kinase C and nitric oxide synthase, which are involved in the generation of ROS and the blockade of the dopamine transporter. This, together with caspase 3 activation, must play a role in MDMA-induced cytotoxicity.« less

Pharmacologic study of calcium influx pathways in rabbit aortic smooth muscle

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lukeman, D.S.

1987-01-01

Functional characteristics and pharmacologic domains of receptor-operated and potential-sensitive calcium (Ca/sup 2 +/) channels (ROCs and PSCs, respectively) were derived via measurements of /sup 45/Ca/sup 2 +/ influx (M/sup Ca/) during activation by the neurotransmitters norepinephrine (NE), histamine (HS), and serotonin (5-HT) and by elevated extracellular potassium (K/sup +/) in the individual or combined presence of organic Ca/sup 2 +/ channel antagonists (CAts), calmodulin antagonists (Calm-ants), lanthanum (La/sup 3 +/), and agents that increase intracellular levels of cyclic AMP.

Induction of cell expansion of goldfish melanocytoma cells (GMM-1) by epinephrine and dexamethasone requires external calcium.

PubMed

Shih, Y L; Lo, S J

1993-05-01

Treatment of GMM-1 (a goldfish melanocytoma cell line) cells with epinephrine induced a rapid cell expansion (flattening of cells, extension and broadening of cellular processes) similar to the effect of dexamethasone reported previously (Shih et al., 1990). Studies on the possible involvement of secondary messengers in cell expansion indicated that (i) both 8-bromo-CAMP and forskolin caused cell shrinking (the opposite of cell expansion); (ii) TPA also caused cell shrinking; (iii) phospholipid derivatives, such as 1,2-dioctanoyl-sn-glycerol, lysophosphatidic acid, and arachidonic acid caused cell expansion; and (iv) EGTA (calcium chelator) and nifedipine (calcium channel blocker) inhibited the effect of epinephrine. Together with the previous findings, these observations indicate that epinephrine and dexamethasone may share a common pathway in triggering an external calcium influx to cause cell expansion. The results of the effects of epinephrine agonists and antagonists, together with those of other workers, also show that there are multiple isoforms of adrenoceptor in the goldfish.

Benzodiazepine-induced hippocampal CA1 neuron alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic acid (AMPA) receptor plasticity linked to severity of withdrawal anxiety: differential role of voltage-gated calcium channels and N-methyl-D-aspartic acid receptors.

PubMed

Xiang, Kun; Tietz, Elizabeth I

2007-09-01

Withdrawal from 1-week oral administration of the benzodiazepine, flurazepam (FZP) is associated with increased alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic acid (AMPA) receptor (AMPAR) miniature excitatory postsynaptic currents (mEPSCs) but reduction of N-methyl-D-aspartic acid (NMDA) receptor (NMDAR)-evoked (e)EPSCs in hippocampal CA1 neurons. A positive correlation was observed between increased AMPAR-mediated mEPSC amplitude and anxiety-like behavior in 1-day FZP-withdrawn rats. These effects were disrupted by systemic AMPAR antagonist administration (GYKI-52466, 0.5 mg/kg, intraperitoneal) at withdrawal onset, strengthening the hypothesis that CA1 neuron AMPAR-mediated hyperexcitability is a central component of a functional anatomic circuit associated with the expression of withdrawal anxiety. Abolition of AMPAR current upregulation in 2-day FZP withdrawn rats by GYKI-52466 injection also reversed the reduction in NMDAR-mediated eEPSC amplitude in CA1 neurons from the same rats, suggesting that downregulation of NMDAR function may serve a protective, negative-feedback role to prevent AMPAR-mediated neuronal overexcitation. NMDAR antagonist administration (MK-801, 0.25 mg/kg intraperitoneally) had no effect on modifying increased glutamatergic strength or on withdrawal anxiety, whereas injection of an L-type voltage-gated calcium channel antagonist, nimodipine (10 mg/kg, intraperitoneally) averted AMPAR current enhancement and anxiety-like behavior, suggesting that these manifestations may be initiated by a voltage-gated calcium channel-dependent signal transduction pathway. An evidence-based model of likely cellular mechanisms in the hippocampus contributing to benzodiazepine withdrawal anxiety was proposed implicating regulation of multiple CA1 neuron ion channels.

Nucleotide-mediated relaxation in guinea-pig aorta: selective inhibition by MRS2179

PubMed Central

Kaiser, Robert A; Buxton, Iain L O

2002-01-01

The vasodilatory effects of nucleotides in the guinea-pig thoracic aorta were examined to determine the relationship between molecular expression and function of P2Y receptors. In aortic rings precontracted with norepinephrine, vasodilatory responses to purine nucleotides exhibited a rank-order of potency of 2-methylthio-ATP>ADP>ATP. Responses to UTP, but not UDP suggested a functional role for P2Y4 but not P2Y6 receptors. Aortic endothelial cells express at least four P2Y receptors; P2Y1, P2Y2, P2Y4 and P2Y6. In primary culture, these cells exhibit desensitizing transient calcium responses characteristic of P2Y1, P2Y2 and P2Y4, but not P2Y6 receptors. UDP had no effect on endothelial cell calcium. The pyrimidinergic receptor agonist UTP is capable of eliciting robust vasodilation in aortic rings and causing calcium responses in cultured guineapig aortic endothelial cells. These responses are equivalent to the maximum responses observed to ATP and ADP. Measurement of intracellular calcium release in response to ATP and 2-methylthio-ATP were similar, however only the 2-methylthio-ATP response was sensitive to the P2Y1 antagonist N6-methyl-2′-deoxyadenosine-3′,5′-bisphosphate (MRS2179). In aortic rings, vasodilatory responses to 2-methylthio-ATP, ATP and ADP were all blocked by pre-incubation of tissues with MRS2179. MRS2179 pretreatment had no effect of the ability of UTP to cause relaxation of norepinephrine responses in aortic rings or the ability of UTP to cause calcium release in aortic endothelial cells. We demonstrate robust effects of purine and pyrimidine nucleotides in guineapig aorta and provide functional and biochemical evidence that MRS2179 is a selective P2Y1 antagonist. PMID:11815389

Modulation of chloroplast movement in the green alga Mougeotia by the Ca2+ ionophore A23187 and by calmodulin antagonists.

PubMed Central

Serlin, B S; Roux, S J

1984-01-01

The Ca2+ ionophore A23187 can induce chloroplast rotation within a single nonirradiated Mougeotia cell. The induced turning was dependent on the position of ionophore application and Ca2+ in the external medium. The role of calmodulin in mediating light-induced chloroplast rotation in the alga Mougeotia was investigated by using the paired calmodulin-antagonist drugs W5-W7 and W12-W13. In each pair, the antagonist with the greater affinity for calmodulin had the greater inhibitor effect on the phytochrome-controlled light response. These results support the hypothesis that calcium functions as a chemical messenger to couple the stimulus of phytochrome photoactivation with physiological responses in plants. Images PMID:11536594

Antagonist effects of calcium on borosilicate glass alteration

NASA Astrophysics Data System (ADS)

Mercado-Depierre, S.; Angeli, F.; Frizon, F.; Gin, S.

2013-10-01

Numerous studies have been conducted on glass and cement durability in contact with water, but very little work to date has focused directly on interactions between the two materials. These interactions are mostly controlled by silicon-calcium reactivity. However, the physical and chemical processes involved remain insufficiently understood to predict the evolution of coupled glass-cement systems used in several industrial applications. Results are reported from borosilicate glass alteration in calcium-rich solutions. Our data show that four distinct behaviors can be expected according to the relative importance of three key parameters: the pH, the reaction progress (short- or long-term alteration) and the calcium concentration. Glass alteration is thus controlled by specific mechanisms depending on the solution chemistry: calcium complexation at the glass surface, precipitation of calcium silicate hydrates (C-S-H) or calcium incorporation in the altered layer. These findings highlight the impact of silicon-calcium interactions on glass durability and open the way for a better understanding of glass-cement mixing in civil engineering applications as well as in nuclear waste storage.

Emerging drugs for neuropathic pain.

PubMed

Gilron, Ian; Dickenson, Anthony H

2014-09-01

Neuropathic pain is a costly and disabling condition, which affects up to 8% of the population. Available therapies often provide incomplete pain relief and treatment-related side effects are common. Preclinical neuropathic pain models have facilitated identification of several promising targets, which have progressed to human clinical phases of evaluation. A systematic database search yielded 25 new molecular entities with specified pharmacological mechanisms that have reached Phase II or III clinical trials. These include calcium channel antagonists, vanilloid receptor antagonists, potassium channel agonists, NMDA antagonists, novel opioid receptor agonists, histamine H3 receptor antagonists, a novel sodium channel antagonist, serotonin modulators, a novel acetylcholine receptor agonist, α-2b adrenoreceptor agonist, cannabinoid CB2 receptor agonist, nitric oxide synthase inhibitor, orexin receptor antagonist, angiotensin II 2 antagonist, imidazoline I2 receptor agonist, apoptosis inhibitor and fatty acid amide hydrolase inhibitor. Although the diversity of pharmacological mechanisms of interest emphasise the complexity of neuropathic pain transmission, the considerable number of agents under development reflect a continued enthusiasm in drug development for neuropathic pain. Ongoing enhancements in methodology of both preclinical and clinical research and closer translation in both directions are expected to more efficiently identify new agents, which will improve the management of neuropathic pain.

Spasmolytic effect of citral and extracts of Cymbopogon citratus on isolated rabbit ileum.

PubMed

Devi, Ramachandran Chitra; Sim, Si Mui; Ismail, Rosnah

2011-01-01

Cymbopogon citratus, commonly known as lemongrass, has been shown to have antioxidant, antimicrobial and chemo-protective properties. Citral, a monoterpenoid, is the major constituent of C. citratus that gives off a lemony scent and is postulated to be responsible for most of its actions. In addition, C. citratus has been traditionally used to treat gastrointestinal discomforts, however, the scientific evidence for this is still lacking. Thus, the aim of the present study was to investigate the effect of the extracts of various parts of C. citratus (leaves, stems and roots) and citral on the visceral smooth muscle activity of rabbit ileum. The effect of the test substances were tested on the spontaneous contraction, acetylcholine (ACh)- and KCl-induced contractions. Citral at doses between 0.061 mM to 15.6 mM and the extract of leaves at doses between 0.001 mg/mL to 1 mg/mL significantly reduced the spontaneous, ACh- and KCl-induced ileal contractions. When the ileum was incubated in K(+)-rich-Ca(2+)-free Tyrode's solution, it showed only minute contractions. However, the strength of contraction was increased with the addition of increasing concentrations of CaCl(2). The presence of citral almost abolished the effect of adding CaCl(2), while the leaf extract shifted the calcium concentration-response curve to the right, suggesting a calcium antagonistic effect. These results were similar to that elicited by verapamil, a known calcium channel blocker. In addition, the spasmolytic effect of citral was observed to be reduced by the nitric oxide synthase inhibitor, L-NAME. In conclusion, citral and the leaf extract of C. citratus exhibited spasmolytic activity and it appeared that they may act as calcium antagonists. Furthermore, the relaxant effect of citral, but not that of the leaf extract may be mediated by nitric oxide suggesting the presence of other chemical components in the leaf extract other than citral.

Modulation of the activity of midbrain central gray substance neurons by calcium channel agonists and antagonists in vitro.

PubMed

Yakhnitsa, V A; Pilyavskii, A I; Limansky, Y P; Bulgakova, N V

1996-01-01

Changes in the background impulse activity of midbrain central gray substance neurons have been studied on slice preparations from the rat midbrain upon application of calcium-free solution, an activator of calcium channels, BAY-K 8644 (10 nM), organic (verapamil, 40 microM; D600, 10 microM; nifedipine, 1-10 microM; amiloride, 1 microM) and inorganic (Co2+, 1.5 mM) calcium channel blockers. Besides BAY-K 8644, all the substances inhibited most of the neurons studied. Verapamil, BAY-K 8644 and Co2+ also revealed facilitatory effects. Facilitatory action of BAY-K was most effective in silent neurons and in those previously inhibited by amiloride. Latent period values of inhibition in calcium-free solution and upon application of organic and inorganic blockers have the following sequence: D600 > amiloride > verapamil > Co2+ > nifedipine > calcium-free solution. Maximum rise time had the following order: amiloride > D600 > nifedipine > verapamil > Co2+ > calcium-free solution. Complete suppression of the neuronal activity induced by amiloride lasted twice as long as that induced by calcium-free solution, Co2+ and nifedipine, and six times as long as verapamil-induced suppression. Preliminary application of calcium channel blockers reduced facilitatory and increased inhibitory effects of serotonin and substance P. Data obtained are discussed with the supposition in mind that inhibition of the function of calcium channels in central gray substance neurons could be one of the mechanisms underlying the analgesic effect of a series of neurotropic agents after their introduction into this structure.

Impact of magnesium:calcium ratio on calcification of the aortic wall

PubMed Central

2017-01-01

Objective An inverse relationship between serum magnesium concentration and vascular calcification has been reported following observational clinical studies. Moreover, several studies have been suggesting a protective effect of magnesium on the vascular calcification. However, the exact mechanism remains elusive, and investigators have speculated among a myriad of potential actions. The effect of magnesium on calcification of the aortic wall is yet to be investigated. In the present study, the effects of magnesium and calcium on the metabolism of extracellular PPi, the main endogenous inhibitor of vascular calcification, were investigated in the rat aorta. Approach and results Calcium and magnesium have antagonist effects on PPi hydrolysis in the aortic wall. Km and Ki values for PPi hydrolysis in rat aortic rings were 1.1 mmol/L magnesium and 32 μmol/L calcium, respectively, but ATP hydrolysis was not affected with calcium. Calcium deposition in the rat aortic wall dramatically increased when the magnesium concentration was increased (ratio of Mg:Ca = 1:1; 1.5 mmol/L calcium and 1.5 mmol/L magnesium) respect to low magnesium concentration (ratio Mg:Ca = 1:3, 1.5 mmol/L calcium and 0.75 mmol/L magnesium). Conclusion Data from observational clinical studies showing that the serum magnesium concentration is inversely correlated with vascular calcification could be reinterpreted as a compensatory regulatory mechanism that reduces both PPi hydrolysis and vascular calcification. The impact of magnesium in vascular calcification in humans could be studied in association with calcium levels, for example, as the magnesium:calcium ratio. PMID:28570619

Preventive treatment of migraine.

PubMed

Silberstein, Steven D

2005-01-01

Migraine preventive therapy, even in the absence of a headache, is given in an attempt to reduce the frequency, duration, or severity of attacks. Circumstances that might warrant preventive treatment include disabling migraine attacks, the overuse of acute medications or failure of or contraindication to acute medications, troublesome side effects from medication, hemiplegic migraine, or very frequent headaches (more than 2 a week). The major medication groups for preventive treatment include anticonvulsants, antidepressants, b-adrenergic blockers, calcium channel antagonists, serotonin antagonists, neurotoxins, nonsteroidal anti-inflammatory drugs, and others. If preventive medication is indicated, the agent preferentially should be chosen from one of the first-line categories, based on the drug's side-effect profile and the patient's coexistent and comorbid conditions.

Calcium antagonistic activity of Bacopa monniera in guinea-pig trachea.

PubMed

Channa, Shabana; Dar, Ahsana

2012-01-01

To demonstrate the calcium antagonistic property of ethanol extract of Bacopa monniera in guinea-pig trachea. The dose response curves of CaCl(2) (1 × 10(-5) to 1 × 10(-1) M) were constructed in the absence and presence of ethanol extract of Bacopa monniera (100, 500 and 700 μg/ml) or nifedipine (1 × 10(-6) M) in guinea-pig trachea in calcium free high K(+)-MOPS-PSS (3-(N-morpholino)-propanesulphonic acid physiological salt solution). The data was analyzed by ANOVA followed by least significant difference test or by Student's 't' test for unequal variance when appropriate. A probability of at least P < 0.05 was considered statistically significant. The plant extract (500 and 700 μg/ml) significantly (P < 0.05) depressed and shifted the calcium concentration-response curves (1 × 10(-3)- 1 × 10(-1) M) to rightward similar to that of nifedipine. Bacopa monniera extract exhibited calcium channel blocking activity in guinea-pig tracheal smooth muscles that may rationalize its relaxant action on guinea-pig trachea and its traditional use in respiratory disorders.

The effects of crustacean cardioactive peptide on locust oviducts are calcium-dependent.

PubMed

Donini, Andrew; Lange, Angela B

2002-04-01

The role of calcium as a second messenger in the crustacean cardioactive peptide (CCAP)-induced contractions of the locust oviducts was investigated. Incubation of the oviducts in a calcium-free saline containing, a preferential calcium cation chelator, or an extracellular calcium channel blocker, abolished CCAP-induced contractions, indicating that the effects of CCAP on the oviducts are calcium-dependent. In contrast, sodium free saline did not affect CCAP-induced contractions. Co-application of CCAP to the oviducts with preferential L-type voltage-dependent calcium channel blockers reduced CCAP-induced contractions by 32-54%. Two preferential T-type voltage-dependent calcium channel blockers both inhibited CCAP-induced oviduct contractions although affecting different components of the contractions. Amiloride decreased the tonic component of CCAP-induced contractions by 40-55% and flunarizine dihydrochloride decreased the frequency of CCAP-induced phasic contractions by as much as 65%, without affecting tonus. Flunarizine dihydrochloride did not alter the proctolin-induced contractions of the oviducts. Results suggest that the actions of CCAP are partially mediated by voltage-dependent calcium channels similar to vertebrate L-type and T-type channels. High-potassium saline does not abolish CCAP-induced contractions indicating the presence of receptor-operated calcium channels that mediate the actions of CCAP on the oviducts. The involvement of calcium from intracellular stores in CCAP-induced contractions of the oviducts is likely since, an intracellular calcium antagonist decreased CCAP-induced contractions by 30-35%.

Effects of calcium antagonists on isolated bovine cerebral arteries: inhibition of constriction and calcium-45 uptake induced by potassium or serotonin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wendling, W.W.; Harakal, C.

1987-05-01

The purpose of this study was to determine the mechanisms by which organic calcium channel blockers inhibit cerebral vasoconstriction. Isolated bovine middle cerebral arteries were cut into rings to measure contractility or into strips to measure radioactive calcium (/sup 45/Ca) influx and efflux. Calcium channel blockers (10(-5) M verapamil or 3.3 X 10(-7) M nifedipine) and calcium-deficient solutions all produced near-maximal inhibition of both potassium- and serotonin-induced constriction. In calcium-deficient solutions containing potassium or serotonin, verapamil and nifedipine each blocked subsequent calcium-induced constriction in a competitive manner. Potassium and serotonin significantly increased /sup 45/Ca uptake into cerebral artery strips duringmore » 5 minutes of /sup 45/Ca loading; for potassium /sup 45/Ca uptake increased from 62 to 188 nmol/g, and for serotonin from 65 to 102 nmol/g. Verapamil or nifedipine had no effect on basal /sup 45/Ca uptake but significantly blocked the increase in /sup 45/Ca uptake induced by potassium or serotonin. Potassium, and to a lesser extent serotonin, each induced a brief increase in the rate of /sup 45/Ca efflux into calcium-deficient solutions. Verapamil or nifedipine had no effect on basal or potassium-stimulated /sup 45/Ca efflux. The results demonstrate that verapamil and nifedipine block /sup 45/Ca uptake through both potential-operated (potassium) and receptor-operated (serotonin) channels in bovine middle cerebral arteries.« less

Novel Therapeutic Strategies for Reducing Right Heart Failure Associated Mortality in Fibrotic Lung Diseases

PubMed Central

Levy, Matthew; Oyenuga, Olusegun

2015-01-01

Fibrotic lung diseases carry a significant mortality burden worldwide. A large proportion of these deaths are due to right heart failure and pulmonary hypertension. Underlying contributory factors which appear to play a role in the mechanism of progression of right heart dysfunction include chronic hypoxia, defective calcium handling, hyperaldosteronism, pulmonary vascular alterations, cyclic strain of pressure and volume changes, elevation of circulating TGF-β, and elevated systemic NO levels. Specific therapies targeting pulmonary hypertension include calcium channel blockers, endothelin (ET-1) receptor antagonists, prostacyclin analogs, phosphodiesterase type 5 (PDE5) inhibitors, and rho-kinase (ROCK) inhibitors. Newer antifibrotic and anti-inflammatory agents may exert beneficial effects on heart failure in idiopathic pulmonary fibrosis. Furthermore, right ventricle-targeted therapies, aimed at mitigating the effects of functional right ventricular failure, include β-adrenoceptor (β-AR) blockers, angiotensin-converting enzyme (ACE) inhibitors, antioxidants, modulators of metabolism, and 5-hydroxytryptamine-2B (5-HT2B) receptor antagonists. Newer nonpharmacologic modalities for right ventricular support are increasingly being implemented. Early, effective, and individualized therapy may prevent overt right heart failure in fibrotic lung disease leading to improved outcomes and quality of life. PMID:26583148

Does calcium influx regulate melatonin production through the circadian pacemaker in chick pineal cells? Effects of nitrendipine, Bay K 8644, Co2+, Mn2+, and low external Ca2+.

PubMed

Zatz, M; Mullen, D A

1988-11-01

We have recently described a system, using dispersed chick pineal cells in static culture, which displays a persistent, photosensitive, circadian rhythm of melatonin production and release. Here, we describe the effects of nitrendipine (NTR) (a dihydropyridine 'antagonist' of L-type calcium channels), Bay K 8644 (BK) (a dihydropyridine calcium channel 'agonist'), cobalt and manganese ions (both inorganic calcium channel blockers), and low external calcium concentrations, on the melatonin rhythm. NTR inhibited and BK stimulated melatonin output; they were potent and effective. Co2+, Mn2+, and low external Ca2+ markedly inhibited melatonin output. These results support a role for calcium influx through voltage-dependent calcium channels (L-type) in the regulation of melatonin production. Four or 8 h pulses of white light or darkness, in otherwise constant red light, cause, in addition to acute effects, phase-dependent phase shifts of the melatonin rhythm in subsequent cycles. Such phase shifts indicate an effect on (proximal to) the pacemaker generating the rhythm. Four or 8 h pulses of NTR, BK, Co2+, or low Ca2+, however, did not appreciably alter the phase of subsequent melatonin cycles. Neither did BK interfere with phase shifts induced by light pulses. Mn2+ pulses did induce phase-dependent phase shifts, but, unlike those evoked by light or dark pulses, these were all delays. Such effects of Mn2+ in other systems have been attributed to, and are characteristic of, 'metabolic inhibitors'. On balance, the results fail to support a prominent role for calcium influx in regulating the pacemaker underlying the circadian rhythm in chick pineal cells. Rather, calcium influx appears to regulate melatonin production primarily by acting on the melatonin-synthesizing apparatus, distal to the pacemaker.

Brain Damage from Soman-Induced Seizures Is Greatly Exacerbated by Dimethyl sulfoxide (DMSO): Modest Neuroprotection by 2-Aminoethyl diphenylborinate (2- APB), a Transient Receptor Potential Channel Inhibitor and Inositol 1,4,5-triphosphate Receptor Antagonist

DTIC Science & Technology

2008-03-04

whereby the depletion of calcium from intracellular stores (e.g., via IP3 or ryanodine receptor activation ) signals the opening of calcium permeable TRP...system and allowed at least 30 min to acclimate. Baseline ECoG activity and behavior were monitored for at least 15 min. Following baseline recordings...used because the former does not cross the blood-brain-barrier and will not diminish seizure activity or interfere with central effects of the

Metabolic changes of cultured DRG neurons induced by adenosine using confocal microscopy imaging

NASA Astrophysics Data System (ADS)

Zheng, Liqin; Huang, Yimei; Chen, Jiangxu; Wang, Yuhua; Yang, Hongqin; Zhang, Yanding; Xie, Shusen

2012-12-01

Adenosine exerts multiple effects on pain transmission in the peripheral nervous system. This study was performed to use confocal microscopy to evaluate whether adenosine could affect dorsal root ganglia (DRG) neurons in vitro and test which adenosine receptor mediates the effect of adenosine on DRG neurons. After adding adenosine with different concentration, we compared the metabolic changes by the real time imaging of calcium and mitochondria membrane potential using confocal microscopy. The results showed that the effect of 500 μM adenosine on the metabolic changes of DRG neurons was more significant than others. Furthermore, four different adenosine receptor antagonists were used to study which receptor mediated the influences of adenosine on the cultured DRG neurons. All adenosine receptor antagonists especially A1 receptor antagonist (DPCPX) had effect on the Ca2+ and mitochondria membrane potential dynamics of DRG neurons. The above studies demonstrated that the effect of adenosine which may be involved in the signal transmission on the sensory neurons was dose-dependent, and all the four adenosine receptors especially the A1R may mediate the transmission.

An integrated platform for simultaneous multi-well field potential recording and Fura-2-based calcium transient ratiometry in human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes.

PubMed

Rast, Georg; Weber, Jürgen; Disch, Christoph; Schuck, Elmar; Ittrich, Carina; Guth, Brian D

2015-01-01

Human induced pluripotent stem cell-derived cardiomyocytes are available from various sources and they are being evaluated for safety testing. Several platforms are available offering different assay principles and read-out parameters: patch-clamp and field potential recording, imaging or photometry, impedance measurement, and recording of contractile force. Routine use will establish which assay principle and which parameters best serve the intended purpose. We introduce a combination of field potential recording and calcium ratiometry from spontaneously beating cardiomyocytes as a novel assay providing a complementary read-out parameter set. Field potential recording is performed using a commercial multi-well multi-electrode array platform. Calcium ratiometry is performed using a fiber optic illumination and silicon avalanche photodetectors. Data condensation and statistical analysis are designed to enable statistical inference of differences and equivalence with regard to a solvent control. Simultaneous recording of field potentials and calcium transients from spontaneously beating monolayers was done in a nine-well format. Calcium channel blockers (e.g. nifedipine) and a blocker of calcium store release (ryanodine) can be recognized and discriminated based on the calcium transient signal. An agonist of L-type calcium channels, FPL 64176, increased and prolonged the calcium transient, whereas BAY K 8644, another L-type calcium channel agonist, had no effect. Both FPL 64176 and various calcium channel antagonists have chronotropic effects, which can be discriminated from typical "chronotropic" compounds, like (±)isoprenaline (positive) and arecaidine propargyl ester (negative), based on their effects on the calcium transient. Despite technical limitations in temporal resolution and exact matching of composite calcium transient with the field potential of a subset of cells, the combined recording platform enables a refined interpretation of the field potential recording and a more reliable identification of drug effects on calcium handling. Copyright © 2015 Elsevier Inc. All rights reserved.

Neurotoxicity Induced by Bupivacaine via T-Type Calcium Channels in SH-SY5Y Cells

PubMed Central

Wen, Xianjie; Xu, Shiyuan; Liu, Hongzhen; Zhang, Quinguo; Liang, Hua; Yang, Chenxiang; Wang, Hanbing

2013-01-01

There is concern regarding neurotoxicity induced by the use of local anesthetics. A previous study showed that an overload of intracellular calcium is involved in the neurotoxic effect of some anesthetics. T-type calcium channels, which lower the threshold of action potentials, can regulate the influx of calcium ions. We hypothesized that T-type calcium channels are involved in bupivacaine-induced neurotoxicity. In this study, we first investigated the effects of different concentrations of bupivacaine on SH-SY5Y cell viability, and established a cell injury model with 1 mM bupivacaine. The cell viability of SH-SY5Y cells was measured following treatment with 1 mM bupivacaine and/or different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride, an antagonist of T-type calcium channels for 24 h. In addition, we monitored the release of lactate dehydrogenase, cytosolic Ca2+ ([Ca2+]i), cell apoptosis and caspase-3 expression. SH-SY5Y cells pretreated with different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride improved cell viability, reduced lactate dehydrogenase release, inhibited apoptosis, and reduced caspase-3 expression following bupivacaine exposure. However, the protective effect of NNC 55-0396 dihydrochloride plateaued. Overall, our results suggest that T-type calcium channels may be involved in bupivacaine neurotoxicity. However, identification of the specific subtype of T calcium channels involved requires further investigation. PMID:23658789

Effect of clazosentan, a selective endothelin A receptor antagonist, and tezosentan, a dual endothelin A/B antagonist, on pulsatile shear stress induced constriction of the iliac in the anaesthetized pig.

PubMed

Ruane-O'Hora, Therese; Rae, Mark George; Markos, Farouk

2011-08-01

1. The effects of changes in mean and pulsatile shear stress on the diameter of the iliac of the anaesthetized pig were investigated in the presence of clazosentan and tezosentan. 2. A total of 17 pigs were used. Mean shear stress was increased by infusing acetylcholine downstream (2-20 μg/min) through the deep femoral artery. Pulsatile shear stress was enhanced first by injecting varying volumes (1-10 mL) of calcium gluconate (stock 10 mg/mL) directly into the left ventricle. Second, by electrical stimulation of the left sympathetic nerves to the heart (1-16 Hz, 4 min duration, supramaximal voltage). 3. An increase in mean shear stress induced a vasodilation that was not altered significantly by the selective endothelin A antagonist clazosentan (10 mg/kg i.v.). Similarly, the vasoconstriction induced by an increase in pulsatile shear stress brought about by either calcium gluconate injections or left sympathetic nerve stimulation was unaffected by clazosentan. However, tezosentan (10 mg/kg i.v.), significantly attenuated the vasoconstriction induced by an increase in pulsatile shear stress. 4. In conclusion, an increase in pulsatile shear stress causes vasoconstriction of the pig iliac artery, which is attenuated by dual endothelin receptor antagonism, but not by specific endothelin A blockade. © 2011 The Authors. Clinical and Experimental Pharmacology and Physiology © 2011 Blackwell Publishing Asia Pty Ltd.

Calcium antagonistic activity of Bacopa monniera in guinea-pig trachea

PubMed Central

Channa, Shabana; Dar, Ahsana

2012-01-01

Objective: To demonstrate the calcium antagonistic property of ethanol extract of Bacopa monniera in guinea-pig trachea. Materials and Methods: The dose response curves of CaCl2 (1 × 10-5 to 1 × 10-1 M) were constructed in the absence and presence of ethanol extract of Bacopa monniera (100, 500 and 700 μg/ml) or nifedipine (1 × 10-6 M) in guinea-pig trachea in calcium free high K+-MOPS-PSS (3-(N-morpholino)-propanesulphonic acid physiological salt solution). The data was analyzed by ANOVA followed by least significant difference test or by Student's ‘t’ test for unequal variance when appropriate. A probability of at least P < 0.05 was considered statistically significant. Results: The plant extract (500 and 700 μg/ml) significantly (P < 0.05) depressed and shifted the calcium concentration-response curves (1 × 10-3- 1 × 10-1 M) to rightward similar to that of nifedipine. Conclusions: Bacopa monniera extract exhibited calcium channel blocking activity in guinea-pig tracheal smooth muscles that may rationalize its relaxant action on guinea-pig trachea and its traditional use in respiratory disorders. PMID:23087517

Calcium-sensing receptor (CASR) is involved in porcine in vitro fertilisation and early embryo development.

PubMed

Liu, C; Liu, Y; Larsen, K; Hou, Y P; Callesen, H

2018-01-01

It has been demonstrated that extracellular calcium is necessary in fertilisation and embryo development but the mechanism is still not well understood. The present study mainly focussed on the extracellular calcium effector called the calcium-sensing receptor (CASR) and examined its expression in porcine gametes and embryos and its function during fertilisation and early embryo development. By using reverse transcription polymerase chain reaction, CASR was found to be expressed in porcine oocytes, spermatozoa and embryos at different developmental stages. Functionally, medium supplementation with a CASR agonist or an antagonist during in vitro fertilisation (IVF) and in vitro culture (IVC) was tested. During fertilisation, the presence of a CASR agonist increased sperm penetration rate and decreased polyspermy rate leading to an increased normal fertilisation rate. During embryo development, for the IVF embryos, agonist treatment during IVC significantly increased cleavage rate and blastocyst formation rate compared with the control group. Furthermore, parthenogenetically activated embryos showed similar results with lower cleavage and blastocyst formation rates in the antagonist group than in the other groups. It was concluded that CASR, as the effector of extracellular calcium, modulates porcine fertilisation and early embryo development.

NMDA receptor-antagonistic properties of hyperforin, a constituent of St. John's Wort.

PubMed

Kumar, Vikas; Mdzinarishvili, Alexander; Kiewert, Cornelia; Abbruscato, Thomas; Bickel, Ulrich; van der Schyf, Cornelis J; Klein, Jochen

2006-09-01

Extracts of the medicinal plant St. John's wort (Hypericum perforatum) are widely used for the treatment of affective disorders. Hyperforin, a constituent of St. John's wort, is known to modulate the release and re-uptake of various neurotransmitters, an action that likely underlies its antidepressive activity. We now report that hyperforin also has N-methyl-D-aspartate (NMDA)-antagonistic effects. Hyperforin (10 microM) was found to inhibit the NMDA-induced calcium influx into cortical neurons. In rat hippocampal slices, hyperforin inhibited the NMDA-receptor-mediated release of choline from phospholipids. Hyperforin also antagonized the increase of water content in freshly isolated hippocampal slices, and it counteracted, at 3 and 10 microM, the increase of water content induced by NMDA. Hyperforin was inactive, however, in two in vivo models of brain edema formation, middle cerebral artery occlusion and water intoxication in mice. In conclusion, hyperforin has NMDA-receptor-antagonistic and potential neuroprotective effects in vitro. This effect may contribute to the therapeutic effectiveness of St. John's wort extracts in some situations, for example, for relapse prevention in alcoholism.

The inhibitory effect of somatostatin peptides on the rat anococcygeus muscle in vitro.

PubMed Central

Priestley, T.; Woodruff, G. N.

1988-01-01

1. Electrically evoked contractions of the rat anococcygeus muscle were inhibited in a concentration-dependent manner by somatostatin-14 (SS14), -28 (SS28) and two synthetic hexapeptide analogues: L-363,301 (Pro-Phe-D-Trp-Lys-Thr-Phe) and L-363,586 (N-Me-Ala-Tyr-D-Trp-Lys-Val-Phe), with pIC50 values of 7.41, 7.38, 7.07 and 8.34, respectively. 2. The inhibitory effects of SS14 were dependent on stimulation frequency and external calcium ion concentration. Calcium behaved as a non-competitive antagonist of SS14, it reduced the maximal inhibitory effect of the peptide and at a concentration of 5.08 mM it significantly affected the pIC50 value. 3. SS14 (3 x 10(-7) M) did not affect the tonic actions of bath-applied noradrenaline in the absence of field stimulation. 4. The effects of SS14 persisted in naloxone (10(-5) M) and were, therefore, not due to an action at opiate receptors. Furthermore, experiments involving the lyophilization of bath contents, showed no evidence to support an indirect mechanism involving the release of an endogenous inhibitory substance. 5. High concentrations (10(-5) M) of SS14 or L-363,301 inhibited the relaxation response evoked by electrical stimulation of guanethidine (3 x 10(-4) M)-treated preparations. 6. These results are consistent with similar actions of SS14 on other smooth muscle preparations and are presumed to reflect a presynaptic inhibition of transmitter release by a direct action on somatostatin receptors. The antagonistic effect of calcium on this response is discussed with reference to a possible role in receptor desensitization. PMID:2900039

Evaluation of the calcium-antagonist, antidiarrhoeic and central nervous system activities of Baccharis serraefolia.

PubMed

Tortoriello, J; Aguilar-Santamaría, L

1996-09-01

Baccharis serraefolia is a widely used plant to treat diarrhoea in Mexican traditional medicine. Although the methanolic extract of this plant has shown an important dose-dependent spasmolytic activity, its underlying mechanism has not been studied. In the present work, the methanolic extract of B. serraefolia significantly delayed the onset of tonic seizures induced by strychnine and pentylenetetrazol; besides, it diminished the death rate and number of animals that exhibited convulsions. It produced potentiation of the hypnotic effect of pentobarbital. Oral administration produced an inhibition of gastrointestinal transit in mice as effective as that produced by loperamide. As to the effect on smooth muscles, the active extract produced an inhibition of contraction induced electrically, which could not be reversed by naloxone. The calcium concentration-contraction curve showed a rightward displacement when the extract was added to isolated guinea pig ileum depolarized with high K+ and cumulative concentrations of Ca2+. The results suggest that the methanolic extract does not interact with classical opiate receptors and its effects, at least that produced on smooth muscle, may be due to a probable interference with calcium influx and/or calcium release from an intra-cellular store.

Gonadotrophin-releasing activity of neurohypophysial hormones: II. The pituitary oxytocin receptor mediating gonadotrophin release differs from that of corticotrophs.

PubMed

Evans, J J; Catt, K J

1989-07-01

Neurohypophysial hormones stimulate gonadotrophin release from dispersed rat anterior pituitary cells in vitro, acting through receptors distinct from those which mediate the secretory response to gonadotrophin-releasing hormone (GnRH). The LH response to oxytocin was not affected by the presence of the phosphodiesterase inhibitor, methyl isobutylxanthine, but was diminished in the absence of extracellular calcium and was progressively increased as the calcium concentration in the medium was raised to normal. In addition, the calcium channel antagonist, nifedipine, suppressed oxytocin-stimulated secretion of LH. It is likely that the mechanisms of LH release induced by GnRH and neurohypophysial hormones are similar, although stimulation of gonadotrophin secretion is mediated by separate receptor systems. Oxytocin was more active than vasopressin in releasing LH, but less active in releasing ACTH. The highly selective oxytocin agonist, [Thr4,Gly7]oxytocin, elicited concentration-dependent secretion of LH but had little effect on corticotrophin secretion. The neurohypophysial hormone antagonist analogues, [d(CH2)5Tyr(Me)2]vasopressin, [d(CH2)5Tyr(Me)2,Orn8]vasotocin and [d(CH2)5D-Tyr(Et)2Val4,Cit8]vasopressin, inhibited the LH response to both oxytocin and vasopressin. However, [d(CH2)5Tyr(Me)2]vasopressin was much less effective in inhibiting the ACTH response to the neurohypophysial hormones, and [d(CH2)5Tyr-(Me)2,Orn8]vasotocin and [d(CH2)5D-Tyr(Et)2,Val4,Cit8]vasopressin exhibited no inhibitory activity against ACTH release. Thus, agonist and antagonist analogues of neurohypophysial hormones display divergent activities with regard to LH and ACTH responses, and the neuropeptide receptor mediating gonadotroph activation is clearly different from that on the corticotroph. Whereas the corticotroph receptor is a vasopressin-type receptor an oxytocin-type receptor is responsible for gonadotrophin release by neurohypophysial hormones.

Induction of calcium-dependent nitric oxide synthases by sex hormones.

PubMed

Weiner, C P; Lizasoain, I; Baylis, S A; Knowles, R G; Charles, I G; Moncada, S

1994-05-24

We have examined the effects of pregnancy and sex hormones on calcium-dependent and calcium-independent nitric oxide synthases (NOSs) in the guinea pig. Pregnancy (near term) caused a > 4-fold increase in the activity of calcium-dependent NOS in the uterine artery and at least a doubling in the heart, kidney, skeletal muscle, esophagus, and cerebellum. The increase in NOS activity in the cerebellum during pregnancy was inhibited by the estrogen-receptor antagonist tamoxifen. Treatment with estradiol (but not progesterone) also increased calcium-dependent NOS activity in the tissues examined from both females and males. Testosterone increased calcium-dependent NOS only in the cerebellum. No significant change in calcium-independent NOS activity was observed either during pregnancy or after the administration of any sex hormone. Both pregnancy and estradiol treatment increased the amount of mRNAs for NOS isozymes eNOS and nNOS in skeletal muscle, suggesting that the increases in NOS activity result from enzyme induction. Thus both eNOS and nNOS are subject to regulation by estrogen, an action that could explain some of the changes that occur during pregnancy and some gender differences in physiology and pathophysiology.

P2X1 Receptor-Mediated Ca2+ Influx Triggered by DA-9801 Potentiates Nerve Growth Factor-Induced Neurite Outgrowth.

PubMed

Back, Moon Jung; Lee, Hae Kyung; Lee, Joo Hyun; Fu, Zhicheng; Son, Mi Won; Choi, Sang Zin; Go, Hyo Sang; Yoo, Sungjae; Hwang, Sun Wook; Kim, Dae Kyong

2016-11-16

Nerve growth factor (NGF)-induced neuronal regeneration has emerged as a strategy to treat neuronal degeneration-associated disorders. However, direct NGF administration is limited by the occurrence of adverse effects at high doses of NGF. Therefore, development of a therapeutic strategy to promote the NGF trophic effect is required. In view of the lack of understanding of the mechanism for potentiating the NGF effect, this study investigated molecular targets of DA-9801, a well-standardized Dioscorea rhizome extract, which has a promoting effect on NGF. An increase in intracellular calcium ion level was induced by DA-9801, and chelation of extracellular calcium ions with ethylene-bis(oxyethylenenitrilo)tetraacetic acid (EGTA) suppressed the potentiating effect of DA-9801 on NGF-induced neurite outgrowth. In addition, EGTA treatment reduced the DA-9801-induced phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2), the major mediators of neurite outgrowth. To find which calcium ion-permeable channel contributes to the calcium ion influx induced by DA-9801, we treated PC12 cells with various inhibitors of calcium ion-permeable channels. NF449, a P2X1 receptor selective antagonist, significantly abolished the potentiating effect of DA-9801 on NGF-induced neurite outgrowth and abrogated the DA-9801-induced ERK1/2 phosphorylation. In addition, transfection with siRNA of P2X1 receptor significantly reduced the DA-9801-enhanced neurite outgrowth. In conclusion, calcium ion influx through P2X1 receptor mediated the promoting effect of DA-9801 on NGF-induced neurite outgrowth via ERK1/2 phosphorylation.

The PLC/IP3R/PKC Pathway is Required for Ethanol-enhanced GABA Release

PubMed Central

Kelm, M. Katherine; Weinberg, Richard J.; Criswell, Hugh E.; Breese, George R.

2010-01-01

Summary Research on the actions of ethanol at the GABAergic synapse has traditionally focused on postsynaptic mechanisms, but recent data demonstrate that ethanol also increases both evoked and spontaneous GABA release in many brain regions. Using whole-cell voltage-clamp recordings, we previously showed that ethanol increases spontaneous GABA release at the rat interneuron-Purkinje cell synapse. This presynaptic ethanol effect is dependent on calcium release from internal stores, possibly through activation of inositol 1,4,5-trisphosphate receptors (IP3Rs). After confirming that ethanol targets vesicular GABA release, in the present study we used electron microscopic immunohistochemistry to demonstrate that IP3Rs are located in presynaptic terminals of cerebellar interneurons. Activation of IP3Rs requires binding of IP3, generated through activation of phospholipase C (PLC). We find that the PLC antagonist edelfosine prevents ethanol from increasing spontaneous GABA release. Diacylglycerol generated by PLC and calcium released by activation of the IP3R activate protein kinase C (PKC). Ethanol-enhanced GABA release was blocked by two PKC antagonists, chelerythrine and calphostin C. When a membrane impermeable PKC antagonist, PKC (19-36), was delivered intracellularly to the postsynaptic neuron, ethanol continued to increase spontaneous GABA release. Overall, these results suggest that activation of the PLC/IP3R/PKC pathway is necessary for ethanol to increase spontaneous GABA release from presynaptic terminals onto Purkinje cells. PMID:20206640

Vasorelaxant effect of the analgesic clonixin on rat aorta.

PubMed

Morales, M A; Silva, A; Brito, G; Bustamante, S; Ponce, H; Paeile, C

1995-03-01

1. A novel vasorelaxant effect of clonixinate of L-lysine (Clx), analgesic and anti-inflammatory, was studied in rat aortic rings. 2. Clx completely relaxed aortic rings contracted by KCl 70 mM and together with its analog flunixin exhibited lesser potency but equal efficacy than verapamil. In comparison, indomethacin, which is a more potent cyclo-oxygenase inhibitor relaxed only about 40% of the maximal contraction of aortic rings. 3. Furthermore, Clx antagonized Ca2+ dependent aortic contraction and BAY K-8644 induced aortic contraction suggesting its calcium antagonist character. 4. From these results it can be concluded that the hypotensive effect seen in rats in vivo after Clx i.v. injection arises because of vasodilatory effect of Clx and gives further support to the proposal that the pharmacological mechanism of action of Clx should be calcium antagonism.

Is calcium excess in western diet a major cause of arterial disease?

PubMed

Seely, S

1991-11-01

The daily requirement of a young adult for calcium is 300-400 mg, the quantity consumed in many third-world countries. The dietary intake can be doubled or trebled by consumption of milk, thus half a litre of milk, consumed by many individuals daily in prosperous countries, adds 600 mg. The need for calcium is greatly reduced in old age, so that the excess from a given intake becomes more pronounced. The potential hazard of a high intake is that a small fraction finds its way into soft tissues. The aorta is notably prone to calcification, resulting in loss of elasticity. The aorta and its large branches constitute an elastic reservoir, distended during systole and contracting in diastole. This contraction provides the energy for the maintenance of diastolic pressure, which decreases with the deterioration of elasticity and needs a continually increasing systolic pressure to restore its normal value. The heart is disadvantaged in two ways. Its work is increased by having to eject the systolic volume into a stiffer reservoir, and the diastolic filling of the coronary arteries is reduced. This is the main cause of hypertension in old age--there is no increase in blood pressure with age in undeveloped countries where intake of calcium is low. The best cure would be prevention: the reduction of intake of calcium in prosperous countries. Failing that, phytic acid is suggested as the best calcium antagonist. Phytic acid, a natural product present in grains, converts dietary calcium into insoluble phosphates which pass unabsorbed through the digestive tract. The presently used calcium antagonists are not so satisfactory.

T-type calcium channel antagonism decreases motivation for nicotine and blocks nicotine- and cue-induced reinstatement for a response previously reinforced with nicotine.

PubMed

Uslaner, Jason M; Vardigan, Joshua D; Drott, Jason M; Uebele, Victor N; Renger, John J; Lee, Ariel; Li, Zhaoxia; Lê, A D; Hutson, Pete H

2010-10-15

Recent evidence suggests an involvement of T-type calcium channels in the effects of drugs of abuse. We examined the influence of the novel, potent, and selective T-type calcium channel antagonist [2-(4-cyclopropylphenyl)-N-((1R)-1-{5-[2,2,2-trifluoroethyl]oxo}pyridine-2-yl)ethyl]acetamide] (TTA-A2) (.3, 1, or 3 mg/kg) on motivation for nicotine, as measured by nicotine self-administration on a progressive ratio (PR) schedule, and nicotine- and cue-induced reinstatement for a response previously reinforced with nicotine delivery (n = 11 or 12 Long Evans rats/group). Furthermore, we examined the specificity of the TTA-A2 effects by characterizing its influence on PR responding for food (in the absence or presence of nicotine-potentiated responding), food- versus nicotine-induced cue-potentiated reinstatement for a response previously reinforced by food administration (n = 11 or 12 Wistar Hannover rats/group), and its ability to induce a conditioned place aversion. TTA-A2 dose-dependently decreased self-administration of nicotine on a PR schedule and the ability of both nicotine and a cue paired with nicotine to reinstate responding. The effects were specific for nicotine's incentive motivational properties, as TTA-A2 did not influence responding for food on a PR schedule but did attenuate the ability of nicotine to potentiate responding for food. Likewise, TTA-A2 did not alter food-induced cue-potentiated reinstatement for a response previously reinforced by food but did decrease nicotine-induced cue-potentiated reinstatement. Finally, TTA-A2 did not produce an aversive state, as indicated by a lack of ability to induce conditioned place aversion. These data suggest that T-type calcium channel antagonists have potential for alleviating nicotine addiction by selectively decreasing the incentive motivational properties of nicotine. Copyright © 2010 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Novel cellular bouton structure activated by ATP in the vascular wall of porcine retinal arterioles.

PubMed

Misfeldt, Mikkel Wölck; Aalkjaer, Christian; Simonsen, Ulf; Bek, Toke

2010-12-01

The retinal blood flow is regulated by the tone of resistance arterioles, which is influenced by purinergic compounds such as adenosine and adenosine 5'-triphosphate (ATP) released from the retinal tissue. However, it is unknown what cellular elements in the perivascular retina are responsible for the effect of purines on the tone of retinal arterioles. Porcine retinal arterioles were loaded with the calcium-sensitive fluorophore Oregon green. The vessels were mounted in a confocal myograph for simultaneous recordings of tone and calcium activity in cells of the vascular wall during stimulation with ATP and adenosine, with and without modifiers of these compounds. Additionally, immunohistochemistry was used to localize elements with calcium activity in the vascular wall. Hyperfluorescence indicating calcium activity was recorded in a population of abundant round boutons interspersed in a network of vimentin-positive processes located immediately external to the smooth muscle cell layer but internal to the perivascular glial cells. These structures showed calcium activity when the vessel was relaxed with ATP but not when it was relaxed with adenosine. Ryanodine reduced calcium activity in the boutons, whereas the ATP antagonist adenosine-5'-O-(α, β- methylene diphosphate) reduced calcium activity in both the boutons and vascular tone. The vasodilating effect of purines in porcine retinal tissue involves ATP-dependent calcium activity in a layer of cellular boutons located external to the vascular smooth muscle cells and internal to the perivascular glial cells.

Synthetic peptides corresponding to human follicle-stimulating hormone (hFSH)-beta-(1-15) and hFSH-beta-(51-65) induce uptake of 45Ca++ by liposomes: evidence for calcium-conducting transmembrane channel formation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Grasso, P.; Santa-Coloma, T.A.; Reichert, L.E. Jr.

1991-06-01

We have previously described FSH receptor-mediated influx of 45Ca++ in cultured Sertoli cells from immature rats and receptor-enriched proteoliposomes via activation of voltage-sensitive and voltage-independent calcium channels. We have further shown that this effect of FSH does not require cholera toxin- or pertussis toxin-sensitive guanine nucleotide binding protein or activation of adenylate cyclase. In the present study, we have identified regions of human FSH-beta-subunit which appear to be involved in mediating calcium influx. We screened 11 overlapping peptide amides representing the entire primary structure of hFSH-beta-subunit for their effects on 45Ca++ flux in FSH receptor-enriched proteoliposomes. hFSH-beta-(1-15) and hFSH-beta-(51-65) inducedmore » uptake of 45Ca++ in a concentration-related manner. This effect of hFSH-beta-(1-15) and hFSH-beta-(51-65) was also observed in liposomes lacking incorporated FSH receptor. Reducing membrane fluidity by incubating liposomes (containing no receptor) with hFSH-beta-(1-15) or hFSH-beta-(51-65) at temperatures lower than the transition temperatures of their constituent phospholipids resulted in no significant (P greater than 0.05) difference in 45Ca++ uptake. The effectiveness of the calcium ionophore A23187, however, was abolished. Ruthenium red, a voltage-independent calcium channel antagonist, was able to completely block uptake of 45Ca++ induced by hFSH-beta-(1-15) and hFSH-beta-(51-65) whereas nifedipine, a calcium channel blocker specific for L-type voltage-sensitive calcium channels, was without effect. These results suggest that in addition to its effect on voltage-sensitive calcium channel activity, interaction of FSH with its receptor may induce formation of transmembrane aqueous channels which also facilitate influx of extracellular calcium.« less

Calcium channel blockers and esophageal sclerosis: should we expect exacerbation of interstitial lung disease?

PubMed

Seretis, Charalampos; Seretis, Fotios; Gemenetzis, George; Liakos, Nikolaos; Pappas, Apostolos; Gourgiotis, Stavros; Lagoudianakis, Emmanuel; Keramidaris, Dimitrios; Salemis, Nikolaos

2012-01-01

Esophageal sclerosis is the most common visceral manifestation of systemic sclerosis, resulting in impaired esophageal clearance and retention of ingested food; in addition, co-existence of lung fibrosis with esophageal scleroderma is not uncommon. Both the progression of generalized connective tissue disorders and the damaging effect of chronic aspiration due to esophageal dysmotility appear to be involved in this procedure of interstitial fibrosis. Nifedipine is a widely prescribed calcium antagonist in a significant percentage of rheumatologic patients suffering from Raynaud syndrome, in order to inhibit peripheral vasospasm. Nevertheless, blocking calcium channels has proven to contribute to exacerbation of gastroesophageal reflux, which consequently can lead to chronic aspiration. We describe the case of severe exacerbation of interstitial lung disease in a 76-year-old female with esophageal sclerosis who was treated with oral nifedipine for Raynaud syndrome.

L-type Voltage-Gated Calcium Channels in Conditioned Fear: A Genetic and Pharmacological Analysis

ERIC Educational Resources Information Center

McKinney, Brandon C.; Sze, Wilson; White, Jessica A.; Murphy, Geoffrey G.

2008-01-01

Using pharmacological approaches, others have suggested that L-type voltage-gated calcium channels (L-VGCCs) mediate both consolidation and extinction of conditioned fear. In the absence of L-VGCC isoform-specific antagonists, we have begun to investigate the subtype-specific role of LVGCCs in consolidation and extinction of conditioned fear…

Comparison of the effect of the antacid Rennie versus low-dose H2-receptor antagonists (ranitidine, famotidine) on intragastric acidity.

PubMed

Netzer, P; Brabetz-Höfliger, A; Bründler, R; Flogerzi, B; Hüsler, J; Halter, F

1998-04-01

Symptoms of functional dyspepsia are common and patients often self-medicate with antacids, or with low-dose H2-antagonists which are available as over-the-counter medications. To date, there has been limited information available comparing the effects on intragastric acidity of these two types of over-the-counter medication. Therefore we studied the effect of the antacid Rennie and two H2-antagonists on the intragastric pH of fasting volunteers. Sixteen healthy, fasting volunteers were randomized into a double-blind, placebo-controlled, four-way crossover study comparing Rennie (calcium-magnesium carbonate) 1360 mg, ranitidine 75 mg, famotidine 10 mg and placebo. Their effect on gastric pH was monitored by a 4-h gastric pH-metry. The primary efficacy parameter was the time lag before an intragastric pH > 3.0 was reached after drug administration. The median time lag before pH > 3.0 was reached after drug administration was 5.8 min for Rennie, 64.9 min for ranitidine, 70.1 min for famotidine and 240.0 min for placebo. The percentage of time with values of pH > 3.0 was 10.4% for Rennie, 61.4% for ranitidine, 56.6% for famotidine and 1.4% for placebo. The onset of action in fasting volunteers was significantly faster with the antacid than with the two H2-antagonists. The duration of action was significantly longer with an H2-antagonist than with the antacid. This suggests that the two products should be used for different indications: antacids are superior for rapid pain relief, whereas H2-antagonists might be better for symptom prophylaxis--for example for nocturnal dyspepsia.

PDGF-mediated protection of SH-SY5Y cells against Tat toxin involves regulation of extracellular glutamate and intracellular calcium

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhu Xuhui; Department of Laboratory Medicine, Tongji Hospital and Tongji Medical College of Huazhong University of Science and Technology, Wuhan; Yao Honghong

2009-10-15

The human immunodeficiency virus (HIV-1) protein Tat has been implicated in mediating neuronal apoptosis, one of the hallmark features of HIV-associated dementia (HAD). Mitigation of the toxic effects of Tat could thus be a potential mechanism for reducing HIV toxicity in the brain. In this study we demonstrated that Tat-induced neurotoxicity was abolished by NMDA antagonist-MK801, suggesting the role of glutamate in this process. Furthermore, we also found that pretreatment of SH-SY5Y cells with PDGF exerted protection against Tat toxicity by decreasing extracellular glutamate levels. We also demonstrated that extracellular calcium chelator EGTA was able to abolish PDGF-mediated neuroprotection, therebymore » underscoring the role of calcium signaling in PDGF-mediated neuroprotection. We also showed that Erk signaling pathway was critical for PDGF-mediated protection of cells. Additionally, blocking calcium entry with EGTA resulted in suppression of PDGF-induced Erk activation. These findings thus underscore the role of PDGF-mediated calcium signaling and Erk phosphorylation in the protection of cells against HIV Tat toxicity.« less

β-adrenergic Receptor Blocker ICI 118,551 Selectively Increases Intermediate-Conductance Calcium-Activated Potassium Channel (IKCa )-Mediated Relaxations in Rat Main Mesenteric Artery.

PubMed

Ozkan, Melike Hacer; Uma, Serdar

2018-06-01

Endothelial IK C a and/or SK C a channels play an important role in the control of vascular tone by participating in endothelium-dependent relaxation. Whether β-AR antagonists, mainly used in hypertension, affect endothelial K C a channel function is unknown. In this study, we examined the effect of the β2-AR antagonist and inverse agonist ICI 118,551 on the IK C a /SK C a channel activity by assessing functional relaxation responses to several agonists that stimulate these channels. Mesenteric arterial rings isolated from male Sprague Dawley mounted to organ baths. Acetylcholine elicited IK C a - and SK C a -mediated relaxations that were abolished by TRAM-34 and apamin, respectively. ICI 118,551, which did not dilate the arteries per se, increased the IK C a -mediated relaxations, whereas SK C a -mediated relaxations remained unaltered. Same potentiating effect was also detected on the IK C a -mediated relaxations to carbachol and A23187, but not to NS309. Neither acetylcholine-induced nitric oxide-mediated relaxations nor SNP relaxations changed with ICI 118,551. The PKA inhibitor KT-5720, the selective β2-AR agonist salbutamol, the selective β2-AR antagonist butoxamine, the non-selective β-AR antagonist propranolol, and the inverse agonists carvedilol or nadolol failed to affect the IK C a -mediated relaxations. ICI 118,551-induced increase was not reversed by salbutamol or propranolol as well. Besides, low potassium-induced relaxations in endothelium-removed arteries remained the same in the presence of ICI 118,551. These data demonstrate a previously unrecognized action of ICI 118,551, the ability to potentiate endothelial IK C a channel-mediated vasodilation, through a mechanism independent of β2-AR antagonistic or inverse agonistic action. Instead, the enhancement of acetylcholine relaxation seems likely to occur by a mechanism secondary to endothelial calcium increase. © 2017 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

Resveratrol-induced antinociception is involved in calcium channels and calcium/caffeine-sensitive pools.

PubMed

Pan, Xiaoyu; Chen, Jiechun; Wang, Weijie; Chen, Ling; Wang, Lin; Ma, Quan; Zhang, Jianbo; Chen, Lichao; Wang, Gang; Zhang, Meixi; Wu, Hao; Cheng, Ruochuan

2017-02-07

Resveratrol has been widely investigated for its potential health properties, although little is known about its mechanism in vivo. Previous studies have indicated that resveratrol produces antinociceptive effects in mice. Calcium channels and calcium/caffeine-sensitive pools are reported to be associated with analgesic effect. The present study was to explore the involvement of Ca2+ channel and calcium/caffeine-sensitive pools in the antinociceptive response of resveratrol. Tail-flick test was used to assess antinociception in mice treated with resveratrol or the combinations of resveratrol with MK 801, nimodipine, CaCl2, ryanodine and ethylene glycol tetraacetic acid (EGTA), respectively. The Ca2+/calmodulin-dependent protein kinase II (CaMKII) and brain-derived neurotrophic factor (BDNF) levels in the spinal cord were also investigated when treated with the above drugs. The results showed that resveratrol increased the tail flick latency in the tail-flick test, in dose-dependent manner. N-methyl-D-aspartate (NMDA) glutamate receptor antagonist MK 801 potentiated the antinociceptive effects of sub-threshold dose of resveratrol at 10 mg/kg. Ca2+ channel blocker, however, abolished the antinociceptive effects of resveratrol. In contrast to these results, EGTA or ryanodine treatment (i.c.v.) potentiated resveratrol-induced antinociception. There was a significant decrease in p-CaMKII and an increase in BDNF expression in the spinal cord when combined with MK 801, nimodipine, ryanodine and EGTA. While an increase in p-CaMKII level and a decrease in BDNF expression were observed when high dose of resveratrol combined with CaCl2. These findings suggest that resveratrol exhibits the antinociceptive effects by inhibition of calcium channels and calcium/caffeine-sensitive pools.

The effects of vasoactive peptide urocortin 2 on hemodynamics in spontaneous hypertensive rat and the role of L-type calcium channel and CRFR2.

PubMed

Liu, Chunna; Liu, Xinyu; Yang, Jing; Duan, Yan; Yao, Hongyue; Li, Fenghua; Zhang, Xia

2015-04-01

Urocortin (UCN) is a newly identified vascular-active peptide that has been shown to reverse cardiovascular remodeling and improve left ventricular (LV) function. The effects and mechanism of urocortin 2 (UCN2) in vivo on the electrical remodeling of left ventricle and the hemodynamics of hypertensive objectives have not been investigated. UCN2 (1 μg/kg/d, 3.5 μg/kg/d or 7 μg/kg/d) was intravenously injected for 2 weeks and its effects on hemodynamics in spontaneously hypertensive rats (SHRs) observed. The whole-cell patch clamp technique was used to explore the effects of UCN2 on the electrical remodeling of left ventricular cardiomyocytes. The flow cytometry method was used to determine the content of fluorescence calcium in myocardium. UCN2 improved the systolic and diastolic function of SHRs as demonstrated by decreased left ventricular systolic pressure (LVSP), left ventricular end diastolic pressure (LVEDP), increased +dp/dtmax and -dp/dtmax and decreased cAMP level. UCN2 inhibited the opening of L-type calcium channel and decreased the calcium channel current of cardiomyocytes. In addition, UCN2 also decreased the contents of fluorescence calcium in SHR myocardium. However, astressin2-B (AST-2B), the antagonist of corticotropin-releasing factor receptor 2 (CRFR2), could reverse the inhibitory effects of UCN2 on calcium channel. UCN2 can modulate electrical remodeling of the myocardium and hemodynamics in an experimental model of SHR via inhibition of L-type calcium channel and CRFR2 in cardiomyocytes. Copyright © 2014 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

Neuronal calcium sensor proteins are direct targets of the insulinotropic agent repaglinide.

PubMed Central

Okada, Miki; Takezawa, Daisuke; Tachibanaki, Shuji; Kawamura, Satoru; Tokumitsu, Hiroshi; Kobayashi, Ryoji

2003-01-01

The NCS (neuronal calcium sensor) proteins, including neurocalcins, recoverins and visinin-like proteins are members of a family of Ca2+-sensitive regulators, each with three Ca2+-binding EF-hand motifs. In plants, lily CCaMK [chimaeric Ca2+/CaM (calmodulin)-dependent protein kinase] and its PpCaMK ( Physcomitrella patens CCaMK) homologue are characterized by a visinin-like domain with three EF-hands. In the present study, in an effort to discover NCS antagonists, we screened a total of 43 compounds using Ca2+-dependent drug affinity chromatography and found that the insulinotropic agent repaglinide targets the NCS protein family. Repaglinide was found to bind to NCS proteins, but not to CaM or S100 proteins, in a Ca2+-dependent manner. Furthermore, the drug antagonized the inhibitory action of recoverin in a rhodopsin kinase assay with IC50 values of 400 microM. Moreover, repaglinide tightly bound to the visinin-like domain of CCaMK and PpCaMK in a Ca2+-dependent manner and antagonized the regulatory function of the domain with IC50 values of 55 and 4 microM for CCaMK and PpCaMK respectively. Although both repaglinide and a potent insulin secretagogue, namely glibenclamide, blocked K(ATP) channels with similar potency, glibenclamide had no antagonizing effect on the Ca2+-stimulated CCaMK and PpCaMK autophosphorylation, mediated by their visinin-like domain. In addition, a typical CaM antagonist, trifluoperazine, had no effect on the CCaMK and PpCaMK autophosphorylation. Repaglinide appears to be the first antagonist of NCS proteins and visinin-like domain-bearing enzymes. It may serve as a useful tool for evaluating the physiological functions of the NCS protein family. In addition, since repaglinide selectively targets NCS proteins among the EF-hand Ca2+-binding proteins, it is a potential lead compound for the development of more potent NCS antagonists. PMID:12844348

Low-concentration hydrogen peroxide can upregulate keratinocyte intracellular calcium and PAR-2 expression in a human keratinocyte-melanocyte co-culture system.

PubMed

Li, Jian; Tang, Lu-Yan; Fu, Wen-Wen; Yuan, Jin; Sheng, You-Yu; Yang, Qin-Ping

2016-12-01

Hydrogen peroxide (H 2 O 2 ) may have a biphasic effect on melanin synthesis and melanosome transfer. High H 2 O 2 concentrations are involved in impaired melanosome transfer in vitiligo. However, low H 2 O 2 concentration promotes the beneficial proliferation and migration of melanocytes. The aim of this study was to explore low H 2 O 2 and its mechanism in melanosome transfer, protease-activated receptor-2 (PAR-2) expression and calcium balance. Melanosomes were fluorescein-labeled for clear visualization of their transfer. The expression of protease-activated receptor-2 (PAR-2) in keratinocytes was determined by western blot analysis. Flow cytometry was employed to evaluate the effects of H 2 O 2 on calcium levels in keratinocytes. Fluorescence microscopy showed the upregulation of melanosome transfer into keratinocytes following 0.3 mM H 2 O 2 treatment in the co-cultures rather than in the untreated control groups, which was associated with higher expression of PAR-2 protein and increased calcium concentration. The addition of a PAR-2 antagonist inhibited the positive activity of H 2 O 2 and calcium flow in keratinocytes. When calcium flow was blocked by a calcium chelator, the addition of H 2 O 2 did not increase the PAR-2 expression level in keratinocytes, therefore, inhibiting dendrite formation and melanosome transfer. Low H 2 O 2 concentration promotes melanosome transfer with increased PAR-2 expression level and calcium concentration in keratinocytes. In addition, the interaction between melanocytes and keratinocytes is more beneficial to enhance calcium levels in keratinocytes which mediate melanin transfer. Moreover, low H 2 O 2 concentration promotes dendrite formation, in which extracellular calcium and Par-2 were involved.

The impact of diets with different magnesium contents on magnesium and calcium in serum and tissues of the rat.

PubMed

Zimmermann, P; Weiss, U; Classen, H G; Wendt, B; Epple, A; Zollner, H; Temmel, W; Weger, M; Porta, S

2000-07-14

The impact of three different magnesium diets (70, 1,000 and 9,000 ppm) on total, ionized and bound magnesium as well as ionized calcium in serum and total calcium and magnesium in femoral bone, skeletal muscle, heart and liver of male Sprague-Dawley rats was investigated. The percentage of ionized serum magnesium was unproportionally high in rats fed a low magnesium (70 ppm) diet. Femoral magnesium was correlated with ionized and total serum magnesium. In contrast, there was generally no correlation between total serum magnesium and the magnesium fractions in skeletal muscle, heart and liver. In rats fed the magnesium deficient diet, total cardiac concentration of magnesium was even significantly increased along with total calcium content, while there were no effects on total muscle and liver magnesium. Within the single groups, ionized serum calcium was never proportional to dietary magnesium, but in all three magnesium diet groups together, it was inversely correlated with dietary magnesium. Moreover, ionized serum calcium was inversely correlated with both ionized and total serum magnesium. In all 3 groups together, the concentrations of total calcium and magnesium in heart and skeletal muscle were correlated, within the single groups correlation existed only in the 1000 ppm group. Magnesium influx via calcium channels during low magnesium intake has been seen in non cardiac tissues [35,36], but nothing similar is known about non selective channels for divalent cations in the heart [33]. Thus, magnesium uptake by cardiac cells along with calcium seems to be possible, especially at low intracellular magnesium concentrations, but is still poorly investigated. We suggest that the calcium-antagonistic effect of magnesium is related to the turnover rate of magnesium rather than to its tissue concentrations.

Transcription factor CREB is involved in CaSR-mediated cytoskeleton gene expression.

PubMed

Huang, Shuaishuai; Ren, Yu; Wang, Ping; Li, Yanyuan; Wang, Xue; Zhuang, Haihui; Fang, Rong; Wang, Yuduo; Liu, Ningsheng; Hehir, Michael; Zhou, Jeff X

2015-03-01

Our previous studies illustrated that a steady increase of intracellular calcium concentration ([Ca2+]i) was important for maintaining microtubules (MTs) rearrangement in apoptotic cells. However, little is known about the effect of calcium sensing receptor (CaSR)-mediated increase in [Ca2+]i on cytoskeleton gene expression. We examined the impact of taxol or CaSR agonist/antagonist on the regulation of [Ca2+]i concentration, cytoskeleton arrangement, phosphorylated CREB and cytoskeleton gene expressions in HeLa cells with dominant negative plasmid of CREB (PM). This study demonstrated that Gdcl3 (a specific CaSR agonist) evoked a rapid increase of [Ca2+]i, formed a rigid bundle of MTs which surrounded the nucleus and decreased the cytoskeleton gene expressions in HeLa cells. These effects were rescued by addition of NPS2390 (a specific CaSR antagonist). Moreover, CaSR activity affected cytoskeleton gene expression through transcription factor CREB. Histoscores of pCREB immunoreactivity in tissues of cervical adenocarcinoma, renal clear cell carcinoma, and diffuse large B-cell lymphoma were markedly increased compared with non malignant tissue. These data demonstrate, for the first time, that CaSR-mediated increase in [Ca2+]i probably modulate cytoskeleton organization and gene expression via transcription factor. © 2014 Wiley Periodicals, Inc.

Acute haemodynamic effects of felodipine and verapamil in man, singly and with metoprolol.

PubMed

Rönn, O; Bengtsson, B; Edgar, B; Raner, S

1985-01-01

In a single-blind randomised study in 9 healthy men we compared the acute haemodynamic effects of the calcium antagonists felodipine and verapamil, singly and in combination with metoprolol. Three different cumulative intravenous doses of 0.25, 0.75 and 1.5 mg felodipine and of 2.0, 4.0 and 8.0 mg verapamil or placebo were given as constant infusions over 5 minutes on 3 occasions and were followed by intravenous metoprolol (15 mg). Felodipine caused a significant and dose-dependent decrease in the total peripheral resistance, and an increase in the forearm blood flow by 8, 48 and 163% with progressively increasing doses showing that the drug is a potent arteriolar vasodilator. A significant and dose-dependent increase in heart rate and a decrease in the pre-ejection period/left ventricular ejection time (PEP/LVET) ratio of up to 15% was also recorded, mainly reflecting a reflexogenic increase in the sympathetic tone. Total peripheral resistance, forearm blood flow, heart rate and the systolic time intervals were mainly unchanged after verapamil, whereas the PQ interval was prolonged. Metoprolol given after the 2 calcium antagonists caused a decrease in heart rate and blood flow and an increase in the total peripheral resistance and PEP/LVET ratio. The tolerability was good to all infusions.

Phosphorylation and calcium antagonistically tune myosin-binding protein C’s structure and function

PubMed Central

Previs, Michael J.; Mun, Ji Young; Michalek, Arthur J.; Previs, Samantha Beck; Gulick, James; Robbins, Jeffrey; Warshaw, David M.; Craig, Roger

2016-01-01

During each heartbeat, cardiac contractility results from calcium-activated sliding of actin thin filaments toward the centers of myosin thick filaments to shorten cellular length. Cardiac myosin-binding protein C (cMyBP-C) is a component of the thick filament that appears to tune these mechanochemical interactions by its N-terminal domains transiently interacting with actin and/or the myosin S2 domain, sensitizing thin filaments to calcium and governing maximal sliding velocity. Both functional mechanisms are potentially further tunable by phosphorylation of an intrinsically disordered, extensible region of cMyBP-C’s N terminus, the M-domain. Using atomic force spectroscopy, electron microscopy, and mutant protein expression, we demonstrate that phosphorylation reduced the M-domain’s extensibility and shifted the conformation of the N-terminal domain from an extended structure to a compact configuration. In combination with motility assay data, these structural effects of M-domain phosphorylation suggest a mechanism for diminishing the functional potency of individual cMyBP-C molecules. Interestingly, we found that calcium levels necessary to maximally activate the thin filament mitigated the structural effects of phosphorylation by increasing M-domain extensibility and shifting the phosphorylated N-terminal fragments back to the extended state, as if unphosphorylated. Functionally, the addition of calcium to the motility assays ablated the impact of phosphorylation on maximal sliding velocities, fully restoring cMyBP-C’s inhibitory capacity. We conclude that M-domain phosphorylation may have its greatest effect on tuning cMyBP-C’s calcium-sensitization of thin filaments at the low calcium levels between contractions. Importantly, calcium levels at the peak of contraction would allow cMyBP-C to remain a potent contractile modulator, regardless of cMyBP-C’s phosphorylation state. PMID:26908872

Amino Alcohol- (NPS-2143) and Quinazolinone-Derived Calcilytics (ATF936 and AXT914) Differentially Mitigate Excessive Signalling of Calcium-Sensing Receptor Mutants Causing Bartter Syndrome Type 5 and Autosomal Dominant Hypocalcemia

PubMed Central

Letz, Saskia; Haag, Christine; Schulze, Egbert; Frank-Raue, Karin; Raue, Friedhelm; Hofner, Benjamin; Mayr, Bernhard; Schöfl, Christof

2014-01-01

Introduction Activating calcium sensing receptor (CaSR) mutations cause autosomal dominant hypocalcemia (ADH) characterized by low serum calcium, inappropriately low PTH and relative hypercalciuria. Four activating CaSR mutations cause additional renal wasting of sodium, chloride and other salts, a condition called Bartter syndrome (BS) type 5. Until today there is no specific medical treatment for BS type 5 and ADH. We investigated the effects of different allosteric CaSR antagonists (calcilytics) on activating CaSR mutants. Methods All 4 known mutations causing BS type 5 and five ADH mutations were expressed in HEK 293T cells and receptor signalling was studied by measurement of intracellular free calcium in response to extracellular calcium ([Ca2+]o). To investigate the effect of calcilytics, cells were stimulated with 3 mM [Ca2+]o in the presence or absence of NPS-2143, ATF936 or AXT914. Results All BS type 5 and ADH mutants showed enhanced signalling activity to [Ca2+]o with left shifted dose response curves. In contrast to the amino alcohol NPS-2143, which was only partially effective, the quinazolinone calcilytics ATF936 and AXT914 significantly mitigated excessive cytosolic calcium signalling of all BS type 5 and ADH mutants studied. When these mutants were co-expressed with wild-type CaSR to approximate heterozygosity in patients, ATF936 and AXT914 were also effective on all mutants. Conclusion The calcilytics ATF936 and AXT914 are capable of attenuating enhanced cytosolic calcium signalling activity of CaSR mutations causing BS type 5 and ADH. Quinazolinone calcilytics might therefore offer a novel treatment option for patients with activating CaSR mutations. PMID:25506941

Calcium and the heat-shock response in the human monocytic line U-937.

PubMed

Kantengwa, S; Capponi, A M; Bonventre, J V; Polla, B S

1990-07-01

In the human monocytic line U-937, 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] increases cytosolic free calcium concentration ([Ca2+]i). 1,25-(OH)2D3 also enhances the synthesis of heat-shock proteins (HSPs) when U-937 cells are exposed to elevated temperatures. To establish whether these two effects of 1,25-(OH)2D3 are related, we examined the effects of calcium on the heat-shock (HS) response, as well as the influence of 1,25-(OH)2D3 on this system. The equilibrium dissociation constant (Kd) of the fluorescent probe used to measure [Ca2+]i, fura-2, at 37 and 45 degrees C was found to be 191 and 234 nM, respectively. Exposure of U-937 cells to 45 degrees C did not increase [Ca2+]i under conditions in which active efflux of the dye was prevented by the organic anion transport inhibitor probenecid (1 mM). In cells preincubated in calcium-free medium, with subsequent addition of 4 mM EGTA before HS, or exposed to the calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7), the increase in HSPs synthesis was not affected. Cell viability, assessed by [3H]thymidine uptake, was not different between cells exposed to HS in calcium-containing or calcium-free media. Moreover, the effects of 1,25-(OH)2D3 on the HS response were also observed in a calcium-depleted medium, indicating that the effects of 1,25-(OH)2D3 on HSP synthesis were not mediated by [Ca2+]i.

Discovery and Development of Calcium Channel Blockers

PubMed Central

Godfraind, Théophile

2017-01-01

In the mid 1960s, experimental work on molecules under screening as coronary dilators allowed the discovery of the mechanism of calcium entry blockade by drugs later named calcium channel blockers. This paper summarizes scientific research on these small molecules interacting directly with L-type voltage-operated calcium channels. It also reports on experimental approaches translated into understanding of their therapeutic actions. The importance of calcium in muscle contraction was discovered by Sidney Ringer who reported this fact in 1883. Interest in the intracellular role of calcium arose 60 years later out of Kamada (Japan) and Heibrunn (USA) experiments in the early 1940s. Studies on pharmacology of calcium function were initiated in the mid 1960s and their therapeutic applications globally occurred in the the 1980s. The first part of this report deals with basic pharmacology in the cardiovascular system particularly in isolated arteries. In the section entitled from calcium antagonists to calcium channel blockers, it is recalled that drugs of a series of diphenylpiperazines screened in vivo on coronary bed precontracted by angiotensin were initially named calcium antagonists on the basis of their effect in depolarized arteries contracted by calcium. Studies on arteries contracted by catecholamines showed that the vasorelaxation resulted from blockade of calcium entry. Radiochemical and electrophysiological studies performed with dihydropyridines allowed their cellular targets to be identified with L-type voltage-operated calcium channels. The modulated receptor theory helped the understanding of their variation in affinity dependent on arterial cell membrane potential and promoted the terminology calcium channel blocker (CCB) of which the various chemical families are introduced in the paper. In the section entitled tissue selectivity of CCBs, it is shown that characteristics of the drug, properties of the tissue, and of the stimuli are important factors of their action. The high sensitivity of hypertensive animals is explained by the partial depolarization of their arteries. It is noted that they are arteriolar dilators and that they cannot be simply considered as vasodilators. The second part of this report provides key information about clinical usefulness of CCBs. A section is devoted to the controversy on their safety closed by the Allhat trial (2002). Sections are dedicated to their effect in cardiac ischemia, in cardiac arrhythmias, in atherosclerosis, in hypertension, and its complications. CCBs appear as the most commonly used for the treatment of cardiovascular diseases. As far as hypertension is concerned, globally the prevalence in adults aged 25 years and over was around 40% in 2008. Usefulness of CCBs is discussed on the basis of large clinical trials. At therapeutic dosage, they reduce the elevated blood pressure of hypertensive patients but don't change blood pressure of normotensive subjects, as was observed in animals. Those active on both L- and T-type channels are efficient in nephropathy. Alteration of cognitive function is a complication of hypertension recognized nowadays as eventually leading to dementia. This question is discussed together with the efficacy of CCBs in cognitive pathology. In the section entitled beyond the cardiovascular system, CCBs actions in migraine, neuropathic pain, and subarachnoid hemorrhage are reported. The final conclusions refer to long-term effects discovered in experimental animals that have not yet been clearly reported as being important in human pharmacotherapy. PMID:28611661

[The antioxidant prevention of disorders in calcium ion metabolism under the action of glutamate on the synaptosomes of the rat cerebral cortex].

PubMed

Avrova, N F; Shestak, K I; Zakharova, I O; Sokolova, T V; Tiurina, Iu Iu; Tiurin, V A

1999-04-01

An increase of intracellular calcium ion concentration and of the 45Ca2+ entry, a decrease in Na+,K(+)-ATPase activity, and activation of Na+/Ca2+ exchange were shown to be initiated by glutamate in the rat brain cortex synaptosomes. These effects could be prevented with antagonists and blocking agents of the NMDA receptors. Pre-incubation of the synaptosomes with alpha-tocopherol, superoxide dismutase, and ganglioside GM1 was shown to normalise [45Ca2+], the rate of 45Ca2+ entry, and the activity of Na+,K(+)-ATPase in the synaptosomes. The data obtained suggest that calcium ions entering the brain cortex neurones via the NMDA receptors in presence of excessive glutamate, trigger activation of free radical reactions damaging the neurones in ischemia, cerebral lesions, and other pathological conditions.

Discovery of Dual ETA/ETB Receptor Antagonists from Traditional Chinese Herbs through in Silico and in Vitro Screening

PubMed Central

Wang, Xing; Zhang, Yuxin; Liu, Qing; Ai, Zhixin; Zhang, Yanling; Xiang, Yuhong; Qiao, Yanjiang

2016-01-01

Endothelin-1 receptors (ETAR and ETBR) act as a pivotal regulator in the biological effects of ET-1 and represent a potential drug target for the treatment of multiple cardiovascular diseases. The purpose of the study is to discover dual ETA/ETB receptor antagonists from traditional Chinese herbs. Ligand- and structure-based virtual screening was performed to screen an in-house database of traditional Chinese herbs, followed by a series of in vitro bioassay evaluation. Aristolochic acid A (AAA) was first confirmed to be a dual ETA/ETB receptor antagonist based intracellular calcium influx assay and impedance-based assay. Dose-response curves showed that AAA can block both ETAR and ETBR with IC50 of 7.91 and 7.40 μM, respectively. Target specificity and cytotoxicity bioassay proved that AAA is a selective dual ETA/ETB receptor antagonist and has no significant cytotoxicity on HEK293/ETAR and HEK293/ETBR cells within 24 h. It is a feasible and effective approach to discover bioactive compounds from traditional Chinese herbs using in silico screening combined with in vitro bioassay evaluation. The structural characteristic of AAA for its activity was especially interpreted, which could provide valuable reference for the further structural modification of AAA. PMID:26999111

Chronic production of angiotensin IV in the brain leads to hypertension that is reversible with an angiotensin II AT1 receptor antagonist.

PubMed

Lochard, Nadheige; Thibault, Gaétan; Silversides, David W; Touyz, Rhian M; Reudelhuber, Timothy L

2004-06-11

Angiotensin IV (Ang IV) is a metabolite of the potent vasoconstrictor angiotensin II (Ang II). Because specific binding sites for this peptide have been reported in numerous tissues including the brain, it has been suggested that a specific Ang IV receptor (AT4) might exist. Bolus injection of Ang IV in brain ventricles has been implicated in learning, memory, and localized vasodilatation. However, the functions of Ang IV in a physiological context are still unknown. In this study, we generated a transgenic (TG) mouse model that chronically releases Ang IV peptide specifically in the brain. TG mice were found to be hypertensive by the tail-cuff method as compared with control littermates. Treatment with the angiotensin-converting enzyme inhibitor captopril had no effect on blood pressure, but surprisingly treatment with the Ang II AT1 receptor antagonist candesartan normalized the blood pressure despite the fact that the levels of Ang IV in the brains of TG mice were only 4-fold elevated over the normal endogenous level of Ang peptides. Calcium mobilization assays performed on cultured CHO cells chronically transfected with the AT1 receptor confirm that low-dose Ang IV can mobilize calcium via the AT1 receptor only in the presence of Ang II, consistent with an allosteric mechanism. These results suggest that chronic elevation of Ang IV in the brain can induce hypertension that can be treated with angiotensin II AT1 receptor antagonists.

RGS2 is regulated by angiotensin II and functions as a negative feedback of aldosterone production in H295R human adrenocortical cells.

PubMed

Romero, Damian G; Plonczynski, Maria W; Gomez-Sanchez, Elise P; Yanes, Licy L; Gomez-Sanchez, Celso E

2006-08-01

Regulator of G protein signaling (RGS) proteins interact with Galpha-subunits of heterotrimeric G proteins, accelerating the rate of GTP hydrolysis and finalizing the intracellular signaling triggered by the G protein-coupled receptor-ligand interaction. Angiotensin (Ang) II interacts with its G protein-coupled receptor in zona glomerulosa adrenal cells and triggers a cascade of intracellular signals that regulates steroidogenesis and proliferation. We studied Ang II-mediated regulation of RGS2, the role of RGS2 in steroidogenesis, and the intracellular signal events involved in H295R human adrenal cells. We report that both H295R cells and human adrenal gland express RGS2 mRNA. In H295R cells, Ang II caused a rapid and transient increase in RGS2 mRNA levels quantified by real-time RT-PCR. Ang II effects were mimicked by calcium ionophore A23187 and blocked by calcium channel blocker nifedipine. Ang II effects also were blocked by calmodulin antagonists (W-7 and calmidazolium) and calcium/calmodulin-dependent kinase antagonist KN-93. RGS2 overexpression by retroviral infection in H295R cells caused a decrease in Ang II-stimulated aldosterone secretion but did not modify cortisol secretion. In reporter assays, RGS2 decreased Ang II-mediated aldosterone synthase up-regulation. These results suggest that Ang II up-regulates RGS2 mRNA by the calcium/calmodulin-dependent kinase pathway in H295R cells. RGS2 overexpression specifically decreases aldosterone secretion through a decrease in Ang II-mediated aldosterone synthase-induced expression. In conclusion, RGS2 expression is induced by Ang II to terminate the intracellular signaling cascade generated by Ang II. RGS2 alterations in expression levels or functionality could be implicated in deregulations of Ang II signaling and abnormal aldosterone secretion by the adrenal gland.

NMDA receptors in mouse anterior piriform cortex initialize early odor preference learning and L-type calcium channels engage for long-term memory.

PubMed

Mukherjee, Bandhan; Yuan, Qi

2016-10-14

The interactions of L-type calcium channels (LTCCs) and NMDA receptors (NMDARs) in memories are poorly understood. Here we investigated the specific roles of anterior piriform cortex (aPC) LTCCs and NMDARs in early odor preference memory in mice. Using calcium imaging in aPC slices, LTCC activation was shown to be dependent on NMDAR activation. Either D-APV (NMDAR antagonist) or nifedipine (LTCC antagonist) reduced somatic calcium transients in pyramidal cells evoked by lateral olfactory tract stimulation. However, nifedipine did not further reduce calcium in the presence of D-APV. In mice that underwent early odor preference training, blocking NMDARs in the aPC prevented short-term (3 hr) and long-term (24 hr) odor preference memory, and both memories were rescued when BayK-8644 (LTCC agonist) was co-infused. However, activating LTCCs in the absence of NMDARs resulted in loss of discrimination between the conditioned odor and a similar odor mixture at 3 hr. Elevated synaptic AMPAR expression at 3 hr was prevented by D-APV infusion but restored when LTCCs were directly activated, mirroring the behavioral outcomes. Blocking LTCCs prevented 24 hr memory and spared 3 hr memory. These results suggest that NMDARs mediate stimulus-specific encoding of odor memory while LTCCs mediate intracellular signaling leading to long-term memory.

Voltage Dependence of a Neuromodulator-Activated Ionic Current.

PubMed

Gray, Michael; Golowasch, Jorge

2016-01-01

The neuromodulatory inward current (IMI) generated by crab Cancer borealis stomatogastric ganglion neurons is an inward current whose voltage dependence has been shown to be crucial in the activation of oscillatory activity of the pyloric network of this system. It has been previously shown that IMI loses its voltage dependence in conditions of low extracellular calcium, but that this effect appears to be regulated by intracellular calmodulin. Voltage dependence is only rarely regulated by intracellular signaling mechanisms. Here we address the hypothesis that the voltage dependence of IMI is mediated by intracellular signaling pathways activated by extracellular calcium. We demonstrate that calmodulin inhibitors and a ryanodine antagonist can reduce IMI voltage dependence in normal Ca(2+), but that, in conditions of low Ca(2+), calmodulin activators do not restore IMI voltage dependence. Further, we show evidence that CaMKII alters IMI voltage dependence. These results suggest that calmodulin is necessary but not sufficient for IMI voltage dependence. We therefore hypothesize that the Ca(2+)/calmodulin requirement for IMI voltage dependence is due to an active sensing of extracellular calcium by a GPCR family calcium-sensing receptor (CaSR) and that the reduction in IMI voltage dependence by a calmodulin inhibitor is due to CaSR endocytosis. Supporting this, preincubation with an endocytosis inhibitor prevented W7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride)-induced loss of IMI voltage dependence, and a CaSR antagonist reduced IMI voltage dependence. Additionally, myosin light chain kinase, which is known to act downstream of the CaSR, seems to play a role in regulating IMI voltage dependence. Finally, a Gβγ-subunit inhibitor also affects IMI voltage dependence, in support of the hypothesis that this process is regulated by a G-protein-coupled CaSR.

Voltage Dependence of a Neuromodulator-Activated Ionic Current123

PubMed Central

2016-01-01

Abstract The neuromodulatory inward current (IMI) generated by crab Cancer borealis stomatogastric ganglion neurons is an inward current whose voltage dependence has been shown to be crucial in the activation of oscillatory activity of the pyloric network of this system. It has been previously shown that IMI loses its voltage dependence in conditions of low extracellular calcium, but that this effect appears to be regulated by intracellular calmodulin. Voltage dependence is only rarely regulated by intracellular signaling mechanisms. Here we address the hypothesis that the voltage dependence of IMI is mediated by intracellular signaling pathways activated by extracellular calcium. We demonstrate that calmodulin inhibitors and a ryanodine antagonist can reduce IMI voltage dependence in normal Ca2+, but that, in conditions of low Ca2+, calmodulin activators do not restore IMI voltage dependence. Further, we show evidence that CaMKII alters IMI voltage dependence. These results suggest that calmodulin is necessary but not sufficient for IMI voltage dependence. We therefore hypothesize that the Ca2+/calmodulin requirement for IMI voltage dependence is due to an active sensing of extracellular calcium by a GPCR family calcium-sensing receptor (CaSR) and that the reduction in IMI voltage dependence by a calmodulin inhibitor is due to CaSR endocytosis. Supporting this, preincubation with an endocytosis inhibitor prevented W7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride)-induced loss of IMI voltage dependence, and a CaSR antagonist reduced IMI voltage dependence. Additionally, myosin light chain kinase, which is known to act downstream of the CaSR, seems to play a role in regulating IMI voltage dependence. Finally, a Gβγ-subunit inhibitor also affects IMI voltage dependence, in support of the hypothesis that this process is regulated by a G-protein-coupled CaSR. PMID:27257619

The Role of Ca2+ Imbalance in the Induction of Acute Oxidative Stress and Cytotoxicity in Cultured Rat Cerebellar Granule Cells Challenged with Tetrabromobisphenol A.

PubMed

Zieminska, Elzbieta; Lenart, Jacek; Diamandakis, Dominik; Lazarewicz, Jerzy W

2017-03-01

Using primary cultures of rat cerebellar granule cells (CGC) we examined the role of calcium transients induced by tetrabromobisphenol A (TBBPA) in triggering oxidative stress and cytotoxicity. CGC were exposed for 30 min to 10 or 25 µM TBBPA. Changes in intracellular calcium concentration ([Ca 2+ ] i ), in the production of reactive oxygen species (ROS), and in the potential of mitochondria (∆Ψm) were measured fluorometrically during the exposure. The intracellular glutathione (GSH) and catalase activity were determined after the incubation; cell viability was evaluated 24 h later. TBBPA concentration-dependently increased [Ca 2+ ] i and ROS production, and reduced GSH content, catalase activity, ∆Ψm and neuronal viability. The combination of NMDA and ryanodine receptor antagonists, MK-801 and bastadin 12 with ryanodine, respectively, prevented Ca 2+ transients and partially reduced cytotoxicity induced by TBBPA at both concentrations. The antagonists also completely inhibited oxidative stress and depolarization of mitochondria evoked by 10 µM TBBPA, whereas these effects were only partially reduced in the 25 µM TBBPA treatment. Free radical scavengers prevented TBBPA-induced development of oxidative stress and improved CGC viability without having any effect on the rises in Ca 2+ and drop in ∆Ψm. The co-administration of scavengers with NMDA and ryanodine receptor antagonists provided almost complete neuroprotection. These results indicate that Ca 2+ imbalance and oxidative stress both mediate acute toxicity of TBBPA in CGC. At 10 µM TBBPA Ca 2+ imbalance is a primary event, inducing oxidative stress, depolarization of mitochondria and cytotoxicity, whilst at a concentration of 25 µM TBBPA an additional Ca 2+ -independent portion of oxidative stress and cytotoxicity emerges.

Growth differentiation factor-15 promotes glutamate release in medial prefrontal cortex of mice through upregulation of T-type calcium channels.

PubMed

Liu, Dong-Dong; Lu, Jun-Mei; Zhao, Qian-Ru; Hu, Changlong; Mei, Yan-Ai

2016-06-29

Growth differentiation factor-15 (GDF-15) has been implicated in ischemic brain injury and synapse development, but its involvement in modulating neuronal excitability and synaptic transmission remain poorly understood. In this study, we investigated the effects of GDF-15 on non-evoked miniature excitatory post-synaptic currents (mEPSCs) and neurotransmitter release in the medial prefrontal cortex (mPFC) in mice. Incubation of mPFC slices with GDF-15 for 60 min significantly increased the frequency of mEPSCs without effect on their amplitude. GDF-15 also significantly elevated presynaptic glutamate release, as shown by HPLC. These effects were blocked by dual TGF-β type I receptor (TβRI) and TGF-β type II receptor (TβRII) antagonists, but not by a TβRI antagonist alone. Meanwhile, GDF-15 enhanced pERK level, and inhibition of MAPK/ERK activity attenuated the GDF-15-induced increases in mEPSC and glutamate release. Blocking T-type calcium channels reduced the GDF-15 induced up-regulation of synaptic transmission. Membrane-protein extraction and use of an intracellular protein-transport inhibitor showed that GDF-15 promoted CaV3.1 and CaV3.3 α-subunit expression by trafficking to the membrane. These results confirm previous findings in cerebellar granule neurons, in which GDF-15 induces its neurobiological effects via TβRII and activation of the ERK pathway, providing novel insights into the mechanism of GDF-15 function in cortical neurons.

Intravenous strontium gluconate as a kinetic marker for calcium in healthy volunteers.

PubMed Central

Moraes, M E; Aronson, J K; Grahame-Smith, D G

1991-01-01

1. We have studied the pharmacokinetics of stable strontium in 10 healthy male volunteers. We gave each volunteer 5 mmol strontium gluconate by intravenous infusion over 1 h and measured strontium concentrations in plasma and urine samples for 20 days. The plasma strontium concentration vs time data for each volunteer were fitted by a triexponential function using NONLIN. Compartmental model-dependent and model-independent pharmacokinetic variables were then calculated. 2. The mean half-life we report (5.4 days) is longer than that previously reported (about 2 days), since we continued sampling for 20 days. However, the rates of clearance (CL 9.4 ml min-1. CLR 5.4 ml min-1, and CLNR 4.0 ml min-1) are similar to those previously reported, and the apparent volume of distribution at steady state (64 l) is similar to the values previously reported for the size of the exchangeable pool of both strontium and calcium. 3. The similarities in the pharmacokinetic behavior of strontium and calcium suggest that the in vivo disposition of strontium may be used as a marker of calcium disposition and for studying the effects of drugs such as the calcium antagonists. PMID:2049251

Butylidenephthalide blocks potassium channels and enhances basal tension in isolated guinea-pig trachea.

PubMed

Hsu, Hsin-Te; Yang, You-Lan; Chen, Wan-Chen; Chen, Chi-Ming; Ko, Wun-Chang

2014-01-01

Butylidenephthalide (Bdph, 30~300 μM), a constituent of Ligusticum chuanxiong Hort., significantly enhanced tension in isolated guinea-pig trachea. In this study, we investigate the mechanism(s) of Bdph-induced contraction in the tissue. Isolated trachea was bathed in 5 mL of Krebs solution containing indomethacin (3 μM), and its tension changes were isometrically recorded. Cromakalim (3 μM), an ATP-dependent K+ channel opener, significantly antagonized the Bdph-induced enhancement of baseline tension. Bdph (300 μM) also significantly antagonized cromakalim-induced relaxation. Bdph (300 μM) did not significantly influence the antagonistic effects of glibenclamide (GBC, 1 μM) and tetraethylammonium (TEA, 8 mM) against the cromakalim-induced relaxation. However, Bdph (300 μM) and 4-aminopiridine (4-AP, 5 mM), a blocker of K v 1 family of K+ channels, in combination significantly rightward shifted the log concentration-relaxation curve of cromakalim. The antagonistic effect of the combination almost equals the sum of the individual effects of Bdph and 4-AP, suggesting that the antagonistic mechanism of Bdph may be similar to that of 4-AP. All calcium channel blockers influenced neither the baseline tension nor antagonistic effect of Bdph against cromakalim. In conclusion, Bdph may be similar to 4-AP, a blocker of K v 1 family of K+ channels, to enhance the baseline tension of guinea-pig trachea.

Novel mode of action of the calcium antagonist mibefradil (Ro 40-5967): potent immunosuppression by inhibition of T-cell infiltration through allogeneic endothelium.

PubMed Central

Blaheta, R A; Hailer, N P; Brude, N; Wittig, B; Oppermann, E; Leckel, K; Harder, S; Scholz, M; Weber, S; Encke, A; Markus, B H

1998-01-01

Cyclosporin A reduces the mitotic activity of allosensitized lymphocytes, but fails to limit emigration of these cells into the donor organ. However, the modulation of both lymphocyte proliferation and infiltration are desirable characteristics of immunosuppressive therapy. The calcium-channel blocker, verapamil, has recently been shown to effectively prevent the transmigration of CD4+ and CD8+ T cells through allogeneic endothelium. Mibefradil (Ro 40-5967) represents a new generation of calcium antagonists with high potency and long-term activity. To evaluate the immunosuppressive potential of this drug, the influence of mibefradil on lymphocyte adhesion to, horizontal locomotion along, and penetration through allogeneic endothelium (HUVEC) was performed. When lymphocytes were prestimulated for 24 hr with mibefradil, adhesion and penetration were dose-dependently reduced. The adhesion ID50 values were 3.4 microM (CD4+ T cells) versus 9.2 microM (CD8+ T cells) and 2.1 microM (CD4+ T cells) versus 3.9 microM (CD8+ T cells) with regard to penetration. Mibefradil also effectively blocked horizontal locomotion. Specific down-regulation of T-cell binding to the P-selection receptor (ID50: CD4+ T cells, 0.8 microM: CD8+ T cells, 1.2 microM) and to the intracellular adhesion molecule-1 (ICAM-1) receptor (ID50: CD4+ T cells, 1.9 microM; CD8+ T cells, 1.5 microM) by mibefradil seems to be responsible for the decreased adhesion and penetration rates. Reduction of intracellular F-actin in T lymphocytes could diminish cell locomotion. In conclusion, the potent suppressive properties of mibefradil support its use as a co-medication in cyclosporin A-based immunosuppressive therapy. PMID:9741343

Novel mode of action of the calcium antagonist mibefradil (Ro 40-5967): potent immunosuppression by inhibition of T-cell infiltration through allogeneic endothelium.

PubMed

Blaheta, R A; Hailer, N P; Brude, N; Wittig, B; Oppermann, E; Leckel, K; Harder, S; Scholz, M; Weber, S; Encke, A; Markus, B H

1998-06-01

Cyclosporin A reduces the mitotic activity of allosensitized lymphocytes, but fails to limit emigration of these cells into the donor organ. However, the modulation of both lymphocyte proliferation and infiltration are desirable characteristics of immunosuppressive therapy. The calcium-channel blocker, verapamil, has recently been shown to effectively prevent the transmigration of CD4+ and CD8+ T cells through allogeneic endothelium. Mibefradil (Ro 40-5967) represents a new generation of calcium antagonists with high potency and long-term activity. To evaluate the immunosuppressive potential of this drug, the influence of mibefradil on lymphocyte adhesion to, horizontal locomotion along, and penetration through allogeneic endothelium (HUVEC) was performed. When lymphocytes were prestimulated for 24 hr with mibefradil, adhesion and penetration were dose-dependently reduced. The adhesion ID50 values were 3.4 microM (CD4+ T cells) versus 9.2 microM (CD8+ T cells) and 2.1 microM (CD4+ T cells) versus 3.9 microM (CD8+ T cells) with regard to penetration. Mibefradil also effectively blocked horizontal locomotion. Specific down-regulation of T-cell binding to the P-selection receptor (ID50: CD4+ T cells, 0.8 microM: CD8+ T cells, 1.2 microM) and to the intracellular adhesion molecule-1 (ICAM-1) receptor (ID50: CD4+ T cells, 1.9 microM; CD8+ T cells, 1.5 microM) by mibefradil seems to be responsible for the decreased adhesion and penetration rates. Reduction of intracellular F-actin in T lymphocytes could diminish cell locomotion. In conclusion, the potent suppressive properties of mibefradil support its use as a co-medication in cyclosporin A-based immunosuppressive therapy.

High-frequency voltage oscillations in cultured astrocytes

PubMed Central

Fleischer, Wiebke; Theiss, Stephan; Slotta, Johannes; Holland, Christine; Schnitzler, Alfons

2015-01-01

Because of their close interaction with neuronal physiology, astrocytes can modulate brain function in multiple ways. Here, we demonstrate a yet unknown astrocytic phenomenon: Astrocytes cultured on microelectrode arrays (MEAs) exhibited extracellular voltage fluctuations in a broad frequency spectrum (100–600 Hz) after electrical stimulation. These aperiodic high-frequency oscillations (HFOs) could last several seconds and did not spread across the MEA. The voltage-gated calcium channel antagonist cilnidipine dose-dependently decreased the power of the oscillations. While intracellular calcium was pivotal, incubation with bafilomycin A1 showed that vesicular release of transmitters played only a minor role in the emergence of HFOs. Gap junctions and volume-regulated anionic channels had just as little functional impact, which was demonstrated by the addition of carbenoxolone (100 μmol/L) and NPPB (100 μmol/L). Hyperpolarization with low potassium in the extracellular solution (2 mmol/L) dramatically raised oscillation power. A similar effect was seen when we added extra sodium (+50 mmol/L) or if we replaced it with NMDG+ (50 mmol/L). The purinergic receptor antagonist PPADS suppressed the oscillation power, while the agonist ATP (100 μmol/L) had only an increasing effect when the bath solution pH was slightly lowered to pH 7.2. From these observations, we conclude that astrocytic voltage oscillations are triggered by activation of voltage-gated calcium channels and driven by a downstream influx of cations through channels that are permeable for large ions such as NMDG+. Most likely candidates are subtypes of pore-forming P2X channels with a low affinity for ATP. PMID:25969464

Allosteric Effects of the Anti-Psychotic Drug Trifluoperazine on the Energetics of Calcium Binding by Calmodulin

PubMed Central

Feldkamp, Michael D.; O'Donnell, Susan E.; Yu, Liping; Shea, Madeline A.

2010-01-01

Trifluoperazine (TFP; Stelazine™) is an antagonist of calmodulin (CaM), an essential regulator of calcium-dependent signal transduction. Reports differ regarding whether, or where, TFP binds to apo CaM. Three crystallographic structures (1CTR, 1A29, 1LIN) show TFP bound to (Ca2+)4-CaM in ratios of 1, 2 or 4 TFP per CaM. In all of these, CaM domains adopt the “open” conformation seen in CaM-kinase complexes having increased calcium affinity. Most reports suggest TFP also increases calcium affinity of CaM. To compare TFP binding to apo CaM and (Ca2+)4-CaM, and explore differential effects on the N- and C-domains of CaM, stoichiometric TFP titrations of CaM were monitored by 15N-HSQC NMR. Two TFP bound to apo CaM, while four bound to (Ca2+)4-CaM. In both cases, the preferred site was in the C-domain. During the titrations, biphasic responses for some resonances suggested inter-site interactions. TFP-binding sites in apo CaM appeared distinct from those in (Ca2+)4-CaM. In equilibrium calcium titrations at defined ratios of TFP:CaM, TFP reduced calcium affinity at most levels tested; this is similar to the effect of many IQ-motifs on CaM. However, at the highest level tested, TFP raised the calcium affinity of the N-domain of CaM. A model of conformational switching is proposed to explain how TFP can exert opposing allosteric effects on calcium affinity by binding to different sites in the “closed”, “semi-open” and “open” domains of CaM. In physiological processes, apo CaM, as well as (Ca2+)4-CaM, needs to be considered a potential target of drug action. PMID:20544963

Calcium-dependent mechanisms mediate the vasorelaxant effects of Tridax procumbens (Lin) aqueous leaf extract in rat aortic ring.

PubMed

Salahdeen, Hussein M; Idowu, Gbolahan O; Yemitan, Omoniyi K; Murtala, Babatunde A; Alada, Abdul-Rasak A

2014-05-01

Tridax procumbens leaf extract has a folk reputation as an antihypertensive agent in Nigeria. Evidence suggests that it has a relaxant effect on smooth muscles. The present study was designed to investigate the role of calcium in the vasorelaxant effect of this extract. Concentration-response studies with noradrenaline (NA), KCl and CaCl2 were carried out in rat aortic rings with and without the extract in physiological salt solution (PSS) (n=6 each). Also, the role of intracellular calcium mobilization was studied by measuring the phasic response to NA in Ca2+-free N,N-ethylene glycol tetraacetic acid (EGTA) PSS (n=6). The results showed that the contractile responses to either NA or KCl were attenuated (p<0.05) in the presence of the extract. Also, the extract attenuated the contractile response to CaCl2 in the presence of NA or KCl (p<0.05) in the Ca2+-free EGTA PSS, while the phasic response to NA was significantly (p<0.05) diminished. These results suggest that the vasorelaxant effect of T. procumbens leaf extract may be mediated by a non-specific, non-competitive inhibition of Ca2+ influx as well as by inhibition of Ca2+ mobilization from intracellular stores. This implies that it may contain vasorelaxant agents that may have calcium antagonistic potential.

Calcium Channel Antagonists as Disease-Modifying Therapy for Parkinson's Disease: Therapeutic Rationale and Current Status.

PubMed

Swart, Tara; Hurley, Michael J

2016-12-01

Parkinson's disease is a disabling hypokinetic neurological movement disorder in which the aetiology is unknown in the majority of cases. Current pharmacological treatments, though effective at restoring movement, are only symptomatic and do nothing to slow disease progression. Electrophysiological, epidemiological and neuropathological studies have implicated Ca V 1.3 subtype calcium channels in the pathogenesis of the disorder, and drugs with some selectivity for this ion channel (brain-penetrant dihydropyridine calcium channel blockers) are neuroprotective in animal models of the disease. Dihydropyridines have been safely used for decades to treat hypertension and other cardiovascular disorders. A phase II clinical trial found that isradipine was safely tolerated by patients with Parkinson's disease, and a phase III trial is currently underway to determine whether treatment with isradipine is neuroprotective and therefore able to slow the progression of Parkinson's disease. This manuscript reviews the current information about the use of dihydropyridines as therapy for Parkinson's disease and discusses the possible mechanism of action of these drugs, highlighting Ca V 1.3 calcium channels as a potential therapeutic target for neuroprotection in Parkinson's disease.

Role of the JAKs/STATs pathway in the intracellular calcium changes induced by interleukin-6 in hippocampal neurons.

PubMed

Orellana, D I; Quintanilla, R A; Gonzalez-Billault, C; Maccioni, R B

2005-11-01

Recent studies show that inflammation has an active role in the onset of neurodegenerative diseases. It is known that in response to extracellular insults microglia and/or astrocytes produce inflammatory agents. These contribute to the neuropathological events in the aging process and neuronal degeneration. Interleukin-6 (IL-6) has been involved in the pathogenesis of neurodegenerative disorders, such as Alzheimer's and Parkinson's diseases. Here, we show that IL-6 treatment of rat hippocampal neurons increases the calcium influx via NMDA-receptor, an effect that is prevented by the specific NMDA receptor antagonist MK-801 (dizocilpine). We also show that this calcium influx is mediated by the JAKs/STATs pathway, since the inhibitor of JAKs/STATs pathway, JAK 3 inhibitor, blocks calcium influx even in the presence of IL-6. This increase in calcium signal was dependent on external sources, since this signal was not observed in the presence of EGTA. Additional studies indicate that the increase in cytosolic calcium induces tau protein hyperphosphorylation, as revealed by using specific antibodies against Alzheimer phosphoepitopes. This anomalous tau hyperphosphorylation was dependent on both the JAKs/STATs pathway and NMDA receptor. These results suggest that IL-6 would induce a cascade of molecular events that produce a calcium influx through NMDA receptors, mediated by the JAKs/STATs pathway, which subsequently modifies the tau hyperphosphorylation patterns.

Mechanisms of calcium sequestration by isolated Malpighian tubules of the house cricket Acheta domesticus.

PubMed

Browne, Austin; O'Donnell, Michael J

2018-01-01

Hemolymph calcium homeostasis in insects is achieved by the Malpighian tubules, primarily by sequestering excess Ca 2+ within internal calcium stores (Ca-rich granules) most often located within type I (principal) tubule cells. Using both the scanning ion-selective electrode technique and the Ramsay secretion assay this study provides the first measurements of basolateral and transepithelial Ca 2+ fluxes across the Malpighian tubules of an Orthopteran insect, the house cricket Acheta domesticus. Ca 2+ transport was specific to midtubule segments, where 97% of the Ca 2+ entering the tubule is sequestered within intracellular calcium stores and the remaining 3% is secreted into the lumen. Antagonists of voltage-gated (L-type) calcium channels decreased Ca 2+ influx ≥fivefold in adenosine 3',5'-cyclic monophosphate (cAMP)-stimulated tubules, suggesting basolateral Ca 2+ influx is facilitated by voltage-gated Ca 2+ channels. Increasing fluid secretion through manipulation of intracellular levels of cAMP or Ca 2+ had opposite effects on tubule Ca 2+ transport. The adenylyl cyclase-cAMP-PKA pathway promotes Ca 2+ sequestration whereas both 5-hydroxytryptamine and thapsigargin inhibited sequestration. Our results suggest that the midtubules of Acheta domesticus are dynamic calcium stores, which maintain hemolymph calcium concentration by manipulating rates of Ca 2+ sequestration through stimulatory (cAMP) and inhibitory (Ca 2+ ) regulatory pathways. © 2017 Wiley Periodicals, Inc.

Prolonged Attenuation of Amygdala-Kindled Seizure Measures in Rats by Convection-Enhanced Delivery of the N-Type Calcium Channel Antagonists ω-Conotoxin GVIA and ω-Conotoxin MVIIA

PubMed Central

Gasior, Maciej; White, Natalie A.; Rogawski, Michael A.

2008-01-01

Convection-enhanced delivery (CED) permits the homogeneous distribution of therapeutic agents throughout localized regions of the brain parenchyma without causing tissue damage as occurs with bolus injection. Here, we examined whether CED infusion of the N-type calcium channel antagonists ω-conotoxin GVIA (ω-CTX-G) and ω-conotoxin MVIIA (ω-CTX-M) can attenuate kindling measures in fully amygdala-kindled rats. Rats were implanted with a combination infusion cannula-stimulating electrode assembly into the right basolateral amygdala. Fully kindled animals received infusions of vehicle, ω-CTX-G (0.005, 0.05, and 0.5 nmol), ω-CTX-M (0.05, 0.15, and 0.5 nmol), proteolytically inactivated ω-CTX-M (0.5 nmol), or carbamazepine (500 nmol) into the stimulation site. CED of ω-CTX-G and ω-CTX-M over a 20-min period resulted in a dose-dependent increase in the afterdischarge threshold and a decrease in the afterdischarge duration and behavioral seizure score and duration during a period of 20 min to 1 week after the infusion, indicating an inhibitory effect on the triggering and expression of kindled seizures. The protective effects of ω-conotoxins reached a maximum at 48 h postinfusion, and then they gradually resolved over the next 5 days. In contrast, carbamazepine was active at 20 min but not at 24 h after the infusion, whereas CED of vehicle or inactivated ω-CTX-M had no effect. Except for transient tremor in some rats receiving the highest toxin doses, no adverse effects were observed. These results indicate that local CED of high-molecular-weight presynaptic N-type calcium channel blockers can produce long-lasting inhibition of brain excitability and that they may provide prolonged seizure protection in focal seizure disorders. PMID:17717191

The common inhaled anesthetic isoflurane increases aggregation of huntingtin and alters calcium homeostasis in a cell model of Huntington's disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang Qiujun; Department of Anesthesiology, The Third Clinical Hospital, Hebei Medical University, Shijiazhuang, Hebei 050051; Liang Ge

2011-02-01

Isoflurane is known to increase {beta}-amyloid aggregation and neuronal damage. We hypothesized that isoflurane will have similar effects on the polyglutamine huntingtin protein and will cause alterations in intracellular calcium homeostasis. We tested this hypothesis in striatal cells from the expanded glutamine huntingtin knock-in mouse (STHdh{sup Q111/Q111}) and wild type (STHdh{sup Q7/Q7}) striatal neurons. The primary cultured neurons were exposed for 24 h to equipotent concentrations of isoflurane, sevoflurane, and desflurane in the presence or absence of extracellular calcium and with or without xestospongin C, a potent endoplasmic reticulum inositol 1,4,5-trisphosphate (InsP{sub 3}) receptor antagonist. Aggregation of huntingtin protein, cellmore » viability, and calcium concentrations were measured. Isoflurane, sevoflurane, and desflurane all increased the aggregation of huntingtin in STHdh{sup Q111/Q111} cells, with isoflurane having the largest effect. Isoflurane induced greater calcium release from the ER and relatively more cell damage in the STHdh{sup Q111/Q111} huntingtin cells than in the wild type STHdh{sup Q7/Q7} striatal cells. However, sevoflurane and desflurane caused less calcium release from the ER and less cell damage. Xestospongin C inhibited the isoflurane-induced calcium release from the ER, aggregation of huntingtin, and cell damage in the STHdh{sup Q111/Q111} cells. In summary, the Q111 form of huntingtin increases the vulnerability of striatal neurons to isoflurane neurotoxicity through combined actions on the ER IP{sub 3} receptors. Calcium release from the ER contributes to the anesthetic induced huntingtin aggregation in STHdh{sup Q111/Q111} striatal cells.« less

Endothelinergic Contractile Hyperreactivity in Rat Contralateral Carotid to Balloon Injury: Integrated Role for ETB Receptors and Superoxide Anion

PubMed Central

Gimenes, Lilian R.; Gomes, Mayara S.; do Vale, Bruno N.; Cardoso, Cristina R. B.; de Oliveira, Ana M.; Moreira, Josimar D.

2017-01-01

Temporal consequences of neurocompensation to balloon injury on endothelinergic functionality in rat contralateral carotid were evaluated. Rats underwent balloon injury in left carotid and were treated with CP-96345 (NK1 antagonist). Concentration-response curves for endothelin-1 were obtained in contralateral (right) carotid at 2, 8, 16, 30, or 45 days after surgery in the absence or presence of BQ-123 (ETA antagonist), BQ-788 (ETB antagonist), or Tempol (superoxide-dismutase mimic). Endothelin-1-induced calcium mobilization was evaluated in functional assays carried out with BQ-123, BQ-788, or Tempol. Endothelin-1-induced NADPH oxidase-driven superoxide generation was measured by lucigenin chemiluminescence assays performed with BQ-123 or BQ-788. Endothelin-1-induced contraction was increased in contralateral carotid from the sixteenth day after surgery. This response was restored in CP-96345-treated rats. Endothelium removal or BQ-123 did not change endothelin-1-induced contraction in contralateral carotid. This response was restored by BQ-788 or Tempol. Contralateral carotid exhibited an increased endothelin-1-induced calcium mobilization, which was restored by BQ-788 or Tempol. Contralateral carotid exhibited an increased endothelin-1-induced lucigenin chemiluminescence, which was restored by BQ-788. We conclude that the NK1-mediated neurocompensatory response to balloon injury elicits a contractile hyperreactivity to endothelin-1 in rat contralateral carotid by enhancing the muscular ETB-mediated NADPH oxidase-driven generation of superoxide, which activates calcium channels. PMID:29062837

Characterization of Beta-leptinotarsin-h and the Effects of Calcium Flux Antagonists on its Activity

DTIC Science & Technology

2005-04-07

A alone. a IP3R, IP3 receptor ; LO, ligand -operated; RyR, ryanodine receptor ; SERCA, sarcoplasmic reticulum endoplasmic reticulum Ca 2C ATPase; SO...observation eliminated non-selective cation channels such as nicotinic, glutamatergic, purinergic P2X , and serotoni- nergic 5-HT3 ligand -operated Ca 2C...nicardipine, nifedipine, SNX-482) was inhibitory. Selective inhibitors of ligand -operated, store-operated, and transduction-operated channels were also not

Distinct action of aranidipine and its active metabolite on renal arterioles, with special reference to renal protection.

PubMed

Nakamura, A; Hayashi, K; Fujiwara, K; Ozawa, Y; Honda, M; Saruta, T

2000-06-01

Aranidipine, a newly developed calcium antagonist, possesses unique pharmacologic characteristics in that its metabolite (M-1) still has antihypertensive action. We examined the effects of both agents on renal microcirculation using the isolated perfused hydronephrotic rat kidney. During norepinephrine-induced constriction, the addition of aranidipine dilated both afferent and efferent arterioles in a dose-dependent manner; at 10(-6) M, 83 +/- 6% and 90 +/- 6% reversal, respectively. In contrast, its active metabolite exerted dilator action predominantly on the afferent arteriole (79 +/- 4% vs. 44 +/- 17% at 10(-6) M for afferent and efferent arterioles, respectively). We further examined the long-term (8 weeks) effect of these agents on the development of renal injury in salt-loaded subtotally nephrectomized spontaneously hypertensive rats. Both aranidipine and M-1 reduced blood pressure by a similar magnitude. The decreases in proteinuria were observed in the aranidipine-treated group at weeks 6, 8, and 10, whereas in the M-1 group, significant reduction was attained only at week 6. Histopathologic examination revealed that both treatments improved glomerular and arteriolar sclerosis. Glomerular sclerosis, however, was less pronounced in the aranidipine-treated group than in the M-1 group. In conclusion, aranidipine has dilator action on both arterioles, whereas M-1 caused predominant dilation of afferent arterioles. Such metabolic changes may constitute a determinant of efferent arteriolar action of the calcium antagonist.

Inhibition of NMDARs in the Nucleus Reticularis of the Thalamus Produces Delta Frequency Bursting.

PubMed

Zhang, Yuchun; Llinas, Rodolfo R; Lisman, John E

2009-01-01

Injection of NMDAR antagonist into the thalamus can produce delta frequency EEG oscillations in the thalamocortical system. It is surprising that an antagonist of an excitatory neurotransmitter should trigger such activity, and the mechanism is unknown. One hypothesis is that the antagonist blocks excitation of GABAergic cells, thus producing disinhibition. To test this hypothesis, we investigated the effect of NMDAR antagonist (APV) on cells of the nucleus reticularis (nRT) in rat brain slices, a thalamic nucleus that can serve as a pacemaker for thalamocortical delta oscillations and that is composed entirely of GABAergic neurons. We found, unexpectedly, that nRT cells are hyperpolarized by APV. This occurs because these cells have an unusual form of NMDAR (probably NR2C) that contributes inward current at resting potential in response to ambient glutamate. The hyperpolarization produced by APV is sufficient to deinactivate T-type calcium channels, and these trigger rhythmic bursting at delta frequency. The APV-induced delta frequency bursting is abolished by dopamine D2 receptor antagonist, indicating that dopamine and NMDAR antagonist work synergistically to stimulate delta frequency bursting. Our results have significant implications concerning the electrophysiological basis of schizophrenia and bring together the NMDAR hypofunction, dopamine, and GABA theories of the disease. Our results suggest that NMDAR hypofunction and dopamine work synergistically on the GABAergic cells of the nRT to generate the delta frequency EEG oscillations, a thalamocortical dysrhythmia (TCD) in the awake state that is an established abnormality in schizophrenia.

Inhibition of NMDARs in the Nucleus Reticularis of the Thalamus Produces Delta Frequency Bursting

PubMed Central

Zhang, Yuchun; Llinas, Rodolfo R.; Lisman, John E.

2009-01-01

Injection of NMDAR antagonist into the thalamus can produce delta frequency EEG oscillations in the thalamocortical system. It is surprising that an antagonist of an excitatory neurotransmitter should trigger such activity, and the mechanism is unknown. One hypothesis is that the antagonist blocks excitation of GABAergic cells, thus producing disinhibition. To test this hypothesis, we investigated the effect of NMDAR antagonist (APV) on cells of the nucleus reticularis (nRT) in rat brain slices, a thalamic nucleus that can serve as a pacemaker for thalamocortical delta oscillations and that is composed entirely of GABAergic neurons. We found, unexpectedly, that nRT cells are hyperpolarized by APV. This occurs because these cells have an unusual form of NMDAR (probably NR2C) that contributes inward current at resting potential in response to ambient glutamate. The hyperpolarization produced by APV is sufficient to deinactivate T-type calcium channels, and these trigger rhythmic bursting at delta frequency. The APV-induced delta frequency bursting is abolished by dopamine D2 receptor antagonist, indicating that dopamine and NMDAR antagonist work synergistically to stimulate delta frequency bursting. Our results have significant implications concerning the electrophysiological basis of schizophrenia and bring together the NMDAR hypofunction, dopamine, and GABA theories of the disease. Our results suggest that NMDAR hypofunction and dopamine work synergistically on the GABAergic cells of the nRT to generate the delta frequency EEG oscillations, a thalamocortical dysrhythmia (TCD) in the awake state that is an established abnormality in schizophrenia. PMID:20057928

The Role of Dopamine Receptors in the Neurobehavioral Syndrome Provoked by Activation of L-Type Calcium Channels in Rodents

PubMed Central

Kasim, Suhail; Blake, Bonita L.; Fan, Xueliang; Chartoff, Elena; Egami, Kiyoshi; Breese, George R.; Hess, Ellen J.; Jinnah, H.A.

2010-01-01

In rodents, activation of L-type calcium channels with ± BayK 8644 causes an unusual behavioral syndrome that includes dystonia and self-biting. Prior studies have linked both of these behaviors to dysfunction of dopaminergic transmission in the striatum. The current studies were designed to further elucidate the relationship between ± BayK 8644 and dopaminergic transmission in the expression of the behavioral syndrome. The drug does not appear to release presynaptic dopamine stores, since microdialysis of the striatum revealed dopamine release was unaltered by ± BayK 8644. In addition, the behaviors were preserved or even exaggerated in mice or rats with virtually complete dopamine depletion. On the other hand, pretreatment of mice with D3 or D1/5 dopamine receptor antagonists attenuated the behavioral effects of ± BayK 8644, while pretreatment with D2 or D4 antagonists had no effect. In D3 receptor knockout mice, ± BayK 8644 elicited both dystonia and self-biting, but these behaviors were less severe than in matched controls. In D1 receptor knockout mice, behavioral responses to ± BayK 8644 appeared exaggerated. These results argue that the behavioral effects of ± BayK 8644 are not mediated by a presynaptic influence. Instead, the behaviors appear to result from a postsynaptic activation of the drug, which does not require but can be modified by D3 or D1/5 receptors. PMID:17028428

Nondopaminergic treatments for Parkinson's disease: current and future prospects

PubMed Central

Freitas, Maria Eliza; Fox, Susan H

2016-01-01

Parkinson's disease is primarily caused by dysfunction of dopaminergic neurons, however, nondopaminergic (ND) systems are also involved. ND targets are potentially useful to reduce doses of levodopa or to treat nonlevodopa-responsive symptoms. Recent studies have investigated the role of ND drugs for motor and nonmotor symptoms. Adenosine A2A receptor antagonists, mixed inhibitors of sodium/calcium channels and monoamine oxidase-B have recently been found to improve motor fluctuations. N-methyl-d-aspartate receptor antagonists and serotonin 5HT1B receptor agonists demonstrated benefit in levodopa-induced dyskinesia. Conversely, studies using antiepileptic drugs and adrenoreceptor antagonist had conflicting results. Moreover, metabotropic glutamate receptor antagonists also failed to improve symptoms. The current review summarizes the most recent findings on ND drugs over the last 2 years. PMID:27230697

Ozone (O{sub 3}) elicits neurotoxicity in spinal cord neurons (SCNs) by inducing ER Ca{sup 2+} release and activating the CaMKII/MAPK signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Yun; Lin, Xiaowen; Zhao, XueJun

Ozone (O{sub 3}) is widely used in the treatment of spinal cord related diseases. Excess or accumulation of this photochemical air can however be neurotoxic. In this study, in vitro cultured Wister rat spinal cord neurons (SCNs) were used to investigate the detrimental effects and underlying mechanisms of O{sub 3}. Ozone in a dose-dependent manner inhibited cell viability at a range of 20 to 500 μg/ml, with the dose at 40 μg/ml resulting in a decrease of cell viability to 75%. The cell death after O{sub 3} exposure was related to endoplasmic reticulum (ER) calcium (Ca{sup 2+}) release. Intracellular Ca{supmore » 2+} chelator, ER stabilizer (inositol 1,4,5-trisphosphate receptor (IP3R) antagonist and ryanodine receptor (RyR) antagonist) and calcium/calmodulin-dependent protein kinase II (CaMKII) antagonist could effectively block Ca{sup 2+} mobilization and inhibit cell death following 40 μg/ml O{sub 3} exposure. In addition, ER Ca{sup 2+} release due to O{sub 3} exposure enhanced phospho-p38 and phospho-JNK levels and apoptosis of SCNs through activating CaMKII. Based on these results, we confirm that ozone elicits neurotoxicity in SCNs via inducing ER Ca{sup 2+} release and activating CaMKII/MAPK signaling pathway. Therefore, physicians should get attention to the selection of treatment concentrations of oxygen/ozone. And, approaches, such as chelating intracellular Ca{sup 2+} and stabilizing neuronal Ca{sup 2+} homeostasis could effectively ameliorate the neurotoxicity of O{sub 3}. - Highlights: • Exposure to O{sub 3} can reduce the viability of SCNs and cause the cell death. • Exposure to O{sub 3} can trigger RyR and IP3R dependent intracellular Ca{sup 2+} release. • Exposure to O{sub 3} can enhance the phospho-CaMKII, phospho-JNK and phospho-p38 levels.« less

Transient receptor potential ankyrin 1 activation enhances hapten sensitization in a T-helper type 2-driven fluorescein isothiocyanate-induced contact hypersensitivity mouse model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shiba, Takahiro; Tamai, Takuma; Sahara, Yurina

2012-11-01

Some chemicals contribute to the development of allergies by increasing the immunogenicity of other allergens. We have demonstrated that several phthalate esters, including dibutyl phthalate (DBP), enhance skin sensitization to fluorescein isothiocyanate (FITC) in a mouse contact hypersensitivity model, in which the T-helper type 2 (Th2) response is essential. On the other hand, some phthalate esters were found to activate transient receptor potential ankyrin 1 (TRPA1) cation channels on sensory neurons. We then found a positive correlation between the enhancing effects of several types of phthalate esters on skin sensitization to FITC and their ability to activate TRPA1. Here wemore » examined the involvement of TRPA1 in sensitization to FITC by using TRPA1 agonists other than phthalate esters. During skin sensitization to FITC, the TRPA1 agonists (menthol, carvacrol, cinnamaldehyde and DBP) augmented the ear-swelling response as well as trafficking of FITC-presenting dendritic cells to draining lymph nodes. We confirmed that these TRPA1 agonists induced calcium influx into TRPA1-expressing Chinese hamster ovary (CHO) cells. We also found that TRPA1 antagonist HC-030031 inhibited DBP-induced calcium influx into TRPA1-expressing CHO cells. After pretreatment with this antagonist upon skin sensitization to FITC, the enhancing effect of DBP on sensitization was suppressed. These results suggest that TRPA1 activation will become a useful marker to find chemicals that facilitate sensitization in combination with other immunogenic haptens. -- Highlights: ► Role of TRPA1 activation was revealed in a mouse model of skin sensitization to FITC. ► TRPA1 agonists enhanced skin sensitization as well as dendritic cell trafficking. ► Dibutyl phthalate (DBP) has been shown to enhance skin sensitization to FITC. ► TRPA1 activation by DBP was inhibited by a selective antagonist, HC-030031. ► HC-030031 inhibited the enhancing effect of DBP on skin sensitization to FITC.« less

Toll-like receptor 7 agonists are potent and rapid bronchodilators in guinea pigs

PubMed Central

Kaufman, Elad H.; Fryer, Allison D.; Jacoby, David B.

2011-01-01

Background Respiratory tract viral infections result in asthma exacerbations. Toll-like receptor (TLR) 7 is a receptor for viral single-stranded RNA and is expressed at high levels in the lungs. Objective Because TLR7 polymorphisms are associated with asthma, we examined the effects of TLR7 agonists in guinea pig airways. Methods We induced bronchoconstriction in guinea pigs in vivo by means of electrical stimulation of the vagus nerve or intravenous administration of acetylcholine and measured the effect of a TLR7 agonist administered intravenously. We induced contraction of airway smooth muscle in segments of isolated guinea pig tracheas in vitro and measured the effect of TLR7 agonists, antagonists, and pharmacologic inhibitors of associated signaling pathways administered directly to the bath. Results TLR7 agonists acutely inhibited bronchoconstriction in vivo and relaxed contraction of airway smooth muscle in vitro within minutes of administration. Airway relaxation induced by the TLR7 agonist R837 (imiquimod) was partially blocked with a TLR7 antagonist and was also blocked by inhibitors of large-conductance, calcium-activated potassium channels; prostaglandin synthesis; and nitric oxide generation. Another TLR7 agonist, 21-mer single-stranded phosphorothioated polyuridylic acid (PolyUs), mediated relaxation that was completely blocked by a TLR7 antagonist. Conclusions These data demonstrate a novel protective mechanism to limit bronchoconstriction and maintain airflow during respiratory tract viral infections. The fast time frame is inconsistent with canonical TLR7 signaling. R837 mediates bronchodilation by means of TLR7-dependent and TLR7-independent mechanisms, whereas PolyUs does so through only the TLR7-dependent mechanism. TLR7-independent mechanisms involve prostaglandins and large-conductance, calcium-activated potassium channels, whereas TLR7-dependent mechanisms involve nitric oxide. TLR7 is an attractive therapeutic target for its ability to reverse bronchoconstriction within minutes. PMID:21167577

Efficacy and Tolerability of Nilvadipine in Combination with an Angiotensin II Receptor Antagonist in Patients with Essential Hypertension: A Multicenter, Open-Label, Uncontrolled Study

PubMed Central

Noda, Keita; Ideishi, Munehito; Tashiro, Eiichiro; Nakashima, Yoshiyuki; Imamura, Mitsuhide; Seki, Masahiko; Fujino, Masanori; Sou, Toshimitsu; Kohara, Masaki; Kanaya, Hisashi; Saku, Nishiki; Kamei, Ritsu; Yamasaki, Misao; Sakai, Hiroshi; Gondo, Naoki; Saku, Keijiro

2003-01-01

Background: Combination therapy with different classes of antihypertensive drugs often is needed to achieve controlled blood pressure (BP). The combination of an angiotensin II receptor antagonist (AIIA) and a calcium antagonist is a preferred option for reducing uncontrolled BP. Objective: The aim of this study was to assess the clinical efficacy and tolerability of nilvadipine, a dihydropyridine calcium antagonist, in combination with an AIIA. Methods: Patients with essential hypertension whose BP was not controlled by an AIIA alone were eligible for this multicenter, open-label, uncontrolled study. One of 3 AIIAs (candesartan cilexetil, losartan potassium, or valsartan) was given for at least 10 weeks before the addition of nilvadipine (daily dose, 4 or 8 mg orally). This combination therapy was given for 8 weeks. BP and heart rate were measured between 2 and 4 weeks before and 0, 4, and 8 weeks after the start of combination therapy. Adverse events were monitored at each visit. Results: Thirty-one patients (18 women [58.1%], 13 men [41.9%]; mean [SD] age, 58.5 [10.5] years) were enrolled. At weeks 4 and 8 of combination therapy, mean systolic BP (SBP) and diastolic BP (DBP) were significantly decreased (P<0.01) (at week 8, by 22.0 mm Hg and 12.5 mm Hg, respectively). The mean BP-lowering effect did not differ significantly between the 3 AIIAs tested. Pulse pressure also decreased significantly at week 8, by 9.6 mm Hg (P<0.01). The responder rate (ie, the percentage of patients with DBP <90 mm Hg or a decrease in DBP ≥10 mm Hg) was 72.0% at week 8. Three patients experienced a total of 4 adverse events: mild or severe flushing, mild headache, and mild palpitation. All of these symptoms resolved after nilvadipine treatment was discontinued. Conclusions: Nilvadipine in combination with an AIIA showed good antihypertensive efficacy and was well tolerated in the hypertensive patients in this study. This combination also significantly decreased pulse pressure, suggesting that this combination therapy also may have a beneficial effect in elderly patients with isolated systolic hypertension. PMID:25053855

Towards evidence-based emergency medicine: best BETs from the Manchester Royal Infirmary. BET 4: Alpha blockers v calcium blockers to increase spontaneous passage of renal calculi.

PubMed

Stewart, Alexander; Ferguson, Craig

2013-02-01

A short cut review was carried out to establish the administration of an alpha-1 receptor antagonist or a calcium channel blocker would facilitate the most rapid and successful expulsion of a stone from a patient with uncomplicated renal colic. 597 articles were found using the reported search, of which five trials were selected as providing the best evidence to answer this question. The authors, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses of these papers are tabulated. It is concluded that in a patient with an uncomplicated ureteric calculus tamsulosin is more effective than nifedipine in promoting speedy and successful expulsion of the stone.

Release of superoxide and change in morphology by neutrophils in response to phorbol esters: antagonism by inhibitors of calcium-binding proteins

PubMed Central

1985-01-01

The ability of phorbol derivatives to function as stimulating agents for superoxide (O2-) release by guinea pig neutrophils has been evaluated and compared to the known ability of each compound to activate protein kinase C. Those that activate the kinase also stimulate O2- release, while those that are inactive with respect to the kinase have no effect on O2- release. The same correlation was observed with respect to the ability of phorbol esters to induce morphological changes in neutrophils, i.e., vesiculation and reduction in granule content. Certain phenothiazines and naphthalene sulfonamides that are known antagonists of calcium-binding proteins blocked both phorbol ester-induced O2- release and morphological changes in these cells. PMID:2993312

Glutamate modulation of GABA transport in retinal horizontal cells of the skate

PubMed Central

Kreitzer, Matthew A; Andersen, Kristen A; Malchow, Robert Paul

2003-01-01

Transport of the amino acid GABA into neurons and glia plays a key role in regulating the effects of GABA in the vertebrate retina. We have examined the modulation of GABA-elicited transport currents of retinal horizontal cells by glutamate, the likely neurotransmitter of vertebrate photoreceptors. Enzymatically isolated external horizontal cells of skate were examined using whole-cell voltage-clamp techniques. GABA (1 mm) elicited an inward current that was completely suppressed by the GABA transport inhibitors tiagabine (10 μm) and SKF89976-A (100 μm), but was unaffected by 100 μm picrotoxin. Prior application of 100 μm glutamate significantly reduced the GABA-elicited current. Glutamate depressed the GABA dose-response curve without shifting the curve laterally or altering the voltage dependence of the current. The ionotropic glutamate receptor agonists kainate and AMPA also reduced the GABA-elicited current, and the effects of glutamate and kainate were abolished by the ionotropic glutamate receptor antagonist 6-cyano-7-nitroquinoxaline. NMDA neither elicited a current nor modified the GABA-induced current, and metabotropic glutamate analogues were also without effect. Inhibition of the GABA-elicited current by glutamate and kainate was reduced when extracellular calcium was removed and when recording pipettes contained high concentrations of the calcium chelator BAPTA. Caffeine (5 mm) and thapsigargin (2 nm), agents known to alter intracellular calcium levels, also reduced the GABA-elicited current, but increases in calcium induced by depolarization alone did not. Our data suggest that glutamate regulates GABA transport in retinal horizontal cells through a calcium-dependent process, and imply a close physical relationship between calcium-permeable glutamate receptors and GABA transporters in these cells. PMID:12562999

Effects of nifedipine on anorectal smooth muscle in vitro.

PubMed

Cook, T A; Brading, A F; Mortensen, N J

1999-06-01

Glyceryl trinitrate reduces anal resting pressure and aids the healing of anal fissures. However, some patients develop tachyphylaxis and the fissure fails to heal, suggesting that other agents are needed. This study assesses the effects of nifedipine (a calcium channel antagonist) in modulating resting tone and agonist-induced contractions in human internal anal sphincter (IAS) and rectal circular muscle. Smooth muscle strips from the IAS and rectal circular muscle from ten patients undergoing surgical resection were mounted for isometric tension recording in a superfusion organ bath. The effects of noradrenaline and carbachol were assessed in the presence of various perfusates. LAS strips developed tone and spontaneous activity. Noradrenaline produced dose-dependent contractions. In calcium-free Krebs solution, tone and activity were abolished and no contractions were elicited in response to noradrenaline. Nifedipine also abolished tone and spontaneous activity, but contractions to noradrenaline were only slightly attenuated. In contrast, rectal smooth muscle strips developed spontaneous activity but no resting tone and contracted in response to carbachol. In calcium-free Krebs solution, the spontaneous activity and carbachol contractions were abolished. Addition of nifedipine to the perfusate abolished spontaneous activity and greatly reduced contractions. These data suggest that spontaneous activity and resting tone are dependent on extracellular calcium and flux across the cells. Agonist-induced contraction in the IAS is attributable mainly to the release of calcium from intracellular stores, whereas rectal circular smooth muscle depends principally on extracellular calcium entering the cell for contraction. The attenuation of contractions in both tissues and the abolition of resting tone in the IAS suggest that nifedipine may be useful in the management of patients with anorectal disorders.

GABAB receptor modulation of the release of substance P from capsaicin-sensitive neurones in the rat trachea in vitro.

PubMed Central

Ray, N. J.; Jones, A. J.; Keen, P.

1991-01-01

1. The role of gamma-aminobutyric acid (GABA) as an inhibitory transmitter in the central nervous system is well documented. Recently, GABAA and GABAB receptors have been identified in the peripheral nervous system, notably on primary afferent neurones (PAN). We have utilised a multi-superfusion system to investigate the effect of selective GABA receptor agonists and antagonists on the release of substance P (SP) from the rat trachea in vitro. 2. GABA (1-100 microM) did not affect spontaneous release of SP-like immunoreactivity (LI) but caused dose-related inhibition of calcium-dependent potassium (60 mM)-stimulated SP-LI release. The greatest inhibition of 77.7 +/- 18.8% was observed at 100 microM. 3. The inhibitory effect of GABA was mimicked by the GABAB receptor agonist, (+/-)-baclofen (1-100 microM), but not the GABAA receptor agonist, 3-amino-1-propane-sulphonic acid (3-APS, 1-100 microM). Baclofen (100 microM) had no effect on SP-LI release stimulated by capsaicin (1 microM). 4. The inhibitory effect of baclofen (30 microM) was significantly reduced by prior and concomitant exposure to the GABAB receptor antagonist, phacolofen (100 microM) but not the GABAA receptor antagonist, bicuculline (10 microM). Neither antagonist, alone, affected spontaneous or potassium-stimulated SP-LI release. 5. We conclude that activation of pre-synaptic GABAB receptors on the peripheral termini of PANs in the rat trachea inhibits SP-LI release and suggest that GABAB receptor agonists may be of value in the therapeutic treatment of asthma. PMID:1713105

Growth hormone-releasing hormone as an agonist of the ghrelin receptor GHS-R1a

PubMed Central

Casanueva, Felipe F.; Camiña, Jesus P.; Carreira, Marcos C.; Pazos, Yolanda; Varga, Jozsef L.; Schally, Andrew V.

2008-01-01

Ghrelin synergizes with growth hormone-releasing hormone (GHRH) to potentiate growth hormone (GH) response through a mechanism not yet fully characterized. This study was conducted to analyze the role of GHRH as a potential ligand of the ghrelin receptor, GHS-R1a. The results show that hGHRH(1–29)NH2 (GHRH) induces a dose-dependent calcium mobilization in HEK 293 cells stably transfected with GHS-R1a an effect not observed in wild-type HEK 293 cells. This calcium rise is also observed using the GHRH receptor agonists JI-34 and JI-36. Radioligand binding and cross-linking studies revealed that calcium response to GHRH is mediated by the ghrelin receptor GHS-R1a. GHRH activates the signaling route of inositol phosphate and potentiates the maximal response to ghrelin measured in inositol phosphate turnover. The presence of GHRH increases the binding capacity of 125I-ghrelin in a dose dependent-fashion showing a positive binding cooperativity. In addition, confocal microscopy in CHO cells transfected with GHS-R1a tagged with enhanced green fluorescent protein shows that GHRH activates the GHS-R1a endocytosis. Furthermore, the selective GHRH-R antagonists, JV-1–42 and JMR-132, act also as antagonists of the ghrelin receptor GHS-R1a. Our findings suggest that GHRH interacts with ghrelin receptor GHS-R1a, and, in consequence, modifies the ghrelin-associated intracellular signaling pathway. This interaction may represent a form of regulation, which could play a putative role in the physiology of GH regulation and appetite control. PMID:19088192

Growth hormone-releasing hormone as an agonist of the ghrelin receptor GHS-R1a.

PubMed

Casanueva, Felipe F; Camiña, Jesus P; Carreira, Marcos C; Pazos, Yolanda; Varga, Jozsef L; Schally, Andrew V

2008-12-23

Ghrelin synergizes with growth hormone-releasing hormone (GHRH) to potentiate growth hormone (GH) response through a mechanism not yet fully characterized. This study was conducted to analyze the role of GHRH as a potential ligand of the ghrelin receptor, GHS-R1a. The results show that hGHRH(1-29)NH(2) (GHRH) induces a dose-dependent calcium mobilization in HEK 293 cells stably transfected with GHS-R1a an effect not observed in wild-type HEK 293 cells. This calcium rise is also observed using the GHRH receptor agonists JI-34 and JI-36. Radioligand binding and cross-linking studies revealed that calcium response to GHRH is mediated by the ghrelin receptor GHS-R1a. GHRH activates the signaling route of inositol phosphate and potentiates the maximal response to ghrelin measured in inositol phosphate turnover. The presence of GHRH increases the binding capacity of (125)I-ghrelin in a dose dependent-fashion showing a positive binding cooperativity. In addition, confocal microscopy in CHO cells transfected with GHS-R1a tagged with enhanced green fluorescent protein shows that GHRH activates the GHS-R1a endocytosis. Furthermore, the selective GHRH-R antagonists, JV-1-42 and JMR-132, act also as antagonists of the ghrelin receptor GHS-R1a. Our findings suggest that GHRH interacts with ghrelin receptor GHS-R1a, and, in consequence, modifies the ghrelin-associated intracellular signaling pathway. This interaction may represent a form of regulation, which could play a putative role in the physiology of GH regulation and appetite control.

Pharmacological characterization of desensitization in scratching behavior induced by intrathecal administration of hemokinin-1 in the rat.

PubMed

Naono, R; Nakayama, T; Ikeda, T; Matsusima, O; Nishimori, T

2008-02-01

Desensitization is induced by the repeated administration of high doses of substance P (SP) or hemokinin-1 (HK-1). However, little information is available about the mechanisms involved in the induction of desensitization by these peptides. Thus, to characterize this desensitization, we examined the dose-dependent effect of these peptides, the effect of pretreatment with neurokinin 1(NK1) receptor antagonists, and the effect of pretreatment with inhibitors of protein kinases such as protein kinase A (PKA), protein kinase C (PKC), calcium/calmodulin kinase II (CaMKII) and mitogen-activated protein kinase kinase (MEK). The number of scratchings induced by 10(-3)M SP or HK-1 decreased following pretreatment with 10(-11)-10(-3)M SP or HK-1 with a marked reduction at 10(-3) and 10(-6)M SP or HK-1. The effect of NK1 receptor antagonists on desensitization induced by pretreatment with 10(-6)M SP was marked, whereas there was little effect of pretreatment with these antagonists on 10(-6)M HK-1-induced desensitization. Additionally, 10(-6)M SP- and HK-1-induced desensitization was attenuated by pretreatment with PKA, PKC and MEK inhibitors, except a CaMKII inhibitor that inhibited SP-induced desensitization. These results indicate that the receptor and kinases involved in HK-1-induced desensitization are partially different from those of SP.

Calcitonin gene-related peptide (CGRP) activates human neutrophils--inhibition by chemotactic peptide antagonist BOC-MLP.

PubMed Central

Richter, J; Andersson, R; Edvinsson, L; Gullberg, U

1992-01-01

The effect of the neuropeptides substance P, neurokinin A and alpha-calcitonin gene-related peptide (CGRP) on human neutrophil granulocytes was investigated. Substance P induced secondary granule secretion at a concentration of 100 microM. CGRP induced a significant secretory response at 10 microM and thus appeared to be about 10 times more potent than substance P. Calcitonin and a fragment of CGRP, CGRP(8-37), had no effect on neutrophil degranulation. The chemotactic peptide antagonist BOC-MLP (100 microM) inhibited lactoferrin secretion mediated both by CGRP and chemotactic peptide FMLP almost completely, while secretion in response to tumour necrosis factor (TNF) was unaffected. Results from receptor binding studies showed that CGRP and N-formyl-methionyl-leucyl-phenylalanine (FMLP) do not compete for binding. This indicates that CGRP does not exert its effects by binding to the chemotactic peptide receptor. CGRP induced a rapid increase in the cytosolic-free calcium concentration and this increase was not, unlike that induced by FMLP, abolished by preincubation of the cells with pertussis toxin (1000 ng/ml). Therefore CGRP signal transduction in neutrophils appears to involve rapid changes in the cytosolic-free calcium concentration but not a pertussis toxin-sensitive G-protein. In summary, this is the first report to show that CGRP can directly activate neutrophil granulocytes, and this probably occurs via a cell surface receptor which is distinct from that of FMLP although both the CGRP and FMLP-mediated effects can be blocked by BOC-MLP. Images Figure 3 PMID:1282494

Enhancement of mesenchymal stem cell chondrogenesis with short-term low intensity pulsed electromagnetic fields.

PubMed

Parate, Dinesh; Franco-Obregón, Alfredo; Fröhlich, Jürg; Beyer, Christian; Abbas, Azlina A; Kamarul, Tunku; Hui, James H P; Yang, Zheng

2017-08-25

Pulse electromagnetic fields (PEMFs) have been shown to recruit calcium-signaling cascades common to chondrogenesis. Here we document the effects of specified PEMF parameters over mesenchymal stem cells (MSC) chondrogenic differentiation. MSCs undergoing chondrogenesis are preferentially responsive to an electromagnetic efficacy window defined by field amplitude, duration and frequency of exposure. Contrary to conventional practice of administering prolonged and repetitive exposures to PEMFs, optimal chondrogenic outcome is achieved in response to brief (10 minutes), low intensity (2 mT) exposure to 6 ms bursts of magnetic pulses, at 15 Hz, administered only once at the onset of chondrogenic induction. By contrast, repeated exposures diminished chondrogenic outcome and could be attributed to calcium entry after the initial induction. Transient receptor potential (TRP) channels appear to mediate these aspects of PEMF stimulation, serving as a conduit for extracellular calcium. Preventing calcium entry during the repeated PEMF exposure with the co-administration of EGTA or TRP channel antagonists precluded the inhibition of differentiation. This study highlights the intricacies of calcium homeostasis during early chondrogenesis and the constraints that are placed on PEMF-based therapeutic strategies aimed at promoting MSC chondrogenesis. The demonstrated efficacy of our optimized PEMF regimens has clear clinical implications for future regenerative strategies for cartilage.

Depletion of calcium stores regulates calcium influx and signal transmission in rod photoreceptors

PubMed Central

Szikra, Tamas; Cusato, Karen; Thoreson, Wallace B; Barabas, Peter; Bartoletti, Theodore M; Krizaj, David

2008-01-01

Tonic synapses are specialized for sustained calcium entry and transmitter release, allowing them to operate in a graded fashion over a wide dynamic range. We identified a novel plasma membrane calcium entry mechanism that extends the range of rod photoreceptor signalling into light-adapted conditions. The mechanism, which shares molecular and physiological characteristics with store-operated calcium entry (SOCE), is required to maintain baseline [Ca2+]i in rod inner segments and synaptic terminals. Sustained Ca2+ entry into rod cytosol is augmented by store depletion, blocked by La3+ and Gd3+ and suppressed by organic antagonists MRS-1845 and SKF-96365. Store depletion and the subsequent Ca2+ influx directly stimulated exocytosis in terminals of light-adapted rods loaded with the activity-dependent dye FM1–43. Moreover, SOCE blockers suppressed rod-mediated synaptic inputs to horizontal cells without affecting presynaptic voltage-operated Ca2+ entry. Silencing of TRPC1 expression with small interference RNA disrupted SOCE in rods, but had no effect on cone Ca2+ signalling. Rods were immunopositive for TRPC1 whereas cone inner segments immunostained with TRPC6 channel antibodies. Thus, SOCE modulates Ca2+ homeostasis and light-evoked neurotransmission at the rod photoreceptor synapse mediated by TRPC1. PMID:18755743

Adrenomedullin increases the short-circuit current in the rat prostate: Receptors, chloride channels, the effects of cAMP and calcium ions and implications on fluid secretion.

PubMed

Liao, S B; Cheung, K H; Cheung, M P L; Wong, P F; O, W S; Tang, F

2014-05-01

In this study, we have investigated the effects of adrenomedullin on chloride and fluid secretion in the rat prostate. The presence of adrenomedullin (ADM) in rat prostate was confirmed using immunostaining, and the molecular species was determined using gel filtration chromatography coupled with an enzyme-linked assay for ADM. The effects of ADM on fluid secretion were studied by short-circuit current technique in a whole mount preparation of the prostate in an Ussing chamber. The results indicated that the ADM level was higher in the ventral than the dorso-lateral prostate and the major molecular species was the active peptide. ADM increased the short-circuit current through both the cAMP- and calcium-activated chloride channels in the ventral lobe, but only through the calcium-activated channels in the dorso-lateral lobe. These stimulatory effects were blocked by the calcitonin gene-related peptide (CGRP) receptor antagonist, hCGRP8-37. We conclude that ADM may regulate prostatic fluid secretion through the chloride channels, which may affect the composition of the seminal plasma bathing the spermatozoa and hence fertility. © 2014 American Society of Andrology and European Academy of Andrology.

Hyperinsulin therapy for calcium channel antagonist poisoning: a seven-year retrospective study.

PubMed

Espinoza, Tamara R; Bryant, Sean M; Aks, Steve E

2013-01-01

The use of hyperinsulin therapy (HIT) in severe calcium channel antagonist (CCA) poisoning has become a more common therapy within the last decade. The objective of this study is to report 7 years of experience recommending HIT. This was a retrospective chart review utilizing our regional poison center (RPC) data from January 1, 2002, through December 31, 2008. All cases of CCA poisoning receiving HIT were searched. Endpoints included the number of CCA cases utilizing HIT, insulin dose, time of initiation of HIT, patient outcome, adverse events, age, glucose concentration, and lowest systolic blood pressure recorded. Forty-six cases of CCA poisoning were managed with HIT over 7 years. All the patients received standard antidotal therapy (= intravenous fluids, calcium salts, glucagon, and pressors). HIT administration followed our RPC recommendation 23 times (50%), and no hypoglycemic events occurred. Means (age, highest glucose measured, and lowest systolic blood pressure measured) were 51 years, 282 mg/dL, and 74 mm Hg, respectively. Our RPC recommendations for HIT were followed 50% of the time over the last 7 years. In light of the lack of hypoglycemia associated with HIT in our study population, we recommend HIT as an early and safe antidote in significant CCA poisoning.

PURIFICATION OF AN INFLUENZA VIRUS SUBSTRATE, AND DEMONSTRATION OF ITS COMPETITIVE ANTAGONISM TO APPLE PECTIN

PubMed Central

Woolley, D. W.

1949-01-01

A substance was demonstrated in erythrocytes which antagonized the inhibiting effect of apple pectin on influenza virus hemagglutination. This substance was purified and found to be a water-soluble material, rather labile, and with some properties which suggested that it contained a polysaccharide. It was destroyed in vitro by highly purified preparations of the virus. It occurred in greatest amount in human erythrocytes and to a lesser extent in the red cells of species not susceptible to the virus. It was also found in normal rabbit serum. Calcium ions were found to be essential to the action of apple pectin in causing inhibition of virus hemagglutination. A second substance was purified from an alkaline extract of erythrocytes, and shown likewise to have an effect antagonistic to that of pectin. However, this latter material was not destroyed by the virus, and seemed to owe its effect to the binding of calcium ions. PMID:18099162

[Calcium kidney stones. Diagnostic and preventive prospects].

PubMed

Arcidiacono, T; Terranegra, A; Biasion, R; Soldati, L; Vezzoli, G

2007-01-01

Kidney stone disease is one of the main causes of hospitalization in Italy. Its prevalence increased in the last century and is probably still increasing. The pathogenesis of the disease is not known, although two main theories have been elaborated. The first hypothesizes that hydroxyapatite deposition in the interstitium of the renal papillae (Randall's plaque) precedes urinary calcium oxalate precipitation on the ulcered surface of the papilla to form a stone. The second presumes the tubular lumen of Bellini's duct to be the site where calcium-oxalate salts precipitate to form the nucleus for stone formation within the urinary tract. These pathogenetic processes may be favored by different dietary and genetic factors. The genes involved are not known, although many studies have been performed. Polymorphisms of genes coding for the vitamin D receptor, calcium-sensing receptor, interleukin-1 receptor antagonist, and urokinase were found to be associated with kidney stones, but these results have not been replicated. Different nutrients are suspected to predispose patients to calcium kidney stone disease. A high intake of animal proteins, sodium, vitamin C and oxalate has been implicated in stone formation, whereas calcium, alkalis and phytate may have a protective effect. The prevention of calcium stone formation is based on the recognition of risk factors like those already mentioned here. Furthermore, a family history of kidney stones may be useful in identifying subjects predisposed to become calcium stone formers. However, the expectations of the scientific community are turned to the advances in genetics and to the findings of genetic studies, which may provide diagnostic tools and criteria to define the risk profile of the single individual.

Activation of muscarinic acetylcholine receptors elicits pigment granule dispersion in retinal pigment epithelium isolated from bluegill.

PubMed

González, Alfredo; Crittenden, Elizabeth L; García, Dana M

2004-07-13

In fish, melanin pigment granules in the retinal pigment epithelium disperse into apical projections as part of the suite of responses the eye makes to bright light conditions. This pigment granule dispersion serves to reduce photobleaching and occurs in response to neurochemicals secreted by the retina. Previous work has shown that acetylcholine may be involved in inducing light-adaptive pigment dispersion. Acetylcholine receptors are of two main types, nicotinic and muscarinic. Muscarinic receptors are in the G-protein coupled receptor superfamily, and five different muscarinic receptors have been molecularly cloned in human. These receptors are coupled to adenylyl cyclase, calcium mobilization and ion channel activation. To determine the receptor pathway involved in eliciting pigment granule migration, we isolated retinal pigment epithelium from bluegill and subjected it to a battery of cholinergic agents. The general cholinergic agonist carbachol induces pigment granule dispersion in isolated retinal pigment epithelium. Carbachol-induced pigment granule dispersion is blocked by the muscarinic antagonist atropine, by the M1 antagonist pirenzepine, and by the M3 antagonist 4-DAMP. Pigment granule dispersion was also induced by the M1 agonist 4-[N-(4-chlorophenyl) carbamoyloxy]-4-pent-2-ammonium iodide. In contrast the M2 antagonist AF-DX 116 and the M4 antagonist tropicamide failed to block carbachol-induced dispersion, and the M2 agonist arecaidine but-2-ynyl ester tosylate failed to elicit dispersion. Our results suggest that carbachol-mediated pigment granule dispersion occurs through the activation of Modd muscarinic receptors, which in other systems couple to phosphoinositide hydrolysis and elevation of intracellular calcium. This conclusion must be corroborated by molecular studies, but suggests Ca2+-dependent pathways may be involved in light-adaptive pigment dispersion.

Inhibition of Listeria monocytogenes by food antimicrobials applied singly and in combination.

PubMed

Brandt, Alex L; Castillo, Alejandro; Harris, Kerri B; Keeton, Jimmy T; Hardin, Margaret D; Taylor, Thomas M

2010-01-01

Combining food antimicrobials can enhance inhibition of Listeria monocytogenes in ready-to-eat (RTE) meats. A broth dilution assay was used to compare the inhibition of L. monocytogenes resulting from exposure to nisin, acidic calcium sulfate, ε-poly-L-lysine, and lauric arginate ester applied singly and in combination. Minimum inhibitory concentrations (MICs) were the lowest concentrations of single antimicrobials producing inhibition following 24 h incubation at 35 °C. Minimum bactericidal concentrations (MBCs) were the lowest concentrations that decreased populations by ≥3.0 log(10) CFU/mL. Combinations of nisin with acidic calcium sulfate, nisin with lauric arginate ester, and ɛ-poly-L-lysine with acidic calcium sulfate were prepared using a checkerboard assay to determine optimal inhibitory combinations (OICs). Fractional inhibitory concentrations (FICs) were calculated from OICs and were used to create FIC indices (FIC(I)s) and isobolograms to classify combinations as synergistic (FIC(I) < 1.00), additive/indifferent (FIC(I)= 1.00), or antagonistic (FIC(I) > 1.00). MIC values for nisin ranged from 3.13 to 6.25 μg/g with MBC values at 6.25 μg/g for all strains except for Natl. Animal Disease Center (NADC) 2045. MIC values for ε-poly-L-lysine ranged from 6.25 to 12.50 μg/g with MBCs from 12.50 to 25.00 μg/g. Lauric arginate ester at 12.50 μg/g was the MIC and MBC for all strains; 12.50 mL/L was the MIC and MBC for acidic calcium sulfate. Combining nisin with acidic calcium sulfate synergistically inhibited L. monocytogenes; nisin with lauric arginate ester produced additive-type inhibition, while ε-poly-L-lysine with acidic calcium sulfate produced antagonistic-type inhibition. Applying nisin along with acidic calcium sulfate should be further investigated for efficacy on RTE meat surfaces. © 2010 Institute of Food Technologists®

Characterization of postsynaptic calcium signals in the pyramidal neurons of anterior cingulate cortex

PubMed Central

Li, Xu-Hui; Song, Qian; Chen, Tao; Zhuo, Min

2017-01-01

Calcium signaling is critical for synaptic transmission and plasticity. N-methyl-D-aspartic acid (NMDA) receptors play a key role in synaptic potentiation in the anterior cingulate cortex. Most previous studies of calcium signaling focus on hippocampal neurons, little is known about the activity-induced calcium signals in the anterior cingulate cortex. In the present study, we show that NMDA receptor-mediated postsynaptic calcium signals induced by different synaptic stimulation in anterior cingulate cortex pyramidal neurons. Single and multi-action potentials evoked significant suprathreshold Ca2+ increases in somas and spines. Both NMDA receptors and voltage-gated calcium channels contributed to this increase. Postsynaptic Ca2+signals were induced by puff-application of glutamate, and a NMDA receptor antagonist AP5 blocked these signals in both somas and spines. Finally, long-term potentiation inducing protocols triggered postsynaptic Ca2+ influx, and these influx were NMDA receptor dependent. Our results provide the first study of calcium signals in the anterior cingulate cortex and demonstrate that NMDA receptors play important roles in postsynaptic calcium signals in anterior cingulate cortex pyramidal neurons. PMID:28726541

P2X1 Receptor Antagonists Inhibit HIV-1 Fusion by Blocking Virus-Coreceptor Interactions

PubMed Central

Giroud, Charline; Marin, Mariana; Hammonds, Jason; Spearman, Paul

2015-01-01

ABSTRACT HIV-1 Env glycoprotein-mediated fusion is initiated upon sequential binding of Env to CD4 and the coreceptor CXCR4 or CCR5. Whereas these interactions are thought to be necessary and sufficient to promote HIV-1 fusion, other host factors can modulate this process. Previous studies reported potent inhibition of HIV-1 fusion by selective P2X1 receptor antagonists, including NF279, and suggested that these receptors play a role in HIV-1 entry. Here we investigated the mechanism of antiviral activity of NF279 and found that this compound does not inhibit HIV-1 fusion by preventing the activation of P2X1 channels but effectively blocks the binding of the virus to CXCR4 or CCR5. The notion of an off-target effect of NF279 on HIV-1 fusion is supported by the lack of detectable expression of P2X1 receptors in cells used in fusion experiments and by the fact that the addition of ATP or the enzymatic depletion of ATP in culture medium does not modulate viral fusion. Importantly, NF279 fails to inhibit HIV-1 fusion with cell lines and primary macrophages when added at an intermediate stage downstream of Env-CD4-coreceptor engagement. Conversely, in the presence of NF279, HIV-1 fusion is arrested downstream of CD4 binding but prior to coreceptor engagement. NF279 also antagonizes the signaling function of CCR5, CXCR4, and another chemokine receptor, as evidenced by the suppression of calcium responses elicited by specific ligands and by recombinant gp120. Collectively, our results demonstrate that NF279 is a dual HIV-1 coreceptor inhibitor that interferes with the functional engagement of CCR5 and CXCR4 by Env. IMPORTANCE Inhibition of P2X receptor activity suppresses HIV-1 fusion and replication, suggesting that P2X signaling is involved in HIV-1 entry. However, mechanistic experiments conducted in this study imply that P2X1 receptor is not expressed in target cells or involved in viral fusion. Instead, we found that inhibition of HIV-1 fusion by a specific P2X1 receptor antagonist, NF279, is due to the blocking of virus interactions with both the CXCR4 and CCR5 coreceptors. The ability of NF279 to abrogate cellular calcium signaling induced by the respective chemokines showed that this compound acts as a dual-coreceptor antagonist. P2X1 receptor antagonists could thus represent a new class of dual-coreceptor inhibitors with a structure and a mechanism of action that are distinct from those of known HIV-1 coreceptor antagonists. PMID:26136569

P2X1 Receptor Antagonists Inhibit HIV-1 Fusion by Blocking Virus-Coreceptor Interactions.

PubMed

Giroud, Charline; Marin, Mariana; Hammonds, Jason; Spearman, Paul; Melikyan, Gregory B

2015-09-01

HIV-1 Env glycoprotein-mediated fusion is initiated upon sequential binding of Env to CD4 and the coreceptor CXCR4 or CCR5. Whereas these interactions are thought to be necessary and sufficient to promote HIV-1 fusion, other host factors can modulate this process. Previous studies reported potent inhibition of HIV-1 fusion by selective P2X1 receptor antagonists, including NF279, and suggested that these receptors play a role in HIV-1 entry. Here we investigated the mechanism of antiviral activity of NF279 and found that this compound does not inhibit HIV-1 fusion by preventing the activation of P2X1 channels but effectively blocks the binding of the virus to CXCR4 or CCR5. The notion of an off-target effect of NF279 on HIV-1 fusion is supported by the lack of detectable expression of P2X1 receptors in cells used in fusion experiments and by the fact that the addition of ATP or the enzymatic depletion of ATP in culture medium does not modulate viral fusion. Importantly, NF279 fails to inhibit HIV-1 fusion with cell lines and primary macrophages when added at an intermediate stage downstream of Env-CD4-coreceptor engagement. Conversely, in the presence of NF279, HIV-1 fusion is arrested downstream of CD4 binding but prior to coreceptor engagement. NF279 also antagonizes the signaling function of CCR5, CXCR4, and another chemokine receptor, as evidenced by the suppression of calcium responses elicited by specific ligands and by recombinant gp120. Collectively, our results demonstrate that NF279 is a dual HIV-1 coreceptor inhibitor that interferes with the functional engagement of CCR5 and CXCR4 by Env. Inhibition of P2X receptor activity suppresses HIV-1 fusion and replication, suggesting that P2X signaling is involved in HIV-1 entry. However, mechanistic experiments conducted in this study imply that P2X1 receptor is not expressed in target cells or involved in viral fusion. Instead, we found that inhibition of HIV-1 fusion by a specific P2X1 receptor antagonist, NF279, is due to the blocking of virus interactions with both the CXCR4 and CCR5 coreceptors. The ability of NF279 to abrogate cellular calcium signaling induced by the respective chemokines showed that this compound acts as a dual-coreceptor antagonist. P2X1 receptor antagonists could thus represent a new class of dual-coreceptor inhibitors with a structure and a mechanism of action that are distinct from those of known HIV-1 coreceptor antagonists. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Expression and coupling of neurokinin receptor subtypes to inositol phosphate and calcium signaling pathways in human airway smooth muscle cells

PubMed Central

Mizuta, Kentaro; Gallos, George; Zhu, Defen; Mizuta, Fumiko; Goubaeva, Farida; Xu, Dingbang; Panettieri, Reynold A.; Yang, Jay; Emala, Charles W.

2013-01-01

Neuropeptide tachykinins (substance P, neurokinin A, and neurokinin B) are present in peripheral terminals of sensory nerve fibers within the respiratory tract and cause airway contractile responses and hyperresponsiveness in humans and most mammalian species. Three subtypes of neurokinin receptors (NK1R, NK2R, and NK3R) classically couple to Gq protein-mediated inositol 1,4,5-trisphosphate (IP3) synthesis and liberation of intracellular Ca2+, which initiates contraction, but their expression and calcium signaling mechanisms are incompletely understood in airway smooth muscle. All three subtypes were identified in native and cultured human airway smooth muscle (HASM) and were subsequently overexpressed in HASM cells using a human immunodeficiency virus-1-based lentivirus transduction system. Specific NKR agonists {NK1R, [Sar9,Met(O2)11]-substance P; NK2R, [β-Ala8]-neurokinin A(4–10); NK3R, senktide} stimulated inositol phosphate synthesis and increased intracellular Ca2+ concentration ([Ca2+]i) in native HASM cells and in HASM cells transfected with each NKR subtype. These effects were blocked by NKR-selective antagonists (NK1R, L-732138; NK2R, GR-159897; NK3R, SB-222200). The initial transient and sustained phases of increased [Ca2+]i were predominantly inhibited by the IP3 receptor antagonist 2-aminoethoxydiphenyl borate (2-APB) or the store-operated Ca2+ channel antagonist SKF-96365, respectively. These results show that all three subtypes of NKRs are expressed in native HASM cells and that IP3 levels are the primary mediators of NKR-stimulated initial [Ca2+]i increases, whereas store-operated Ca2+ channels mediate the sustained phase of the [Ca2+]i increase. PMID:18203813

Expression and coupling of neurokinin receptor subtypes to inositol phosphate and calcium signaling pathways in human airway smooth muscle cells.

PubMed

Mizuta, Kentaro; Gallos, George; Zhu, Defen; Mizuta, Fumiko; Goubaeva, Farida; Xu, Dingbang; Panettieri, Reynold A; Yang, Jay; Emala, Charles W

2008-03-01

Neuropeptide tachykinins (substance P, neurokinin A, and neurokinin B) are present in peripheral terminals of sensory nerve fibers within the respiratory tract and cause airway contractile responses and hyperresponsiveness in humans and most mammalian species. Three subtypes of neurokinin receptors (NK1R, NK2R, and NK3R) classically couple to Gq protein-mediated inositol 1,4,5-trisphosphate (IP3) synthesis and liberation of intracellular Ca2+, which initiates contraction, but their expression and calcium signaling mechanisms are incompletely understood in airway smooth muscle. All three subtypes were identified in native and cultured human airway smooth muscle (HASM) and were subsequently overexpressed in HASM cells using a human immunodeficiency virus-1-based lentivirus transduction system. Specific NKR agonists {NK1R, [Sar9,Met(O2)11]-substance P; NK2R, [beta-Ala8]-neurokinin A(4-10); NK3R, senktide} stimulated inositol phosphate synthesis and increased intracellular Ca2+ concentration ([Ca2+]i) in native HASM cells and in HASM cells transfected with each NKR subtype. These effects were blocked by NKR-selective antagonists (NK1R, L-732138; NK2R, GR-159897; NK3R, SB-222200). The initial transient and sustained phases of increased [Ca2+]i were predominantly inhibited by the IP3 receptor antagonist 2-aminoethoxydiphenyl borate (2-APB) or the store-operated Ca2+ channel antagonist SKF-96365, respectively. These results show that all three subtypes of NKRs are expressed in native HASM cells and that IP3 levels are the primary mediators of NKR-stimulated initial [Ca2+]i increases, whereas store-operated Ca2+ channels mediate the sustained phase of the [Ca2+]i increase.

Role of Hyperkalemia in Heart Failure and the Therapeutic Use of Potassium Binders.

PubMed

Sarwar, Chaudhry M S; Bhagat, Aditi A; Anker, Stefan D; Butler, Javed

2017-01-01

Hyperkalemia can be a life-threatening disorder, especially for at-risk patients with heart failure, chronic kidney disease, with diabetes, and patients on certain drugs like renin-angiotensin-aldosterone system antagonists and mineralocorticoid receptor antagonists. There are limited therapeutic options available for hyperkalemia, and they have narrow effectiveness because of their unfavorable side effects profile in long-term and high cost utilization requiring inpatient care. Patiromersorbitex calcium and sodium zirconium cyclosilicate are novel potassium-lowering compounds for the treatment and prevention of hyperkalemia in at-risk population. These therapeutic agents have shown encouraging results in early phase II and phase III clinical trials. However, there is need to further study their efficacy and safety in heart failure population in order to establish their clinical use. The focus of this chapter will be to promote better understanding of potassium homeostasis in heart failure patients and the mechanistic overview of novel drugs, with emphasis on heart failure population.

Different effect of losartan and amlodipine on penile structures in male spontaneously hypertensive rats.

PubMed

Toblli, Jorge Eduardo; Stella, Inés; Mazza, Osvaldo Néstor; Ferder, León; Inserra, Felipe

2004-01-01

Erectile dysfunction is highly prevalent in hypertensive patients. Since both angiotensin II receptor type-1 blockers (ARBs) and calcium antagonists are current and effective antihypertensive drugs, the aim of this study was to determine possible differences between ARBs and calcium antagonists concerning the protection of penile structures from the deleterious effects of arterial hypertension. During 6 months, 3 groups of male spontaneously hypertensive rats (SHR) and 1 of Wistar-Kyoto (WKY) rats, as a control group, were studied: SHR without treatment; SHR with losartan (L) 30 mg/kg/day; SHR with amlodipine (A) 3 mg/kg/day, and WKY without treatment. Cavernous smooth muscle (CSM) and vascular smooth muscle (VSM) from cavernous arteries, cavernous tissue fibrosis and collagen type III (COL III) were evaluated. After 6 months, SHR+L and SHR+A showed a similar reduction in blood pressure compared with untreated SHR. However, only SHR+L and control WKY presented significantly lower values of: CSM (p < 0.01), VSM (p < 0.01), and COL III (p < 0.01) when compared with either untreated SHR or SHR+A. There was also a positive correlation between left ventricular mass and proteinuria with VSM from cavernous arteries, CSM and COL III in untreated SHR and SHR+A. These relations were not present in SHR+L and WKY. Although losartan and amlodipine achieved similar blood pressure control, losartan but not amlodipine showed a significant protective role against structural changes in the vessels and cavernous spaces of the erectile tissue caused by arterial hypertension. Copyright 2004 S. Karger AG, Basel

Mibefradil reduces blood glucose concentration in db/db mice

PubMed Central

Lu, Yujie; Long, Min; Zhou, Shiwen; Xu, Zihui; Hu, Fuquan; Li, Ming

2014-01-01

OBJECTIVE: Numerous recent studies suggest that abnormal intracellular calcium concentration ([Ca2+]i) is a common defect in diabetic animal models and patients. Abnormal calcium handling is an important mechanism in the defective pancreatic β-cell function in type 2 diabetes. T-type Ca2+ channel antagonists lower blood glucose in type 2 diabetes, but the mechanism remains unknown. METHODS: We examined the effect of the Ca2+ channel antagonist mibefradil on blood glucose in male db/db mice and phenotypically normal heterozygous mice by intraperitoneal injection. RESULTS: Mibefradil (15 mg/kg, i.p., b.i.d.) caused a profound reduction of fasting blood glucose from 430.92±20.46 mg/dl to 285.20±5.74 mg/dl in three days. The hypoglycemic effect of mibefradil was reproduced by NNC 55-0396, a compound structurally similar to mibefradil but more selective for T-type Ca2+ channels, but not by the specific L-type Ca2+ channel blocker nicardipine. Mibefradil did not show such hypoglycemic effects in heterozygous animals. In addition, triglycerides, basal insulin and food intake were significantly decreased by mibefradil treatment in the db/db mice but not in the controls. Western blot analysis, immunohistochemistry and immunofluorescence staining showed a significantly increased expression of T-type Ca2+ channel α-subunits Cav3.1 and Cav3.2 in liver and brain tissues from db/db mice compared to those from heterozygous animals. CONCLUSIONS: Collectively, these results suggest that T-type Ca2+ channels are potential therapeutic targets for antidiabetic drugs. PMID:24473561

Modulation of AMPA receptor mediated current by nicotinic acetylcholine receptor in layer I neurons of rat prefrontal cortex

PubMed Central

Tang, Bo; Luo, Dong; Yang, Jie; Xu, Xiao-Yan; Zhu, Bing-Lin; Wang, Xue-Feng; Yan, Zhen; Chen, Guo-Jun

2015-01-01

Layer I neurons in the prefrontal cortex (PFC) exhibit extensive synaptic connections with deep layer neurons, implying their important role in the neural circuit. Study demonstrates that activation of nicotinic acetylcholine receptors (nAChRs) increases excitatory neurotransmission in this layer. Here we found that nicotine selectively increased the amplitude of AMPA receptor (AMPAR)-mediated current and AMPA/NMDA ratio, while without effect on NMDA receptor-mediated current. The augmentation of AMPAR current by nicotine was inhibited by a selective α7-nAChR antagonist methyllycaconitine (MLA) and intracellular calcium chelator BAPTA. In addition, nicotinic effect on mEPSC or paired-pulse ratio was also prevented by MLA. Moreover, an enhanced inward rectification of AMPAR current by nicotine suggested a functional role of calcium permeable and GluA1 containing AMPAR. Consistently, nicotine enhancement of AMPAR current was inhibited by a selective calcium-permeable AMPAR inhibitor IEM-1460. Finally, the intracellular inclusion of synthetic peptide designed to block GluA1 subunit of AMPAR at CAMKII, PKC or PKA phosphorylation site, as well as corresponding kinase inhibitor, blocked nicotinic augmentation of AMPA/NMDA ratio. These results have revealed that nicotine increases AMPAR current by modulating the phosphorylation state of GluA1 which is dependent on α7-nAChR and intracellular calcium. PMID:26370265

Fullerenol C60(OH)24 nanoparticles decrease relaxing effects of dimethyl sulfoxide on rat uterus spontaneous contraction

NASA Astrophysics Data System (ADS)

Slavic, Marija; Djordjevic, Aleksandar; Radojicic, Ratko; Milovanovic, Slobodan; Orescanin-Dusic, Zorana; Rakocevic, Zlatko; Spasic, Mihajlo B.; Blagojevic, Dusko

2013-05-01

Dimethyl sulfoxide (DMSO) is a widely used solvent and cryoprotectant that can cause impaired blood flow, reduction in intracranial pressure, tissue edema, inflammatory reactions, inhibition of vascular smooth muscle cell migration and proliferation, processes which can lead to atherosclerosis of the coronary, peripheral and cerebral circulation. Although the adverse effects are rare when DMSO is administered in clinically established concentrations, there is no safe antagonist for an overdose. In this work, we treated isolated spontaneous and calcium-induced contractile active rat uteri (Wistar, virgo intacta), with DMSO and fullerenol C60(OH)24 nanoparticle (FNP) in DMSO. FNP is a water-soluble derivative of fullerene C60. Its size is a 1.1 nm in diameter and is a very promising candidate for a drug carrier in nanomedicine. FNP also displays free radical scavenging activity. DMSO decreased both spontaneous and calcium-induced contractions. In contrast, FNP only decreased spontaneous contraction. FNP decreased copper-zinc superoxide dismutase activity and prevented the DMSO-induced increase in glutathione reductase activity. Atomic force microscopy detected that FNP aggregated with calcium ions. Our results indicate that FNP has properties that make it a good candidate to be a modulator of DMSO activity which could minimize side effects of the latter.

Effects of NMDA antagonists on picrotoxin-, low Mg2+- and low Ca2+-induced epileptogenesis and on evoked changes in extracellular Na+ and Ca2+ concentrations in rat hippocampal slices.

PubMed

Köhr, G; Heinemann, U

1989-01-01

The anticonvulsant properties of ketamine and 2-APV were compared on 3 types of convulsant activity in hippocampal area CA1: the 'picrotoxin-epilepsy,' the 'low magnesium epilepsy' and the 'low calcium epilepsy.' In particular the spontaneous activity, the synaptically evoked responses and the changes in [Ca2+]0 were examined, since in many cases of epilepsy, Ca2+ uptake into cells is enhanced. In normal medium, ketamine and 2-APV have nearly no effect on stimulus evoked decreases in [Ca2+]0, although they clearly depress NMDA-induced ionic changes. However, ketamine and 2-APV prevent to some extent the augmentation of stimulus-induced changes in [Ca2+]0, observed after treating slices with picrotoxin or Mg2+-free medium. This extra Ca2+ uptake is probably mediated by NMDA operated channels. Our findings also show that ketamine, like 2-APV, has a stronger anticonvulsant effect on the low Mg-than on the picrotoxin-induced epileptiform activity. Responses to iontophoretically applied NMDA are facilitated in the 'low calcium epilepsy' and can be selectively blocked by ketamine. Spontaneous epileptiform activity occurring in low calcium can be blocked by ketamine only when some synaptic transmission is still present.

Noscapine protects OLN-93 oligodendrocytes from ischemia-reperfusion damage: Calcium and nitric oxide involvement.

PubMed

Nadjafi, S; Ebrahimi, S-A; Rahbar-Roshandel, N

2015-12-01

This study was carried out to evaluate the effects of noscapine, a benzylisoquinoline alkaloid from opium poppy, on oligodendrocyte during ischemia/reperfusion-induced excitotoxic injury. Changes in intracellular calcium levels due to chemical ischemia and nitric oxide (NO) production during ischemia/reperfusion were evaluated as the hallmarks of ischemia-derived excitotoxic event. OLN-93 cell line (a permanent immature rat oligodendrocyte) was used as a model of oligodendrocyte. 30- or 60-minute-oxygen-glucose deprivation/24 hours reperfusion were used to induce excitotoxicity. MTT (3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) assay was used to evaluate cell viability. Ratiometric fluorescence microscopy using Ca(2+)-sensitive indicator Fura-2/AM was utilized to assess intracellular calcium levels. NO production was evaluated by Griess method. Noscapine (4 μM) significantly attenuated intracellular Ca(2+) elevation (P < 0.001). Also, noscapine significantly decreased NO production during a 30-minute oxygen-glucose deprivation/reperfusion (P < 0.01). The inhibitory effect of noscapine (4 μM) on intracellular Ca(2+) was greater than ionotropic glutamate receptors antagonists. Noscapine is protective against ischemia/reperfusion-induced excitotoxic injury in OLN-93 oligodendrocyte. This protective effect seems to be related to attenuation of intracellular Ca(2+) overload and NO production.

Effects of intermittent pressure imitating rolling manipulation on calcium ion homeostasis in human skeletal muscle cells.

PubMed

Zhang, Hong; Liu, Howe; Lin, Qing; Zhang, Guohui; Mason, David C

2016-08-26

Homeostasis imbalance of intracellular Ca(2+) is one of the key pathophysiological factors in skeletal muscle injuries. Such imbalance can cause significant change in the metabolism of Ca(2+)-related biomarkers in skeletal muscle, such as superoxide dismutase (SOD), malondialdehyde (MDA) and creatine kinase (CK). Measurements of these biomarkers can be used to evaluate the degree of damage to human skeletal muscle cells (HSKMCs) injury. Rolling manipulation is the most popular myofascial release technique in Traditional Chinese Medicine. The mechanism of how this technique works in ameliorating muscle injury is unknown. This study aimed to investigate the possible Ca(2+) mediated effects of intermittent pressure imitating rolling manipulation (IPIRM) of Traditional Chinese Medicine in the injured HSKMCs. The normal HSKMCs was used as control normal group (CNG), while the injured HSKMCs were further divided into five different groups: control injured group (CIG), Rolling manipulation group (RMG), Rolling manipulation-Verapamil group (RMVG), static pressure group (SPG) and static pressure-Verapamil group (SPVG). RMG and RMVG cells were cyclically exposed to 9.5-12.5 N/cm(2) of IPIRM at a frequency of 1.0 Hz for 10 min. SPG and SPVG were loaded to a continuous pressure of 12.5 N/cm(2) for 10 min. Verapamil, a calcium antagonist, was added into the culture mediums of both RMVG and SPVG groups to block the influx of calcium ion. Compared with the CNG (normal cells), SOD activity was remarkably decreased while both MDA content and CK activity were significantly increased in the CIG (injured cells). When the injured cells were treated with the intermittent rolling manipulation pressure (RMG), the SOD activity was significantly increased and MDA content and CK activity were remarkably decreased. These effects were suppressed by adding the calcium antagonist Verapamil into the culture medium in RMVG. On the other hand, exposure to static pressure in SPG and SPVG affected neither the SOD activity nor the MDA content and CK activity in the injured muscle cells regardless of the presence of verapamil or not in the culture medium. These data suggest that the intermittent rolling pressure with the manipulation could ameliorate HSKMCs injury through a Ca(2+) dependent pathway. Static pressure did not lead to the same results.

Prostaglandin E2 Stimulates EP2, Adenylate Cyclase, Phospholipase C, and Intracellular Calcium Release to Mediate Cyclic Adenosine Monophosphate Production in Dental Pulp Cells.

PubMed

Chang, Mei-Chi; Lin, Szu-I; Lin, Li-Deh; Chan, Chiu-Po; Lee, Ming-Shu; Wang, Tong-Mei; Jeng, Po-Yuan; Yeung, Sin-Yuet; Jeng, Jiiang-Huei

2016-04-01

Prostaglandin E2 (PGE2) plays a crucial role in pulpal inflammation and repair. However, its induction of signal transduction pathways is not clear but is crucial for future control of pulpal inflammation. Primary dental pulp cells were exposed to PGE2 and 19R-OH PGE2 (EP2 agonist) or sulprostone (EP1/EP3 agonist) for 5 to 40 minutes. Cellular cyclic adenosine monophosphate (cAMP) levels were measured using the enzyme-linked immunosorbent assay. In some experiments, cells were pretreated with SQ22536 (adenylate cyclase inhibitor), H89 (protein kinase A inhibitor), dorsomorphin (adenosine monophosphate-activated protein kinase inhibitor), U73122 (phospholipase C inhibitor), thapsigargin (inhibitor of intracellular calcium release), W7 (calmodulin antagonist), verapamil (L-type calcium channel blocker), and EGTA (extracellular calcium chelator) for 20 minutes before the addition of PGE2. PGE2 and 19R-OH PGE2 (EP2 agonist) stimulated cAMP production, whereas sulprostone (EP1/EP3 agonist) shows little effect. PGE2-induced cAMP production was attenuated by SQ22536 and U73122 but not H89 and dorsomorphin. Intriguingly, thapsigargin and W7 prevented PGE2-induced cAMP production, but verapamil and EGTA showed little effect. These results indicate that PGE2-induced cAMP production is associated with EP2 receptor and adenylate cyclase activation. These events are mediated by phospholipase C, intracellular calcium release, and calcium-calmodulin signaling. These results are helpful for understanding the role of PGE2 in pulpal inflammation and repair and possible future drug intervention. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

The role of Rho-kinase and calcium ions in constriction triggered by ET-1.

PubMed

Wiciński, Michał; Szadujkis-Szadurska, Katarzyna; Węclewicz, Mateusz M; Malinowski, Bartosz; Matusiak, Grzegorz; Walczak, Maciej; Wódkiewicz, Eryk; Grześk, Grzegorz; Pawlak-Osińska, Katarzyna

2018-05-05

Endothelin-1 (ET-1) is one of the key factors regulating tension of smooth muscles in blood vessels. It is believed that ET-1 plays an important role in pathogenesis of hypertension, and cardiovascular diseases; therefore, research in order to limit ET-1-mediated action is still in progress. The main objective of this paper was to evaluate the role of Rho-kinase in the ET-1-induced constriction of arteries. The analysis also included significance of intra- and extracellular pool of calcium ions in constriction triggered by ET-1. The studies were performed on perfused Wistar rat tail arteries. Concentration response curve (CRC) was determined for ET-1 in the presence of increased concentrations of Rho-kinase inhibitor (Y-27632) and IP3-receptor antagonist (2APB), both in reference to constriction triggered by solely ET-1. Afterwards, the influence of calcium ions present in the perfusion fluid was evaluated in terms of the effect triggered by 2APB and occurring in arteries constricted by ET-1. ET-1, in concentration dependent manner, leads to increase in perfusion pressure. Y-27632 and 2APB lead to shift of the concentration response curve for ET-1 to the right with simultaneously lowered maximum effect. There was no difference in reaction of the artery constricted by ET-1 and treated with 2APB in solution containing calcium and in calcium-free solution. Vasoconstrictive action of endothelin is not significantly dependent on the inflow of extracellular calcium, but it is proportional to inflow of Ca 2+ related to activation of IP3 receptors and to Rho-kinase activity. Copyright © 2018. Published by Elsevier Inc.

[Lacidipine efficacy and safety for high blood pressure treatment in pediatric oncohematology].

PubMed

Bernard, E; Mialou, V; Dony, A; Garnier, N; Renard, C; Bleyzac, N

2014-10-01

In adults, lacidipine seems to have no CYP3A4-inhibiting action. This particular characteristic makes it advantageous when combined with drugs metabolized by CYP3A4, such as cyclosporine. Until now, no data on the efficacy or safety of this calcium antagonist have been available in children. Thirty-nine hypertensive children (age: 0.13-14 years) receiving lacidipine in oncohematology for a mean of 75 days were included in this retrospective study. The causes of high blood pressure were renal tumor (n=7), catecholamine-secreting tumor (n=4), corticoid treatment (n=5), and cyclosporine treatment (n=23). An initial dosage of 0.05 mg/kg/day was sufficient for 41% of the patients. The remaining patients needed to increase the dosage, by steps of 0.03 mg/kg/day, until reaching an average effective dosage of 0.1 mg/kg/day. Lacidipine significantly decreased blood pressure by 30 (±14) mmHg for systolic blood pressure and by 26 (±13) mmHg for diastolic blood pressure. A medication plan with twice-daily administration was not significantly more effective than a single administration per day. Lacidipine was well tolerated, and no toxicity-related withdrawal of treatment occurred. For 22 patients treated with both cyclosporine and lacidipine, renal function was not disturbed over time, suggesting its preservation by lacidipine. No significant increase in cyclosporine blood concentration was detected. Lacidipine seems to be an effective calcium antagonist in pediatric oncohematology, is well tolerated, has a kidney-protector effect and no drug interaction when combined with cyclosporine. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Enhancement by vasopressin of adrenergic responses in human mesenteric arteries.

PubMed

Medina, P; Noguera, I; Aldasoro, M; Vila, J M; Flor, B; Lluch, S

1997-03-01

Vasopressin not only acts directly on blood vessels through V1-receptor stimulation but also may modulate adrenergic-mediated responses in animal experiments in vitro and in vivo. The aim of the present study was to investigate whether subpressor concentrations of vasopressin could modify the constrictor responses to norepinephrine and electrical stimulation of the perivascular nerves in human mesenteric arteries. Human mesenteric artery rings (3-3.5 mm long, 0.8-1.2 mm OD) were obtained from 38 patients undergoing abdominal operations. The arterial rings were suspended in organ bath chambers for isometric recording of tension. Vasopressin (3 x 10(-11) M) enhanced the contractions elicited by electrical stimulation at 2, 4, and 8 Hz (by 100, 100, and 72%, respectively) and produced a leftward shift of the concentration-response curves to norepinephrine (half-maximal effective concentration decreased from 2.2 x 10(-6) to 5.0 x 10(-7) M; P < 0.05) without any alteration in maximal contractions. Vasopressin also potentiated KCl- and calcium-induced contractions. The V1-receptor antagonist 1-[beta-mercapto-beta,beta-cyclopentamethylenepropionic acid-2-O-methyl-tyrosine, 8-arginine]vasopressin (10(-6) M) prevented the potentiation evoked by vasopressin in all cases. The calcium antagonist nifedipine (10(-6) M) did not affect the potentiation of electrical stimulation and norepinephrine induced by vasopressin but abolished KCl-induced contractions. The results suggest that vasopressin, in addition to its direct vasoconstrictor effect, strongly potentiates the responses to adrenergic stimulation and KCl depolarization. Both the direct and indirect effects of vasopressin appear to be mediated by V1-receptor stimulation. The amplifying effect of vasopressin on constrictor responses may be relevant in those clinical situations characterized by increased plasma vasopressin levels.

Verapamil Blocks Scopolamine Enhancement Effect on Memory Consolidation in Passive Avoidance Task in Rats

PubMed Central

Giménez De Béjar, Verónica; Caballero Bleda, María; Popović, Natalija; Popović, Miroljub

2017-01-01

Our recent data have indicated that scopolamine, a non-selective muscarinic receptor antagonist, improves memory consolidation, in a passive avoidance task, tested in rats. It has been found that verapamil, a phenylalkylamine class of the L-type voltage-dependent calcium channel antagonist, inhibits [3H] N-methyl scopolamine binding to M1 muscarinic receptors. However, there are no data about the effect of verapamil on memory consolidation in the passive avoidance task, in rats. The purpose of the present study was to examine the effects of verapamil (0.5, 1.0, 2.5, 5.0, 10, or 20 mg/kg i.p.) as well as the interaction between scopolamine and verapamil on memory consolidation in the step-through passive avoidance task, in Wistar rats. Our results showed that verapamil (1.0 and 2.5 mg/kg) administered immediately after the acquisition task significantly increased the latency of the passive avoidance response, on the 48 h retested trial, improving memory consolidation. On the other hand, verapamil in a dose of 5 mg/kg, that per se does not affect memory consolidation, significantly reversed the memory consolidation improvement induced by scopolamine (1 mg/kg, i.p., administered immediately after verapamil treatment) but did not change the passive avoidance response in rats treated by an ineffective dose of scopolamine (30 mg/kg). In conclusion, the present data suggest that (1) the post-training administration of verapamil, dose-dependently, improves the passive avoidance response; (2) verapamil, in ineffective dose, abolished the improvement of memory consolidation effect of scopolamine; and (3) exists interaction between cholinergic muscarinic receptors and calcium homeostasis-related mechanisms in the consolidation of emotional memory. PMID:28878678

The effect of fixed oil and water extracts of Nigella sativa on sickle cells: an in vitro study.

PubMed

Ibraheem, N K; Ahmed, J H; Hassan, M K

2010-03-01

Various drugs have been investigated in the treatment of sickle cell disease (SCD), such as hydroxyurea, piracetam and calcium antagonists. Most of these drugs are potentially toxic and are not suitable for long-term therapy. Recently, Nigella sativa (NS) has been reported to have calcium antagonist and antioxidant activities, both of which play a role in the management of the disease. This study aimed to investigate the in vitro antisickling effect of extracts from NS. Thirty-two patients with SCD, aged 7-47 years old, were recruited for the study. A total of 3 ml of venous blood was collected from each patient and divided into six tubes with heparin. The blood was mixed with 0.5 ml of either 0.1 percent, 0.05 percent or 0.01 percent v/v of the oil extract of NS. A slide was prepared by spreading a drop of treated blood, covered with a cover slide to ensure the complete deoxygenation condition. The separation of irreversibly sickled cells (ISCs) was performed on eight patients by a density gradient (Percoll-Renografin) centrifugation method. The 0.1 percent v/v concentration of the oil extract of NS resulted in an approximately 80 percent reduction in the formation of sickle cells. The 0.05 percent v/v concentration of NS produced an intermediate effect, while the 0.01 percent v/v concentration had no effect on the formation of sickle cells. The 0.1 percent v/v concentration of the fixed oil of NS led to a considerable reduction in the formation of ISCs. The fixed oil extracted from NS seeds has an in vitro antisickling activity.

Nuclear calcium is required for human T cell activation

PubMed Central

Samstag, Yvonne

2016-01-01

Calcium signals in stimulated T cells are generally considered single entities that merely trigger immune responses, whereas costimulatory events specify the type of reaction. Here we show that the “T cell calcium signal” is a composite signal harboring two distinct components that antagonistically control genomic programs underlying the immune response. Using human T cells from healthy individuals, we establish nuclear calcium as a key signal in human T cell adaptogenomics that drives T cell activation and is required for signaling to cyclic adenosine monophosphate response element–binding protein and the induction of CD25, CD69, interleukin-2, and γ-interferon. In the absence of nuclear calcium signaling, cytosolic calcium activating nuclear factor of activated T cells translocation directed the genomic response toward enhanced expression of genes that negatively modulate T cell activation and are associated with a hyporesponsive state. Thus, nuclear calcium controls the T cell fate decision between a proliferative immune response and tolerance. Modulators of nuclear calcium–driven transcription may be used to develop a new type of pro-tolerance immunosuppressive therapy. PMID:27810914

Physiological and anatomical studies of the development of the sympathetic innervation to rat iris arterioles.

PubMed

Sandow; Hill

1999-09-24

The development of the sympathetic innervation to rat irideal arterioles has been investigated using histochemical and in vitro pharmacological and electrophysiological methods. A plexus of fibres and varicosities appeared over the surface of the vessels after the first postnatal week and increased to reach a maximum density during the fourth postnatal week. Transmural nerve stimulation produced small, consistent contractions that were first recorded in arterioles of 7-day old rats. Contractions became larger and faster, reaching the adult form during the fourth postnatal week. Contractions became more sensitive to the alpha1-adrenoceptor antagonists, prazosin and naftopidil, and less sensitive to the alpha1A/D antagonist, WB4101 and alpha2 antagonist, yohimbine, during development. At both 10 and 21 days, contractile responses resulted from the release of intracellular calcium as they were abolished by caffeine (10(-3) M), thapsigargin (2 x 10(-6) M) and cyclopiazonic acid (3 x 10(-6) M), but not by nifedipine (10(-6) M). Intracellular recordings showed that nerve stimulation produced large, slow depolarizations at all ages tested. Time to peak potential decreased during development, while the amplitude of the depolarizations did not vary significantly. Results suggest that, throughout development, sympathetic nerves cause constriction of iris arterioles due to the release of noradrenaline and activation of alpha-adrenoceptors on the smooth muscle cells. Early responses involved both alpha1- and alpha2-adrenoceptors, while later responses were due to alpha1-adrenoceptors only. Irrespective of these changes in adrenoceptor subtypes, smooth muscle contraction resulted from the mobilization of intracellular calcium suggesting that both alpha1- and alpha2-adrenoceptors were coupled to pathways which accessed this source of calcium.

Glycine Receptor Activation Impairs ATP-Induced Calcium Transients in Cultured Cortical Astrocytes

PubMed Central

Morais, Tatiana P.; Coelho, David; Vaz, Sandra H.; Sebastião, Ana M.; Valente, Cláudia A.

2018-01-01

In central nervous system, glycine receptor (GlyR) is mostly expressed in the spinal cord and brainstem, but glycinergic transmission related elements have also been identified in the brain. Astrocytes are active elements at the tripartite synapse, being responsible for the maintenance of brain homeostasis and for the fine-tuning of synaptic activity. These cells communicate, spontaneously or in response to a stimulus, by elevations in their cytosolic calcium (calcium transients, Ca2+T) that can be propagated to other cells. How these Ca2+T are negatively modulated is yet poorly understood. In this work, we evaluated GlyR expression and its role on calcium signaling modulation in rat brain astrocytes. We first proved that GlyR, predominantly subunits α2 and β, was expressed in brain astrocytes and its localization was confirmed in the cytoplasm and astrocytic processes by immunohistochemistry assays. Calcium imaging experiments in cultured astrocytes showed that glycine (500 μM), a GlyR agonist, caused a concentration-dependent reduction in ATP-induced Ca2+T, an effect abolished by the GlyR antagonist, strychnine (0.8 μM), as well as by nocodazole (1 μM), known to impair GlyR anchorage to the plasma membrane. This effect was mimicked by activation of GABAAR, another Cl--permeable channel. In summary, we demonstrated that GlyR activation in astrocytes mediates an inhibitory effect upon ATP induced Ca2+T, which most probably involves changes in membrane permeability to Cl- and requires GlyR anchorage at the plasma membrane. GlyR in astrocytes may thus be part of a mechanism to modulate astrocyte-to-neuron communication. PMID:29386993

Mercaptoacetate and fatty acids exert direct and antagonistic effects on nodose neurons via GPR40 fatty acid receptors.

PubMed

Darling, Rebecca A; Zhao, Huan; Kinch, Dallas; Li, Ai-Jun; Simasko, Steven M; Ritter, Sue

2014-07-01

β-mercaptoacetate (MA) is a drug known to block mitochondrial oxidation of medium- and long-chain fatty acids (FAs) and to stimulate feeding. Because MA-induced feeding is vagally dependent, it has been assumed that the feeding response is mediated by MA's antimetabolic action at a peripheral, vagally innervated site. However, MA's site of action has not yet been identified. Therefore, we used fluorescent calcium measurements in isolated neurons from rat nodose ganglia to determine whether MA has direct effects on vagal sensory neurons. We found that MA alone did not alter cytosolic calcium concentrations in nodose neurons. However, MA (60 μM to 6 mM) significantly decreased calcium responses to both linoleic acid (LA; 10 μM) and caprylic acid (C8; 10 μM) in all neurons responsive to LA and C8. GW9508 (40 μM), an agonist of the FA receptor, G protein-coupled receptor 40 (GPR40), also increased calcium levels almost exclusively in FA-responsive neurons. MA significantly inhibited this response to GW9508. MA did not inhibit calcium responses to serotonin, high K(+), or capsaicin, which do not utilize GPRs, or to CCK, which acts on a different GPR. GPR40 was detected in nodose ganglia by RT-PCR. Results suggest that FAs directly activate vagal sensory neurons via GPR40 and that MA antagonizes this effect. Thus, we propose that MA's nonmetabolic actions on GPR40 membrane receptors, expressed by multiple peripheral tissues in addition to the vagus nerve, may contribute to or mediate MA-induced stimulation of feeding. Copyright © 2014 the American Physiological Society.

Butyl benzyl phthalate suppresses the ATP-induced cell proliferation in human osteosarcoma HOS cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, P.-S., E-mail: pslediting@mail.scu.edu.t; Chen, C.-Y.

2010-05-01

Butyl benzyl phthalate (BBP), an endocrine disruptor present in the environment, exerts its genomic effects via intracellular steroid receptors and elicits non-genomic effects by interfering with membrane ion-channel receptors. We previously found that BBP blocks the calcium signaling coupled with P2X receptors in PC12 cells (Liu and Chen, 2006). Osteoblast P2X receptors were recently reported to play a role in cell proliferation and bone remodeling. In this present study, the effects of BBP on ATP-induced responses were investigated in human osteosarcoma HOS cells. These receptors mRNA had been detected, named P2X4, P2X7, P2Y2, P2Y4, P2Y5, P2Y9, and P2Y11, in humanmore » osteosarcoma HOS cells by RT-PCR. The enhancement of cell proliferation and the decrease of cytoviability had both been shown to be coupled to stimulation via different concentrations of ATP. BBP suppressed the ATP-induced calcium influx (mainly coupled with P2X) and cell proliferation but not the ATP-induced intracellular calcium release (mainly coupled with P2Y) and cytotoxicity in human osteosarcoma HOS cells. Suramin, a common P2 receptor's antagonist, blocked the ATP-induced calcium signaling, cell proliferation, and cytotoxicity. We suggest that P2X is mainly responsible for cell proliferation, and P2Y might be partially responsible for the observed cytotoxicity. BBP suppressed the calcium signaling coupled with P2X, suppressing cell proliferation. Since the importance of P2X receptors during bone metastasis has recently become apparent, the possible toxic risk of environmental BBP during bone remodeling is a public problem of concern.« less

Radioprotection of Intestinal Stem Cells and Whole Body Radiation Lethality from Photons and Neutrons by Prostaglandins along or in Combination with WR-2721

DTIC Science & Technology

1990-12-01

asthma, calcium metabolic defects, angina pectoris and peptic ulcer . As a result of the array and sometimes antagonistic biologic effects of the family of...Arlington, VA 22202-4302, and to the Office of Management and Budget. Paperwork Reduction Project (0704-0188). Washington. OC 20503 1, AGENCY USE ONLY (Leave... perforated plastic tubes that, in turn, were placed between two A-150 tissue equivalent disks for build-up and backscatter. The disks holding the mice were

Calcium plus vitamin D supplementation and lung cancer incidence among postmenopausal women in the Women's Health Initiative.

PubMed

Tao, Meng-Hua; Dai, Qi; Chen, Shande; Freudenheim, Jo L; Rohan, Thomas; Wakelee, Heather; Datta, Mridul; Wactawski-Wende, Jean

2017-08-01

Magnesium and calcium are antagonistic in many physiologic processes. However, few studies have investigated the associations of supplemental calcium with lung cancer risk taking this antagonism into account. We evaluated the effect of calcium and vitamin D supplementation on lung cancer incidence and explored whether the ratio of baseline calcium to magnesium (Ca:Mg) intake modifies the association in the Women's Health Initiative (WHI) calcium plus vitamin D supplementation (CaD) trial. The intervention phase of the WHI CaD was a double-blinded, randomized, placebo-controlled trial in 36,382 postmenopausal women aged 50-79 years, recruited at 40U.S. centers. Post-intervention follow-up continued among 29,862 (86%) of the surviving participants. Risk of lung cancer in association with CaD supplementation was evaluated using proportional hazard regression models. After 11 years' cumulative follow-up, there were 207 lung cancers (incidence 0.11% per year) in the supplement arm and 241 (0.12%) in the placebo arm (hazard ratio (HR) for the intervention, 0.91; 95% confidence interval (CI), 0.71-1.17). Subgroup analyses suggested that the HR for lung cancer varied by baseline Ca:Mg intake ratio among women who were current smokers at enrollment (p=0.04 for interaction). Over the entire follow-up period, calcium and vitamin D supplementation did not reduce lung cancer incidence among postmenopausal women. In exploratory analyses, an interaction was found for the baseline Ca:Mg intake ratio on lung cancer among current smokers at the trial entry. This findings need to be further studied for the role of calcium with magnesium in lung carcinogenesis in current smokers. Copyright © 2017 Elsevier B.V. All rights reserved.

Histamine receptors in human detrusor smooth muscle cells: physiological properties and immunohistochemical representation of subtypes.

PubMed

Neuhaus, Jochen; Weimann, Annett; Stolzenburg, Jens-Uwe; Dawood, Waled; Schwalenberg, Thilo; Dorschner, Wolfgang

2006-06-01

The potent inflammatory mediator histamine is released from activated mast cells in interstitial cystitis (IC). Here, we report on the histamine receptor subtypes involved in the intracellular calcium response of cultured smooth muscle cells (cSMC). Fura-2 was used to monitor the calcium response in cSMC, cultured from human detrusor biopsies. The distribution of histamine receptor subtypes was addressed by immunocytochemistry in situ and in vitro. Histamine stimulated a maximum of 92% of the cells (n=335), being more effective than carbachol (70%, n=920). HTMT (H1R-agonist), dimaprit (H2R) and MTH (H3R) lead to significant lower numbers of reacting cells (60, 48 and 54%). Histamine receptor immunoreactivity (H1R, H2R, H3R, H4R) was found in situ and in vitro. Histamine-induced calcium increase is mediated by distinct histamine receptors. Thus, pre-therapeutic evaluation of histamine receptor expression in IC patients may help to optimize therapy by using a patient-specific cocktail of subtype-specific histamine receptor antagonists.

Real-time Recording of Cytosolic Calcium Levels in Arabidopsis thaliana Cell Cultures during Parabolic Flights

NASA Astrophysics Data System (ADS)

Neef, Maren; Ecke, Margret; Hampp, Rüdiger

2015-07-01

In plants, like in other organisms, calcium (Ca2+) is an important second messenger which participates in the conversion of environmental signals into molecular responses. There is increasing evidence, that sensing of changes in gravitation or reorientation of tissues is an example for such signaling cascades in which Ca2+ is involved. In order to determine g-dependent changes in the cytosolic calcium (Ca^{2+}_{ {cyt}}) concentration of plant cells, semisolid transgenic callus cell cultures of Arabidopsis thaliana (A.t.), expressing the calcium sensor YC3.6 (cameleon), were exposed to g-forces between 1.8 g and μ g during parabolic flights. Using such cells, intracellular calcium transients can be monitored by FRET in vivo and in real-time. Interestingly we observed a slight decrease of the Ca^{2+}_{ {cyt}} level during the hypergravity phases of a parabola but a significant increase of the Ca^{2+}_{ {cyt}} concentration during microgravity. Application of known Ca2+ inhibitors and antagonists yielded the following effects: nifedipine (Ca2+ channel blocker) showed no effect, whereas LaCl3, GdCl3 (both inhibitors of uptake at the plasma membrane), DPI (inhibitor of NADP oxidase), and DMSO (solvent) diminished the gravity-alteration-related Ca^{2+}_{ {cyt}} response. EGTA (binding of Ca2+) and eosin yellow (inhibitor of a plasma membrane-located Ca2+ pump) suppressed the respective Ca^{2+}_{ {cyt}} changes entirely. We thus conclude that the significant increase in Ca^{2+}_{ {cyt}} under microgravity is largely due to extracellular Ca2+ sources.

Nimodipine, an L-type calcium channel blocker attenuates mitochondrial dysfunctions to protect against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinsonism in mice.

PubMed

Singh, Alpana; Verma, Poonam; Balaji, Gillela; Samantaray, Supriti; Mohanakumar, Kochupurackal P

2016-10-01

Parkinson's disease (PD), the most common progressive neurodegenerative movement disorder, results from loss of dopaminergic neurons of substantia nigra pars compacta. These neurons exhibit Cav1.3 channel-dependent pacemaking activity. Epidemiological studies suggest reduced risk for PD in population under long-term antihypertensive therapy with L-type calcium channel antagonists. These prompted us to investigate nimodipine, an L-type calcium channel blocker for neuroprotective effect in cellular and animal models of PD. Nimodipine (0.1-10 μM) significantly attenuated 1-methyl-4-phenyl pyridinium ion-induced loss in mitochondrial morphology, mitochondrial membrane potential and increases in intracellular calcium levels in SH-SY5Y neuroblastoma cell line as measured respectively employing Mitotracker green staining, TMRM, and Fura-2 fluorescence, but only a feeble neuroprotective effect was observed in MTT assay. Nimodipine dose-dependently reduced 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced parkinsonian syndromes (akinesia and catalepsy) and loss in swimming ability in Balb/c mice. It attenuated MPTP-induced loss of dopaminergic tyrosine hydroxylase positive neurons in substantia nigra, improved mitochondrial oxygen consumption and inhibited reactive oxygen species production in the striatal mitochondria measured using dichlorodihydrofluorescein fluorescence, but failed to block striatal dopamine depletion. These results point to an involvement of L-type calcium channels in MPTP-induced dopaminergic neuronal death in experimental parkinsonism and more importantly provide evidences for nimodipine to improve mitochondrial integrity and function. Copyright © 2016 Elsevier Ltd. All rights reserved.

The use of antioxidants to prevent glutamate-induced derangement of calcium ion metabolism in rat cerebral cortex synaptosomes.

PubMed

Avrova, N F; Shestak, K I; Zakharova, I O; Sokolova, T V; Tyurina, Y Y; Tyurin, V A

2000-01-01

Glutamate is shown to induce increases in intracellular Ca2+ concentrations ([Ca2+]i), increases in 45Ca2+ influx, decreases in the activity of Na+,K+-ATPase activity, and activation of the Na+/Ca2+ exchanger in rat cerebral cortex synaptosomes. NMDA receptor antagonists virtually prevented these effects. Preincubation of synaptosomes with alpha-tocopherol, superoxide dismutase, and ganglioside GM1 normalized [Ca2+]i, 45Ca2+ influx, and Na+,K+-ATPase activity in rat cerebral cortex synaptosomes exposed to glutamate. Glutamate and GM1 activated the Na+/K+ exchanger, and their effects were additive. Calcium ions entering cerebral cortex nerve cells via NMDA receptors during exposure to high glutamate concentrations appeared to be only the trigger for the processes activating free-radical reactions. Activation of these reactions led to increases in Ca2+ influx into cells, decreases in Na+,K+-ATPase activity, and significant increases in [Ca2+]i, though this could be prevented by antioxidants and gangliosides.

Assay of Calcium Transients and Synapses in Rat Hippocampal Neurons by Kinetic Image Cytometry and High-Content Analysis: An In Vitro Model System for Postchemotherapy Cognitive Impairment.

PubMed

McDonough, Patrick M; Prigozhina, Natalie L; Basa, Ranor C B; Price, Jeffrey H

2017-07-01

Postchemotherapy cognitive impairment (PCCI) is commonly exhibited by cancer patients treated with a variety of chemotherapeutic agents, including the endocrine disruptor tamoxifen (TAM). The etiology of PCCI is poorly understood. Our goal was to develop high-throughput assay methods to test the effects of chemicals on neuronal function applicable to PCCI. Rat hippocampal neurons (RHNs) were plated in 96- or 384-well dishes and exposed to test compounds (forskolin [FSK], 17β-estradiol [ES]), TAM or fulvestrant [FUL], aka ICI 182,780) for 6-14 days. Kinetic Image Cytometry™ (KIC™) methods were developed to quantify spontaneously occurring intracellular calcium transients representing the activity of the neurons, and high-content analysis (HCA) methods were developed to quantify the expression, colocalization, and puncta formed by synaptic proteins (postsynaptic density protein-95 [PSD-95] and presynaptic protein Synapsin-1 [Syn-1]). As quantified by KIC, FSK increased the occurrence and synchronization of the calcium transients indicating stimulatory effects on RHN activity, whereas TAM had inhibitory effects. As quantified by HCA, FSK also increased PSD-95 puncta and PSD-95:Syn-1 colocalization, whereas ES increased the puncta of both PSD-95 and Syn-1 with little effect on colocalization. The estrogen receptor antagonist FUL also increased PSD-95 puncta. In contrast, TAM reduced Syn-1 and PSD-95:Syn-1 colocalization, consistent with its inhibitory effects on the calcium transients. Thus TAM reduced activity and synapse formation by the RHNs, which may relate to the ability of this agent to cause PCCI. The results illustrate that KIC and HCA can be used to quantify neurotoxic and neuroprotective effects of chemicals in RHNs to investigate mechanisms and potential therapeutics for PCCI.

Osteoclast cytosolic calcium, regulated by voltage-gated calcium channels and extracellular calcium, controls podosome assembly and bone resorption

NASA Technical Reports Server (NTRS)

Miyauchi, A.; Hruska, K. A.; Greenfield, E. M.; Duncan, R.; Alvarez, J.; Barattolo, R.; Colucci, S.; Zambonin-Zallone, A.; Teitelbaum, S. L.; Teti, A.

1990-01-01

The mechanisms of Ca2+ entry and their effects on cell function were investigated in cultured chicken osteoclasts and putative osteoclasts produced by fusion of mononuclear cell precursors. Voltage-gated Ca2+ channels (VGCC) were detected by the effects of membrane depolarization with K+, BAY K 8644, and dihydropyridine antagonists. K+ produced dose-dependent increases of cytosolic calcium ([Ca2+]i) in osteoclasts on glass coverslips. Half-maximal effects were achieved at 70 mM K+. The effects of K+ were completely inhibited by dihydropyridine derivative Ca2+ channel blocking agents. BAY K 8644 (5 X 10(-6) M), a VGCC agonist, stimulated Ca2+ entry which was inhibited by nicardipine. VGCCs were inactivated by the attachment of osteoclasts to bone, indicating a rapid phenotypic change in Ca2+ entry mechanisms associated with adhesion of osteoclasts to their resorption substrate. Increasing extracellular Ca2+ ([Ca2+]e) induced Ca2+ release from intracellular stores and Ca2+ influx. The Ca2+ release was blocked by dantrolene (10(-5) M), and the influx by La3+. The effects of [Ca2+]e on [Ca2+]i suggests the presence of a Ca2+ receptor on the osteoclast cell membrane that could be coupled to mechanisms regulating cell function. Expression of the [Ca2+]e effect on [Ca2+]i was similar in the presence or absence of bone matrix substrate. Each of the mechanisms producing increases in [Ca2+]i, (membrane depolarization, BAY K 8644, and [Ca2+]e) reduced expression of the osteoclast-specific adhesion structure, the podosome. The decrease in podosome expression was mirrored by a 50% decrease in bone resorptive activity. Thus, stimulated increases of osteoclast [Ca2+]i lead to cytoskeletal changes affecting cell adhesion and decreasing bone resorptive activity.

Inotropic effects of diadenosine tetraphosphate (AP4A) in human and animal cardiac preparations.

PubMed

Vahlensieck, U; Bokník, P; Gombosová, I; Huke, S; Knapp, J; Linck, B; Lüss, H; Müller, F U; Neumann, J; Deng, M C; Scheld, H H; Jankowski, H; Schlüter, H; Zidek, W; Zimmermann, N; Schmitz, W

1999-02-01

Diadenosine tetraphosphate (AP4A) is an endogenous compound and exerts diverse physiological effects in animal systems. However, the effects of AP4A on inotropy in ventricular cardiac preparations have not yet been studied. The effects of AP4A on force of contraction (FOC) were studied in isolated electrically driven guinea pig and human cardiac preparations. Furthermore, the effects of AP4A on L-type calcium current and [Ca]i were studied in isolated guinea pig ventricular myocytes. In guinea pig left atria, AP4A (0.1-100 microM) reduced FOC maximally by 36.5 +/- 4.3%. In guinea pig papillary muscles, AP4A (100 microM) alone was ineffective, but reduced isoproterenol-stimulated FOC maximally by 29.3 +/- 3.4%. The negative inotropic effects of AP4A in atria and papillary muscles were abolished by the A1-adenosine receptor antagonist 1, 3-dipropyl-cyclopentylxanthine. In guinea pig ventricular myocytes, AP4A (100 microM) attenuated isoproterenol-stimulated L-type calcium current and [Ca]i. In human atrial and ventricular preparations, AP4A (100 microM) alone increased FOC to 158.3 +/- 12.4% and 167.5 +/- 25.1%, respectively. These positive inotropic effects were abolished by the P2-purinoceptor antagonist suramin. On the other hand, AP4A (100 microM) reduced FOC by 27.2 +/- 7.4% in isoproterenol-stimulated human ventricular trabeculae. The latter effect was abolished by 1,3-dipropyl-cyclopentylxanthine. In summary, after beta adrenergic stimulation AP4A exerts negative inotropic effects in animal and human ventricular preparations via stimulation of A1-adenosine receptors. In contrast, AP4A alone can exert positive inotropic effects via P2-purinoceptors in human ventricular myocardium. Thus, P2-purinoceptor stimulation might be a new positive inotropic principle in the human myocardium.

Diffusional dynamics of an active rhodamine-labeled 1,4-dihydropyridine in sarcolemmal lipid multibilayers.

PubMed Central

Mason, R P; Chester, D W

1989-01-01

A "membrane bilayer pathway" model, involving ligand partition into the bilayer, lateral diffusion, and receptor binding has been invoked to describe the 1,4-dihydropyridine (DHP) calcium channel antagonist receptor binding mechanism. In an earlier study (Chester et al. 1987. Biophys. J. 52:1021-1030), the diffusional component of this model was examined using an active fluorescence labeled DHP calcium channel antagonist, nisoldipine-lissamine rhodamine B (Ns-R), in purified cardiac sarcolemmal (CSL) lipid multibilayers. Diffusion coefficient measurements on membrane-bound drug and phospholipid at maximum bilayer hydration yielded similar values (3.8 x 10(-8) cm2/s). However, decreases in bilayer hydration resulted in dramatically reduced diffusion coefficient values for both probes with substantially greater impact on Ns-R diffusion. These data suggested that hydration dependent diffusional differences could be a function of relative probe location along the bilayer normal. In this communication, we have addressed the relative effect of the rhodamine substituent on Ns-R diffusion complex by examining the diffusional dynamics of free rhodamine B under the same conditions used to evaluate Ns-R complex and phospholipid diffusion. X-ray diffraction studies were performed to determine the Ns-R location in the membrane and model the CSL lipid bilayer profile structure to give a rationale for the differences in probe diffusional dynamics as a function of interbilayer water space. PMID:2611332

Calcium overloading in traumatic axonal injury by lateral head rotation: a morphological evidence in rat model.

PubMed

He, Xiao-Sheng; Xiang, Zhang; Zhou, Fei; Fu, Luo-An; Shuang, Wang

2004-05-01

The study investigated morphologically axonal calcium overloading and its relationship with axonal structural changes. Twelve SD rats were divided into an injury and a sham group. The rat model of traumatic axonal injury (TAI) by lateral head rotation was produced. The oxalate-pyroantimonate technique for calcium localization was used to process the rat's medulla oblongata tissues with thin sections observed electron-microscopically for axonal structure and calcium precipitates on it. The axonal damage in medulla oblongata appeared at 2 h post-injury, gradually became diffuse and severe, and continued to exist at 24 hours. At 2 hours, calcium precipitates were deposited on separated lamellae and axolemma, but were rarely distributed in the axoplasm. At 6 hours, calcium precipitates occurred on separated lamellae and axolemma in much higher density, but on axoplasm in extremely small amounts. Some axons, though lacking structural changes of the myelin sheath, sequestered plenty of calcium deposits on their swollen mitochondria. At 24 hours, damaged axons presented with much more severe lamellae separation and calcium deposits. Axonal calcium overloading developed in rat TAI model using lateral head rotation. This was significantly related to structural damage in the axons. These findings suggest the feasibility of using calcium antagonists in cope the management of human DAI in its very early stage.

Action of cholecystokinin and cholinergic agents on calcium transport in isolated pancreatic acinar cells.

PubMed Central

Gardner, J D; Conlon, T P; Kleveman, H L; Adams, T D; Ondetti, M A

1975-01-01

COOH-terminal octapeptide of cholecystokinin (CCK-octapeptide) and the cholinergic agent carbamylcholine each produced a fourfold stimulation of calcium outflux in guinea pig isolated pancreatic acinar cells. Neither agent altered calcium influx. Stimulation of calcium outflux was rapid and specific, was abolished by reducing the incubation temperature to 4 degrees C, and was a saturable function of the secretagogue concentration. The concentrations of CCK-octapeptide and carbamylcholine that produced half-maximal stimulation of calcium outflux were 3.1 x 10(-10) M and 4.9 x 10(-5) M, respectively. The cholinergic antagonist antropine competitively inhibited carbamylcholine stimulation of calcium outflux but did not alter stimulation produced by CCK-octapeptide. Stimulation of calcium outflux by maximal concentrations of carbamycholine plus CCK-octapeptide was the same as that produced by a maximal concentration of either agent alone.Calcium outflux became refractory to stimulation by secretagogues, and incubation with either CCK-ostapeptide or carbamylcholine produced a refractoriness to both agents. The relative potencies with CCK and its related fragments stimulated calcium outflux were CCK-octapeptide greater than heptapeptide greater than CCK greater than hexapeptide = gastrin. Secretin, glucagon, and vasoactive intestinal peptide, at concentrations as high as 10(-5) M, failed to alter calcium outflux and did not affect stimulation by CCK-octapeptide or by carbamycholine. Images PMID:1150877

N-arachidonoyl--serine is neuroprotective after traumatic brain injury by reducing apoptosis

PubMed Central

Cohen-Yeshurun, Ayelet; Trembovler, Victoria; Alexandrovich, Alexander; Ryberg, Erik; Greasley, Peter J; Mechoulam, Raphael; Shohami, Esther; Leker, Ronen R

2011-01-01

N-arachidonoyl--serine (AraS) is a brain component structurally related to the endocannabinoid family. We investigated the neuroprotective effects of AraS following closed head injury induced by weight drop onto the exposed fronto-parietal skull and the mechanisms involved. A single injection of AraS following injury led to a significant improvement in functional outcome, and to reduced edema and lesion volume compared with vehicle. Specific antagonists to CB2 receptors, transient receptor potential vanilloid 1 (TRPV1) or large conductance calcium-activated potassium (BK) channels reversed these effects. Specific binding assays did not indicate binding of AraS to the GPR55 cannabinoid receptor. N-arachidonoyl--serine blocked the attenuation in phosphorylated extracellular-signal-regulated kinase 1/2 (ERK) levels and led to an increase in pAkt in both the ipsilateral and contralateral cortices. Increased levels of the prosurvival factor Bcl-xL were evident 24 hours after injury in AraS-treated mice, followed by a 30% reduction in caspase-3 activity, measured 3 days after injury. Treatment with a CB2 antagonist, but not with a CB1 antagonist, reversed this effect. Our results suggest that administration of AraS leads to neuroprotection via ERK and Akt phosphorylation and induction of their downstream antiapoptotic pathways. These protective effects are related mostly to indirect signaling via the CB2R and TRPV1 channels but not through CB1 or GPR55 receptors. PMID:21505478

N-arachidonoyl-L-serine is neuroprotective after traumatic brain injury by reducing apoptosis.

PubMed

Cohen-Yeshurun, Ayelet; Trembovler, Victoria; Alexandrovich, Alexander; Ryberg, Erik; Greasley, Peter J; Mechoulam, Raphael; Shohami, Esther; Leker, Ronen R

2011-08-01

N-arachidonoyl-L-serine (AraS) is a brain component structurally related to the endocannabinoid family. We investigated the neuroprotective effects of AraS following closed head injury induced by weight drop onto the exposed fronto-parietal skull and the mechanisms involved. A single injection of AraS following injury led to a significant improvement in functional outcome, and to reduced edema and lesion volume compared with vehicle. Specific antagonists to CB2 receptors, transient receptor potential vanilloid 1 (TRPV1) or large conductance calcium-activated potassium (BK) channels reversed these effects. Specific binding assays did not indicate binding of AraS to the GPR55 cannabinoid receptor. N-arachidonoyl-L-serine blocked the attenuation in phosphorylated extracellular-signal-regulated kinase 1/2 (ERK) levels and led to an increase in pAkt in both the ipsilateral and contralateral cortices. Increased levels of the prosurvival factor Bcl-xL were evident 24 hours after injury in AraS-treated mice, followed by a 30% reduction in caspase-3 activity, measured 3 days after injury. Treatment with a CB2 antagonist, but not with a CB1 antagonist, reversed this effect. Our results suggest that administration of AraS leads to neuroprotection via ERK and Akt phosphorylation and induction of their downstream antiapoptotic pathways. These protective effects are related mostly to indirect signaling via the CB2R and TRPV1 channels but not through CB1 or GPR55 receptors.

The Antidepressant 5-HT2A Receptor Antagonists Pizotifen and Cyproheptadine Inhibit Serotonin-Enhanced Platelet Function

PubMed Central

Lin, Olivia A.; Karim, Zubair A.; Vemana, Hari Priya; Espinosa, Enma V. P.; Khasawneh, Fadi T.

2014-01-01

There is considerable interest in defining new agents or targets for antithrombotic purposes. The 5-HT2A receptor is a G-protein coupled receptor (GPCR) expressed on many cell types, and a known therapeutic target for many disease states. This serotonin receptor is also known to regulate platelet function. Thus, in our FDA-approved drug repurposing efforts, we investigated the antiplatelet activity of cyproheptadine and pizotifen, two antidepressant 5-HT2A Receptor antagonists. Our results revealed that cyproheptadine and pizotifen reversed serotonin-enhanced ADP-induced platelet aggregation in vitro and ex vivo. And the inhibitory effects of these two agents were found to be similar to that of EMD 281014, a 5-HT2A Receptor antagonist under development. In separate experiments, our studies revealed that these 5-HT2A receptor antagonists have the capacity to reduce serotonin-enhanced ADP-induced elevation in intracellular calcium levels and tyrosine phosphorylation. Using flow cytometry, we also observed that cyproheptadine, pizotifen, and EMD 281014 inhibited serotonin-enhanced ADP-induced phosphatidylserine (PS) exposure, P-selectin expression, and glycoprotein IIb-IIIa activation. Furthermore, using a carotid artery thrombosis model, these agents prolonged the time for thrombotic occlusion in mice in vivo. Finally, the tail-bleeding time was investigated to assess the effect of cyproheptadine and pizotifen on hemostasis. Our findings indicated prolonged bleeding time in both cyproheptadine- and pizotifen-treated mice. Notably, the increases in occlusion and bleeding times associated with these two agents were comparable to that of EMD 281014, and to clopidogrel, a commonly used antiplatelet drug, again, in a fashion comparable to clopidogrel and EMD 281014. Collectively, our data indicate that the antidepressant 5-HT2A antagonists, cyproheptadine and pizotifen do exert antiplatelet and thromboprotective effects, but similar to clopidogrel and EMD 281014, their use may interfere with normal hemostasis. PMID:24466319

Neuropeptide S receptor ligands: a patent review (2005-2016).

PubMed

Ruzza, Chiara; Calò, Girolamo; Di Maro, Salvatore; Pacifico, Salvatore; Trapella, Claudio; Salvadori, Severo; Preti, Delia; Guerrini, Remo

2017-03-01

Neuropeptide S (NPS) is a 20-residue peptide and endogenous ligand of the NPS receptor (NPSR). This receptor was a formerly orphan GPCR whose activation increases calcium and cyclic adenosine monophosphate levels. The NPS/NPSR system is expressed in several brain regions where it controls important biological functions including locomotor activity, arousal and sleep, anxiety, food intake, memory, pain, and drug addiction. Areas covered: This review furnishes an updated overview of the patent literature covering NPSR ligands since 2005, when the first example of an NPSR antagonist was disclosed. Expert opinion: Several potent NPSR antagonists are available as valuable pharmacological tools despite showing suboptimal pharmacokinetic properties in vivo. The optimization of these ligands is needed to speed up their potential clinical advancement as pharmaceuticals to treat drug addiction. In order to support the design of novel NPSR antagonists, we performed a ligand-based conformational analysis recognizing some structural requirements for NPSR antagonism. The identification of small-molecule NPSR agonists now represents an unmet challenge to be addressed. These molecules will allow investigation of the beneficial effects of selective NPSR activation in a large panel of psychiatric disorders and to foresee their therapeutic potential as anxiolytics, nootropics, and analgesics.

Myo-Inositol trisphosphate mobilizes calcium from fusogenic carrot (Daucus carota L. ) protoplasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rincon, M.; Boss, W.F.

1987-02-01

To determine whether or not inositol trisphosphate (IP/sub 3/) mobilizes calcium in higher plant cells; they investigated the effect of IP/sub 3/ on Ca/sup 2 +/ fluxes in fusogenic carrot (Daucus carota L.) protoplasts. The protoplasts were incubated in /sup 45/Ca/sup 2 +/-containing medium and the /sup 45/Ca/sup 2 +/ associated with the protoplasts was monitored with time. Addition of IP/sub 3/ (20 micromolar) caused a 17% net loss of the accumulated /sup 45/Ca/sup 2 +/ within 4 minutes. There was a reuptake of /sup 45/Ca/sup 2 +/ and the protoplasts recovered to their initial value by 10 minutes. Phyticmore » acid (IP/sub 6/), also stimulated /sup 45/Ca/sup 2 +/ efflux from the protoplasts. Both the IP/sub 3/- and the IP/sub 6/-induced /sup 45/Ca/sup 2 +/ efflux were inhibited by the calmodulin antagonist, trifluoperazine.« less

A Novel Hydrogen Sulfide-releasing N-Methyl-d-Aspartate Receptor Antagonist Prevents Ischemic Neuronal Death*

PubMed Central

Marutani, Eizo; Kosugi, Shizuko; Tokuda, Kentaro; Khatri, Ashok; Nguyen, Rebecca; Atochin, Dmitriy N.; Kida, Kotaro; Van Leyen, Klaus; Arai, Ken; Ichinose, Fumito

2012-01-01

Physiological levels of H2S exert neuroprotective effects, whereas high concentrations of H2S may cause neurotoxicity in part via activation of NMDAR. To characterize the neuroprotective effects of combination of exogenous H2S and NMDAR antagonism, we synthesized a novel H2S-releasing NMDAR antagonist N-((1r,3R,5S,7r)-3,5-dimethyladamantan-1-yl)-4-(3-thioxo-3H-1,2-dithiol-4-yl)-benzamide (S-memantine) and examined its effects in vitro and in vivo. S-memantine was synthesized by chemically combining a slow releasing H2S donor 4-(3-thioxo-3H-1,2-dithiol-4-yl)-benzoic acid (ACS48) with a NMDAR antagonist memantine. S-memantine increased intracellular sulfide levels in human neuroblastoma cells (SH-SY5Y) 10-fold as high as that was achieved by ACS48. Incubation with S-memantine after reoxygenation following oxygen and glucose deprivation (OGD) protected SH-SY5Y cells and murine primary cortical neurons more markedly than did ACS48 or memantine. Glutamate-induced intracellular calcium accumulation in primary cortical neurons were aggravated by sodium sulfide (Na2S) or ACS48, but suppressed by memantine and S-memantine. S-memantine prevented glutamate-induced glutathione depletion in SH-SY5Y cells more markedly than did Na2S or ACS48. Administration of S-memantine after global cerebral ischemia and reperfusion more robustly decreased cerebral infarct volume and improved survival and neurological function of mice than did ACS48 or memantine. These results suggest that an H2S-releasing NMDAR antagonist derivative S-memantine prevents ischemic neuronal death, providing a novel therapeutic strategy for ischemic brain injury. PMID:22815476

Role of calcium in the constriction of isolated cerebral arteries

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wendling, W.W.

1987-01-01

Calcium entry blockers (CEB) have been used in the experimental treatment or prevention of many cerebrovascular disorders including stroke, post-ischemic hypoperfusion after cardiac arrest, cerebral vasospasm after subarachnoid hemorrhage, and migraine headache. However, the mechanism of action of these drugs on the cerebral circulation is poorly understood. This study examined the effects of calcium antagonists, Ca/sup 2 +/-deficient solutions, and vasocostrictors on cerebrovascular tone and /sup 45/Ca fluxes, to determine the role of calcium in cerebral arterial constriction. A Scatchard plot of /sup 45/Ca binding to BMCA showed that Ca/sup 2 +/ was bound at either low or high affinitymore » binding sties. The four vasoconstrictors (potassium, serotonin, PGF/sub 2 ..cap alpha../, or SQ-26,655) each increased low affinity /sup 45/Ca uptake into BMCA. The results demonstrate that: (1) Potassium and serotonin constrict BMCA mainly by promoting Ca/sup 2 +/ influx through CEB-sensitive channels; (2) PGF/sub 2 ..cap alpha../ and SQ-26,655 constrict BMCA in part by promoting Ca/sup 2 +/ influx through CEB-sensitive channels, and in part by releasing Ca/sup 2 +/ from depletable internal stores; (3) The major action of CEB on BMCA is to block vasoconstrictor-induced Ca/sup 2 +/ uptake through both potential-operated (K/sup +/-stimulated) and receptor-operated channels.« less

Integrins are Mechanosensors That Modulate Human Eosinophil Activation

PubMed Central

Ahmadzai, Mustafa; Small, Mike; Sehmi, Roma; Gauvreau, Gail; Janssen, Luke J.

2015-01-01

Eosinophil migration to the lung is primarily regulated by the eosinophil-selective family of eotaxin chemokines, which mobilize intracellular calcium (Ca2+) and orchestrate myriad changes in cell structure and function. Eosinophil function is also known to be flow-dependent, although the molecular cognate of this mechanical response has yet to be adequately characterized. Using confocal fluorescence microscopy, we determined the effects of fluid shear stress on intracellular calcium concentration ([Ca2+]i) in human peripheral blood eosinophils by perfusing cells in a parallel-plate flow chamber. Our results indicate that fluid perfusion evokes a calcium response that leads to cell flattening, increase in cell area, shape change, and non-directional migration. None of these changes are seen in the absence of a flow stimulus, and all are blocked by chelation of intracellular Ca2+ using BAPTA. These changes are enhanced by stimulating the cells with eotaxin-1. The perfusion-induced calcium response (PICR) could be blocked by pre-treating cells with selective (CDP-323) and non-selective (RGD tripeptides) integrin receptor antagonists, suggesting that α4β7/α4β1 integrins mediate this response. Overall, our study provides the first pharmacological description of a molecular mechanosensor that may collaborate with the eotaxin-1 signaling program in order to control human eosinophil activation. PMID:26539194

Metabotropic glutamate receptor 5 responses dictate differentiation of neural progenitors to NMDA-responsive cells in fragile X syndrome.

PubMed

Achuta, Venkat Swaroop; Grym, Heli; Putkonen, Noora; Louhivuori, Verna; Kärkkäinen, Virve; Koistinaho, Jari; Roybon, Laurent; Castrén, Maija L

2017-04-01

Disrupted metabotropic glutamate receptor 5 (mGluR5) signaling is implicated in many neuropsychiatric disorders, including autism spectrum disorder, found in fragile X syndrome (FXS). Here we report that intracellular calcium responses to the group I mGluR agonist (S)-3,5-dihydroxyphenylglycine (DHPG) are augmented, and calcium-dependent mGluR5-mediated mechanisms alter the differentiation of neural progenitors in neurospheres derived from human induced pluripotent FXS stem cells and the brains of mouse model of FXS. Treatment with the mGluR5 antagonist 2-methyl-6-(phenylethynyl)-pyridine (MPEP) prevents an abnormal clustering of DHPG-responsive cells that are responsive to activation of ionotropic receptors in mouse FXS neurospheres. MPEP also corrects morphological defects of differentiated cells and enhanced migration of neuron-like cells in mouse FXS neurospheres. Unlike in mouse neurospheres, MPEP increases the differentiation of DHPG-responsive radial glial cells as well as the subpopulation of cells responsive to both DHPG and activation of ionotropic receptors in human neurospheres. However, MPEP normalizes the FXS-specific increase in the differentiation of cells responsive only to N-methyl-d-aspartate (NMDA) present in human neurospheres. Exposure to MPEP prevents the accumulation of intermediate basal progenitors in embryonic FXS mouse brain suggesting that rescue effects of GluR5 antagonist are progenitor type-dependent and species-specific differences of basal progenitors may modify effects of MPEP on the cortical development. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 419-437, 2017. © 2016 Wiley Periodicals, Inc.

Adrenomedullin increased the short-circuit current in the pig oviduct through chloride channels via the CGRP receptor: mediation by cAMP and calcium ions but not by nitric oxide.

PubMed

Liao, S B; Cheung, K H; Cheung, M P L; To, Y T; O, W S; Tang, F

2013-10-01

The oviduct serves as a site for the fertilization of the ovum and the transport of the conceptus down to the uterus for implantation. In this study, we investigated the presence of adrenomedullin (ADM) and its receptor component proteins in the pig oviduct. The effect of ADM on oviductal secretion, the specific receptor, and the mechanisms involved were also investigated. The presence of ADM and its receptor component proteins in the pig oviduct were confirmed using immunostaining. Short-circuit current (I(sc)) technique was employed to study chloride ion secretion in the oviductal epithelium. ADM increased I(sc) through cAMP- and calcium-activated chloride channels, and this effect could be inhibited by the CGRP receptor antagonist, hCGRP8-37. In contrast, the nitric oxide synthase inhibitor, L-NG-nitroarginine methyl ester (L-NAME), could not block the effect of ADM on I(sc). In summary, ADM may increase oviductal fluid secretion via chloride secretion independent of the nitric oxide pathway for the transport of sperm and the conceptus.

Plasticity of calcium-permeable AMPA glutamate receptors in Pro-opiomelanocortin neurons.

PubMed

Suyama, Shigetomo; Ralevski, Alexandra; Liu, Zhong-Wu; Dietrich, Marcelo O; Yada, Toshihiko; Simonds, Stephanie E; Cowley, Michael A; Gao, Xiao-Bing; Diano, Sabrina; Horvath, Tamas L

2017-08-01

POMC neurons integrate metabolic signals from the periphery. Here, we show in mice that food deprivation induces a linear current-voltage relationship of AMPAR-mediated excitatory postsynaptic currents (EPSCs) in POMC neurons. Inhibition of EPSCs by IEM-1460, an antagonist of calcium-permeable (Cp) AMPARs, diminished EPSC amplitude in the fed but not in the fasted state, suggesting entry of GluR2 subunits into the AMPA receptor complex during food deprivation. Accordingly, removal of extracellular calcium from ACSF decreased the amplitude of mEPSCs in the fed but not the fasted state. Ten days of high-fat diet exposure, which was accompanied by elevated leptin levels and increased POMC neuronal activity, resulted in increased expression of Cp-AMPARs on POMC neurons. Altogether, our results show that entry of calcium via Cp-AMPARs is inherent to activation of POMC neurons, which may underlie a vulnerability of these neurons to calcium overload while activated in a sustained manner during over-nutrition.

Comparison of vasculoprotective effects of benidipine and losartan in a rat model of metabolic syndrome.

PubMed

Matsuzaki, Gen; Ishizaka, Nobukazu; Furuta, Kyoko; Hongo, Makiko; Saito, Kan; Sakurai, Ryota; Koike, Kazuhiko; Nagai, Ryozo

2008-06-10

Although antihypertensive drugs confer improvement in endothelial dysfunction and protection from atherogenesis in hypertension, different classes of antihypertensive drugs may elicit different degrees of vasculoprotective effects. We have investigated the effects of a long-acting calcium antagonist, benidipine, and an angiotensin AT(1) receptor antagonist, losartan, on the vascular damage observed in OLETF rats, an animal model of metabolic syndrome. At 34 weeks of age, OLETF rats were treated with either benidipine (3 mg/kg/day, per os) or losartan (25 mg/kg/day, per os) for 8 weeks. The extent of blood pressure reduction, restoration endothelium-dependent aortic relaxation, and elevation of serum nitrite/nitrate concentration did not differ significantly between benidipine- and losartan-treated OLETF rats. Benidipine and losartan also reduced the aortic expression of transforming growth factor-beta1 mRNA and thickening of the vascular wall to a similar extent. Increased cardiac fibrosis was also inhibited by both benidipine and losartan. These data suggest that, when used in an antihypertensive dose, benidipine is as effective as losartan in restoring vascular endothelial function and in suppressing of cardiovascular remodeling in an animal model of metabolic syndrome.

Mechanisms of Pyrethroid Insecticide-Induced Stimulation of Calcium Influx in Neocortical Neurons

PubMed Central

Cao, Zhengyu; Shafer, Timothy J.

2011-01-01

Pyrethroid insecticides bind to voltage-gated sodium channels (VGSCs) and modify their gating kinetics, thereby disrupting neuronal function. Pyrethroids have also been reported to alter the function of other channel types, including activation of voltage-gated calcium channels. Therefore, the present study compared the ability of 11 structurally diverse pyrethroids to evoke Ca2+ influx in primary cultures of mouse neocortical neurons. Nine pyrethroids (tefluthrin, deltamethrin, λ-cyhalothrin, β-cyfluthrin, esfenvalerate, S-bioallethrin, fenpropathrin, cypermethrin, and bifenthrin) produced concentration-dependent elevations in intracellular calcium concentration ([Ca2+]i) in neocortical neurons. Permethrin and resmethrin were without effect on [Ca2+]i. These pyrethroids displayed a range of efficacies on Ca2+ influx; however, the EC50 values for active pyrethroids all were within one order of magnitude. Tetrodotoxin blocked increases in [Ca2+]i caused by all nine active pyrethroids, indicating that the effects depended on VGSC activation. The pathways for deltamethrin- and tefluthrin-induced Ca2+ influx include N-methyl-d-aspartic acid receptors, L-type Ca2+ channels, and reverse mode of operation of the Na+/Ca2+ exchanger inasmuch as antagonists of these sites blocked deltamethrin-induced Ca2+ influx. These data demonstrate that pyrethroids stimulate Ca2+ entry into neurons subsequent to their actions on VGSCs. PMID:20881019

[Results of the Russian EKSPERT program: post-marketing supervision over efficacy and influence of the preparation Ekvator on quality of life at out-patients with arterial hypertension].

PubMed

Glezer, M G; Vygodin, V A; Avakian, A A; Prokof'eva, E B

2014-01-01

Results of an open multicenter prospective postmarketing observational program EKSPERT (post-marketing surveillance of the effectiveness and impact of the EKVATOR treatment on quality of life in patients with arterial hypertension in ambulatory practice). Observation of 10 000 patients conducted in 300 medical center in various regions of the Russian Federation in 1005 doctors. Selected for the final analysis 4954 registration cards. It is shown that in patients with initially insufficient effective antihypertensive treatment has a large number of risk factors: men older than 55 years--56.5%, women older than 65 years--27.8%, unfavorable family history of arterial hypertension (AH)--87.9%, diabetes mellitus (DM)--13.4%, smoking--of patients 18.6%, obesity--35%, angina--35.59%, heart failure--41.3% with a history of myocardial infarction--10.9%, stroke--4.5%, renal disease--11.8%, hypercholesterolemia > 5.0 mmol/l--76.7%. Initially drug antihypertensive treatment was performed in 76.6% of patients, while 43.9% were treated regularly. Prior studies angiotensin converting enzyme inhibitors (ACE) afforded 60.56%, sartans--11% of patients, beta-blockers--41.9%, duretics--41.46%, calcium antagonists used in 21.42% of the patients. After the cancellation of previously used other ACE inhibitors, calcium antagonists and sartans patients were switched to therapy with the EKVATOR (amlodipine and lisinopril). Intensity reduction in systolic and diastolic blood pressure (SBP and DBP) did not depend on sex of the patients, the presence of angina, diabetes. Greater reduction in blood pressure in hypertensive duration more than 5 years, in the presence of congestive heart failure due to more frequent initiation of therapy with full-dose combination (amlodipine 10 mg and lisinopril 20 mg). After 1 months of starting therapy changes uorvney target blood pressure (< 140 and 90 mmHg) reached 51.5% of patients. Target SBP reached 59.7% of patients, the target level of DBP--69.4%. It is important that the majority of patients crossed over lowti graduation SBP and DBP and significantly improved their quality of life assessment. Incidence of adverse events was low--1.5% of them are the most common were swelling in the legs, headache, dizziness, and dry cough. Replacing the previous therapy different ACE inhibitors, sartans and calcium antagonists to the fixed combination amlodipine and lisinopril) (drug EKVATOR), leads to a rapid, pronounced, and safe reduction of BP and improve health in the majority of patients with previously uncorrected BP.

One pot synthesis of some new substituted hexahydro 2H-1,3-benzoxazine derivatives.

PubMed

Safak, C; Simsek, R; Altas, Y; Erol, K; Boydag, S

1996-09-01

In this paper, we synthesized nineteen new compounds having 2,4-diaryl-5-oxohexahydro-2H-1,3-benzoxazine structure by the reaction of 1,3-cyclohexanedione, aromatic aldehyde and ammonium acetate. In addition, we evaluated calcium antagonistic activity of these compounds versus nicardipine.

R-Type Ca2+ channels couple to inhibitory neurotransmission to the longitudinal muscle in the guinea-pig ileum.

PubMed

Rodriguez-Tapia, Eileen S; Naidoo, Vinogran; DeVries, Matthew; Perez-Medina, Alberto; Galligan, James J

2017-03-01

What is the central question of this study? Subtypes of enteric neurons are coded by the neurotransmitters they synthesize, but it is not known whether enteric neuron subtypes might also be coded by other proteins, including calcium channel subtypes controlling neurotransmitter release. What is the main finding and its importance? Our data indicate that guinea-pig ileum myenteric neuron subtypes may be coded by calcium channel subtypes. We found that R-type calcium channels are expressed by inhibitory but not excitatory longitudinal muscle motoneurons. R-Type calcium channels are also not expressed by circular muscle inhibitory motoneurons. Calcium channel subtype-selective antagonists could be used to target subtypes of neurons to treat gastrointestinal motility disorders. There is evidence that R-type Ca 2+ channels contribute to synaptic transmission in the myenteric plexus. It is unknown whether R-type Ca 2+ channels contribute to neuromuscular transmission. We measured the effects of the nitric oxide synthase inhibitor nitro-l-arginine (NLA), Ca 2+ channel blockers and apamin (SK channel blocker) on neurogenic relaxations and contractions of the guinea-pig ileum longitudinal muscle-myenteric plexus (LMMP) in vitro. We used intracellular recordings to measure inhibitory junction potentials. Immunohistochemical techniques localized R-type Ca 2+ channel protein in the LMMP and circular muscle. Cadmium chloride (pan-Ca 2+ channel blocker) blocked and NLA and NiCl 2 (R-type Ca 2+ channel blocker) reduced neurogenic relaxations in a non-additive manner. Nickel chloride did not alter neurogenic cholinergic contractions, but it potentiated neurogenic non-cholinergic contractions. Relaxations were inhibited by apamin, NiCl 2 and NLA and were blocked by combined application of these drugs. Relaxations were reduced by NiCl 2 or ω-conotoxin (N-type Ca 2+ channel blocker) and were blocked by combined application of these drugs. Longitudinal muscle inhibitory junction potentials were inhibited by NiCl 2 but not MRS 2179 (P2Y 1 receptor antagonist). Circular muscle inhibitory junction potentials were blocked by apamin, MRS 2179, ω-conotoxin and CdCl 2 but not NiCl 2 . We conclude that neuronal R-type Ca 2+ channels contribute to inhibitory neurotransmission to longitudinal muscle but less so or not all in the circular muscle of the guinea-pig ileum. © 2016 The Authors. Experimental Physiology © 2016 The Physiological Society.

Identification of a Calcium Signalling Pathway of S-[6]-Gingerol in HuH-7 Cells.

PubMed

Li, Xiao-Hong; McGrath, Kristine C Y; Tran, Van H; Li, Yi-Ming; Mandadi, Sravan; Duke, Colin C; Heather, Alison K; Roufogalis, Basil D

2013-01-01

Calcium signals in hepatocytes control cell growth, proliferation, and death. Members of the transient receptor potential (TRP) cation channel superfamily are candidate calcium influx channels. NF κ B activation strictly depends on calcium influx and often induces antiapoptotic genes favouring cell survival. Previously, we reported that S-[6]-gingerol is an efficacious agonist of the transient receptor potential cation channel subfamily V member 1 (TRPV1) in neurones. In this study, we tested the effect of S-[6]-gingerol on HuH-7 cells using the Fluo-4 calcium assay, RT-qPCR, transient cell transfection, and luciferase measurements. We found that S-[6]-gingerol induced a transient rise in [Ca(2+)] i in HuH-7 cells. The increase in [Ca(2+)] i induced by S-[6]-gingerol was abolished by preincubation with EGTA and was also inhibited by the TRPV1 channel antagonist capsazepine. Expression of TRPV1 in HuH-7 cells was confirmed by mRNA analysis as well as a test for increase of [Ca(2+)] i by TRPV1 agonist capsaicin and its inhibition by capsazepine. We found that S-[6]-gingerol induced rapid NF κ B activation through TRPV1 in HuH-7 cells. Furthermore, S-[6]-gingerol-induced NF κ B activation was dependent on the calcium gradient and TRPV1. The rapid NF κ B activation by S-[6]-gingerol was associated with an increase in mRNA levels of NF κ B-target genes: cIAP-2, XIAP, and Bcl-2 that encode antiapoptotic proteins.

A Critical Role for the Transient Receptor Potential Channel Type 6 in Human Platelet Activation

PubMed Central

Conlon, Christine; Khasawneh, Fadi T.

2015-01-01

While calcium signaling is known to play vital roles in platelet function, the mechanisms underlying its receptor-operated calcium entry component (ROCE) remain poorly understood. It has been proposed, but never proven in platelets, that the canonical transient receptor potential channel-6 (TRPC6) mediates ROCE. Nonetheless, we have previously shown that the mouse TRPC6 regulates hemostasis, thrombogenesis by regulating platelet aggregation. In the present studies, we used a pharmacological approach to characterize the role of TRPC6 in human platelet biology. Thus, interestingly, we observed that a TRPC6 inhibitor exerted significant inhibitory effects on human platelet aggregation in a thromboxane receptor (TPR)-selective manner; no additional inhibition was observed in the presence of the calcium chelator BAPTA. This inhibitor also significantly inhibited human platelet secretion (dense and alpha granules), integrin IIb-IIIa, Akt and ERK phosphorylation, again, in a TPR-selective manner; no effects were observed in response to ADP receptor stimulation. Furthermore, there was a causal relationship between these inhibitory effects, and the capacity of the TRPC6 inhibitor to abrogate elevation in intracellular calcium, that was again found to be TPR-specific. This effect was not found to be due to antagonism of TPR, as the TRPC6 inhibitor did not displace the radiolabeled antagonist [3H]SQ29,548 from its binding sites. Finally, our studies also revealed that TRPC6 regulates human clot retraction, as well as physiological hemostasis and thrombus formation, in mice. Taken together, our findings demonstrate, for the first time, that TRPC6 directly regulates TPR-dependent ROCE and platelet function. Moreover, these data highlight TRPC6 as a novel promising therapeutic strategy for managing thrombotic disorders. PMID:25928636

Verrucotoxin, a stonefish venom, modulates calcium channel activity in guinea-pig ventricular myocytes.

PubMed

Yazawa, K; Wang, J-W; Hao, L-Y; Onoue, Y; Kameyama, M

2007-08-01

Stonefish (Synanceia genus) are commonly found in shallow waters of the Pacific and Indian Oceans. The venom of stonefish is stored in the dorsal fine spines and contains a proteinaceous toxin, verrucotoxin (VTX). The stings produced by the spines induce intense pain, respiratory weakness, damage to the cardiovascular system, convulsions and paralysis, sometimes leading to death. Although there are many studies on VTX, the mechanism(s) underlying the VTX-mediated cardiotoxicity is not yet fully understood. The aim of this study was to investigate the modulation of ion channels in cardiac tissue by VTX. The effects of VTX on changes in the voltage or current in guinea-pig ventricular myocytes were investigated using a patch clamp method. VTX (10 microg ml(-1)) prolonged the action potential duration by 2.5-fold. VTX increased L-type Ca(2+) currents (I (Ca(L))) in a concentration-dependent manner with a EC(50) value of 7 microg ml(-1) and a maximum increase of 3.1-fold. The non-selective beta-adrenoceptor antagonist, propranolol (1 microM) and the selective beta(1)-adrenoceptor antagonist, CGP20712A (10 microM) each abolished the effect of VTX (100 microg ml(-1)) on I (Ca(L)). Furthermore, the protein kinase A (PKA) antagonists H-89 (10 microM) and Rp-8-Br-cAMPS (30 microM) inhibited the effect of VTX on I (Ca(L)). VTX modulates Ca(2+) channel activity through the beta-adrenoceptor-cAMP-PKA pathway.

NGP1-01, a multi-targeted polycyclic cage amine, attenuates brain endothelial cell death in iron overload conditions.

PubMed

Lockman, J A; Geldenhuys, W J; Jones-Higgins, M R; Patrick, J D; Allen, D D; Van der Schyf, C J

2012-12-13

Development and progression of neurodegenerative disorders have, amongst other potential causes, been attributed to a disruption of iron regulatory mechanisms and iron accumulation. Excess extracellular iron may enter cells via nontraditional routes such as voltage-gated calcium channels and N-methyl-d-aspartate (NMDA) receptors leading to intracellular oxidative damage and ultimately mitochondrial failure. Nimodipine, an L-type calcium channel blocker has been shown to reduce iron-induced toxicity in neuronal and brain endothelial cells. Our current study investigates NGP1-01, a multimodal drug acting as an antagonist at both the NMDA receptor and the L-type calcium channel. Our previous studies support NGP1-01 as a promising neuroprotective agent in diseases involving calcium-related excitotoxicity. We demonstrate here that NGP1-01 (1 and 10μM) pretreatment abrogates the effects of iron overload in brain endothelial cells protecting cellular viability. Both concentrations of NGP1-01 were found to attenuate iron-induced reduction in cellular viability to a similar extent, and were statistically significant. To further verify the mechanism, the L-type calcium channel agonist FPL 64176 was administered to promote iron uptake. Addition of NGP1-01 dose-dependently reduced FPL 64176 stimulated uptake of iron. These data support further evaluation of NGP1-01 as a neuroprotective agent, not only in diseases associated with excitotoxicity, but also in those of iron overload. Copyright © 2012 Elsevier B.V. All rights reserved.

Calcium antagonists. A role in the management of cyanide poisoning

DOE Office of Scientific and Technical Information (OSTI.GOV)

Maduh, E.U.; Porter, D.W.; Baskin, S.I.

1993-12-31

The physiological role of calcium was demonstrated by Ringer (1883) when he linked the omission of calcium (Ca++) from the bathing medium to the induction of cardiac arrest in the isolated frog heart. This observation established that Ca++ controlled muscle contraction but it was not until the autumn of 1963 that the specific pharmacological significance of this contribution was realised by Fleckenstein (1964), leading to the development of Ca++ antagonism as a concept in drug action (Fleckenstein 1977). Identifying the precise role of Ca++ ions in toxic cell injury and tissue death attributable to drug and chemical intoxication has laggedmore » behind developments in Ca++ physiology and pharmacology and to date, much remains to be learned, although studies aimed at characterising the role of Ca++ in cytotoxic cell injury are receiving intense attention (Bondy Komulainen 1988; Maduh et al. l988a, l99Oa,b; Orrenius et al. 1989; Trump et al. 1989). On the other hand, the importance of cyanide as a poison has been known from antiquity (for references to earlier literature see Baskin Fricke 1992; Solomonson 1981). In experimental cyanide poisoning, recent studies have examined alterations in cell Ca++ and the influence of Ca++ antagonists in the management of this chemical toxicological emergency. These efforts have principally focused on the cellular Ca++ homeostasis system, its interrelationship with cellular components, and its susceptibility to cyanide action.« less

High pH-Sensitive TRPA1 Activation in Odontoblasts Regulates Mineralization.

PubMed

Kimura, M; Sase, T; Higashikawa, A; Sato, M; Sato, T; Tazaki, M; Shibukawa, Y

2016-08-01

Calcium hydroxide and mineral trioxide aggregate are widely used for indirect and direct pulp capping and root canal filling. Their dissociation into Ca(2+) and OH(-) in dental pulp creates an alkaline environment, which activates reparative/reactionary dentinogenesis. However, the mechanisms by which odontoblasts detect the pH of the extracellular environment remain unclear. We examined the alkali-sensitive intracellular Ca(2+) signaling pathway in rat odontoblasts. In the presence or absence of extracellular Ca(2+), application of alkaline solution increased intracellular Ca(2+) concentration, or [Ca(2+)]i Alkaline solution-induced [Ca(2+)]i increases depended on extracellular pH (8.5 to 10.5) in both the absence and the presence of extracellular Ca(2+) The amplitude was smaller in the absence than in the presence of extracellular Ca(2+) Each increase in [Ca(2+)]i, activated by pH 7.5, 8.5, or 9.5, depended on extracellular Ca(2+) concentration; the equilibrium binding constant for extracellular Ca(2+) concentration decreased as extracellular pH increased (1.04 mM at pH 7.5 to 0.11 mM at pH 9.5). Repeated applications of alkaline solution did not have a desensitizing effect on alkali-induced [Ca(2+)]i increases and inward currents. In the presence of extracellular Ca(2+), alkaline solution-induced [Ca(2+)]i increases were suppressed by application of an antagonist of transient receptor potential ankyrin subfamily member 1 (TRPA1) channels. Ca(2+) exclusion efficiency during alkaline solution-induced [Ca(2+)]i increases was reduced by a Na(+)-Ca(2+) exchanger antagonist. Alizarin red and von Kossa staining revealed increased mineralization levels under repeated high pH stimulation, whereas the TRPA1 antagonist strongly reduced this effect. These findings indicate that alkaline stimuli-such as the alkaline environment inside dental pulp treated with calcium hydroxide or mineral trioxide aggregate-activate Ca(2+) mobilization via Ca(2+) influx mediated by TRPA1 channels and intracellular Ca(2+) release in odontoblasts. High pH-sensing mechanisms in odontoblasts are important for activating dentinogenesis induced by an alkaline environment. © International & American Associations for Dental Research 2016.

The tripeptide aldehyde, Boc-DPhe-Phe-Lysinal, is a novel Ca2+ channel inhibitor in pituitary cells.

PubMed

Makara, G B; Rappay, G; Garamvölgyi, V; Nagy, I; Dankó, S; Bajusz, S

1988-06-22

The effect of Boc-DPhe-Phe-Lysinal (Boc-DPPL) on the 45Ca2+ uptake of rat anterior pituitary monolayer cultures was investigated. The compound decreased the basal Ca2+ uptake at 3 x 10(-4) mol/l. The 45Ca2+ uptake stimulated by potassium-induced depolarization was more sensitive to Boc-DPPL inhibition, a slight decrease was seen with 3 x 10(-6) mol/l and there was a half maximal inhibition at 3 x 10(-5) mol/l. Boc-DPPL is known to inhibit pituitary hormone release in similar concentrations, an effect might also be due to its calcium antagonist property.

TRPV1 mediates cell death in rat synovial fibroblasts through calcium entry-dependent ROS production and mitochondrial depolarization

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hu Fen; Sun Wenwu; Zhao Xiao Ting

Synoviocyte hyperplasia is critical for rheumatoid arthritis, therefore, potentially an important target for therapeutics. It was found in this work that a TRPV1 agonist capsaicin, and acidic solution (pH 5.5) induced increases in cytosolic calcium concentration ([Ca{sup 2+}]{sub c}) and reactive oxygen species (ROS) production in synoviocytes isolated from a rat model of collagen-induced arthritis. The increases in both [Ca{sup 2+}]{sub c} and ROS production were completely abolished in calcium-free buffer or by a TRPV1 antagonist capsazepine. Further experiments revealed that capsaicin and pH 5.5 solution caused mitochondrial membrane depolarization and reduction in cell viability; such effects were inhibited bymore » capsazepine, or the NAD(P)H oxidase inhibitor diphenylene iodonium. Both capsaicin and pH 5.5 buffer induced apoptosis as shown by nuclear condensation and fragmentation. Furthermore, RT-PCR readily detected TRPV1 mRNA expression in the isolated synoviocytes. Taken together, these data indicated that TRPV1 activation triggered synoviocyte death by [Ca{sup 2+}]{sub c} elevation, ROS production, and mitochondrial membrane depolarization.« less

Angiotensin II type 2 receptor (AT2R) localization and antagonist-mediated inhibition of capsaicin responses and neurite outgrowth in human and rat sensory neurons

PubMed Central

Anand, U; Facer, P; Yiangou, Y; Sinisi, M; Fox, M; McCarthy, T; Bountra, C; Korchev, YE; Anand, P

2013-01-01

Background The angiotensin II (AngII) receptor subtype 2 (AT2R) is expressed in sensory neurons and may play a role in nociception and neuronal regeneration. Methods We used immunostaining with characterized antibodies to study the localization of AT2R in cultured human and rat dorsal root ganglion (DRG) neurons and a range of human tissues. The effects of AngII and AT2R antagonist EMA401 on capsaicin responses in cultured human and rat (DRG) neurons were measured with calcium imaging, on neurite length and density with Gap43 immunostaining, and on cyclic adenosine monophosphate (cAMP) expression using immunofluorescence. Results AT2R expression was localized in small-/medium-sized cultured neurons of human and rat DRG. Treatment with the AT2R antagonist EMA401 resulted in dose-related functional inhibition of capsaicin responses (IC50 = 10 nmol/L), which was reversed by 8-bromo-cAMP, and reduced neurite length and density; AngII treatment significantly enhanced capsaicin responses, cAMP levels and neurite outgrowth. The AT1R antagonist losartan had no effect on capsaicin responses. AT2R was localized in sensory neurons of human DRG, and nerve fibres in peripheral nerves, skin, urinary bladder and bowel. A majority sub-population (60%) of small-/medium-diameter neuronal cells were immunopositive in both control post-mortem and avulsion-injured human DRG; some very small neurons appeared to be intensely immunoreactive, with TRPV1 co-localization. While AT2R levels were reduced in human limb peripheral nerve segments proximal to injury, they were preserved in painful neuromas. Conclusions AT2R antagonists could be particularly useful in the treatment of chronic pain and hypersensitivity associated with abnormal nerve sprouting. PMID:23255326

Minoxidil-induced hair growth is mediated by adenosine in cultured dermal papilla cells: possible involvement of sulfonylurea receptor 2B as a target of minoxidil.

PubMed

Li, M; Marubayashi, A; Nakaya, Y; Fukui, K; Arase, S

2001-12-01

The mechanism by which minoxidil, an adenosine-triphosphate-sensitive potassium channel opener, induces hypertrichosis remains to be elucidated. Minoxidil has been reported to stimulate the production of vascular endothelial growth factor, a possible promoter of hair growth, in cultured dermal papilla cells. The mechanism of production of vascular endothelial growth factor remains unclear, however. We hypothesize that adenosine serves as a mediator of vascular endothelial growth factor production. Minoxidil-induced increases in levels of intracellular Ca(2+) and vascular endothelial growth factor production in cultured dermal papilla cells were found to be inhibited by 8-sulfophenyl theophylline, a specific antagonist for adenosine receptors, suggesting that dermal papilla cells possess adenosine receptors and sulfonylurea receptors, the latter of which is a well-known target receptor for adenosine-triphosphate-sensitive potassium channel openers. The expression of sulfonylurea receptor 2B and of the adenosine A1, A2A, and A2B receptors was detected in dermal papilla cells by means of reverse transcription polymerase chain reaction analysis. In order to determine which of the adenosine receptor subtypes contribute to minoxidil-induced hair growth, the effects of subtype-specific antagonists for adenosine receptors were investigated. Significant inhibition in increase in intracellular calcium level by minoxidil or adenosine was observed as the result of pretreatment with 8-cyclopentyl-1,3-dipropylxanthine, an antagonist for adenosine A1 receptor, but not by 3,7-dimethyl-1-propargyl-xanthine, an antagonist for adenosine A2 receptor, whereas vascular endothelial growth factor production was blocked by both adenosine A1 and A2 receptor antagonists. These results indicate that the effect of minoxidil is mediated by adenosine, which triggers intracellular signal transduction via both adenosine A1 and A2 receptors, and that the expression of sulfonylurea receptor 2B in dermal papilla cells might play a role in the production of adenosine.

Cooling-induced contraction in ovine airways smooth muscle.

PubMed

Mustafa, S M; Pilcher, C W; Williams, K I

1999-02-01

The mechanism of cold-induced bronchoconstriction is poorly understood. This prompted the present study whose aim was to determine the step-wise direct effect of cooling on smooth muscle of isolated ovine airways and analyse the role of calcium in the mechanisms involved. Isolated tracheal strips and bronchial segments were suspended in organ baths containing Krebs' solution for isometric tension recording. Tissue responses during stepwise cooling from 37 to 5 degrees C were examined. Cooling induced a rapid and reproducible contraction proportional to cooling temperature in ovine tracheal and bronchial preparations which was epithelium-independent. On readjustment to 37 degrees C the tone returned rapidly to basal level. Maximum contraction was achieved at a temperature of 5 degrees C for trachea and 15 degrees C for bronchiole. Cooling-induced contractions (CIC) was resistant to tetrodotoxin (1; 10 micrometer), and not affected by the muscarinic antagonist atropine (1 micrometer) or the alpha-adrenergic antagonist phentolamine (1 micrometer), or the histamine H1-antagonist mepyramine (1 micrometer) or indomethacin (1 micrometer). Ca2+ antagonists (nifedipine and verapamil) and Mn2+ raised tracheal but not bronchiolar tone and augmented CIC. Incubation in Ca2+-free, EGTA-containing Krebs' solution for 5 min had no effect on CIC, although it significantly reduced KCl-induced contraction by up to 75%. Cooling inhibited Ca2+ influx measured using 45Ca2+ uptake. Caffeine (100 micrometer) significantly inhibited CIC. The results show that cooling-induced contractions do not appear to involve activation of nerve endings, all surface reception systems or Ca2+ influx. However, CIC is mainly dependent on release of intracellular Ca2+. Copyright 1999 The Italian Pharmacological Society.

Calcium currents, charge movement and dihydropyridine binding in fast- and slow-twitch muscles of rat and rabbit.

PubMed Central

Lamb, G D; Walsh, T

1987-01-01

1. The Vaseline-gap technique was used to record slow calcium currents and asymmetric charge movement in single fibres of fast-twitch muscles (extensor digitorum longus (e.d.l.) and sternomastoid) and slow-twitch muscles (soleus) from rat and rabbit, at a holding potential of -90 mV. 2. The slow calcium current in soleus fibres was about one-third of the size of the current in e.d.l. fibres, but was very similar otherwise. In both e.d.l. and soleus fibres, the dihydropyridine (DHP), nifedipine, suppressed the calcium current entirely. 3. In these normally polarized fibres, nifedipine suppressed only part (qns) of the asymmetric charge movement. The proportion of qns suppressed by various concentrations of nifedipine was linearly related to the associated reduction of the calcium current. Half-maximal suppression of both parameters was obtained with about 0.5 microM-nifedipine. The calcium current and the qns component of the charge movement also were suppressed over the same time course by nifedipine. Another DHP calcium antagonist, (+)PN200/110, was indistinguishable from nifedipine in its effects of suppressing calcium currents and qns. 4. In all muscle types, the total amount of qns in each fibre was linearly related to the size of the calcium current (in the absence of DHP). On average, qns was 3.3 times larger in e.d.l. fibres than in soleus fibres. 5. In contrast to the other dihydropyridines, (-)bay K8644, a calcium channel agonist, did not suppress any asymmetric charge movement. 6. The potential dependence of the slow calcium current implied a minimum gating charge of about five or six electronic charges. The movement of qns occurred over a more negative potential range than the change in calcium conductance. 7. Experiments on the binding of (+)PN200/110 indicated that e.d.l. muscles had between about 2 and 3 times more specific DHP binding sites than did soleus muscle. 8. These results point to a close relationship between slow calcium channels, the qns component of the charge movement and DHP binding sites, in both fast- and slow-twitch mammalian muscle. qns appears to be part of the gating current of the T-system calcium channels. PMID:2451745

The analgesic, anti-inflammatory and calcium antagonist potential of Tanacetum artemisioides.

PubMed

Bukhari, Ishfaq Ali; Khan, Rafeeq Alam; Gilani, Anwar-ul Hassan; Shah, Abdul Jabbar; Hussain, Javid; Ahmad, Viqar Uddin

2007-03-01

Several species of the genus Tanacetum are traditionally used in a variety of health conditions including pain, inflammation, respiratory and gastrointestinal disorders. In the current investigation, we evaluated the plant extract of T. artemisioides and some of its pure compounds (flavonoids) for analgesic, anti-inflammatory and calcium antagonist effects in various in-vivo and in vitro studies. Using the actetic acid induced writhing test, intraperitoneal (i.p) administration of the plant extract (25-50 mg/kg) and its flavonoid compounds TA-1 and TA-2 (1-5 mg/kg ) exhibited significant analgesic actvity. The maximum analgesic effect observed with the crude extract of the plant was 71% at 50 mg/kg, while that of compounds TA-1 and TA-2 (5 mg/kg i.p) was 75 and 47%, respectively. The plant extract and its pure compounds caused inhbition of formalin induced paw licking in mice predominatly in the second phase of the test. Diclofenac sodium, a standard reference compound, showed a simlar effect in these chemical induced pain models. In the carrgeenan induced rat paw edema assay, the plant extract (50-200 mg/kg i.p) demonstrated significant (P< 0.01) anti-inflammatory activity which was comparable to that obtained with diclofenac sodium and indomethacin. In isolated rabbit jejunum preprations the plant extract showed an atropine sensitive dose-dependent (0.10-1.0 mg/mL) spasmogenic activity followed by a spasmolytic effect at the next higher doses (3-5 mg/mL). The crude extract of the plant also inhibited the high K+-induced contractions, indicating a calcium channel blocking (CCB) activity, which was further confirmed when the plant extract caused a rightward shift in the Ca++ concentration response curves in the isolated rabbit jejunum preparations, similar to that seen with verapamil. The flavonoid compounds isolated from the plant were devoid of any activity in the isolated tissue preparations. These results indicate that the plant extract of T. artemisioides possesses analgesic, anti-inflammatory and CCB activities. The flavonoid compounds of the plant may have a role in its observed analgesic and antiinflammatory activities, while the CCB activity of the plant may be attributed to some other chemical constituents present. Moreover the findings support the traditional reputation of the genus Tanacetum for its therapeutic benefits in pain and inflammatory conditions.

Memantine Can Reduce Ethanol-Induced Caspase-3 Activity and Apoptosis in H4 Cells by Decreasing Intracellular Calcium.

PubMed

Wang, Xiaolong; Chen, Jiajun; Wang, Hongbo; Yu, Hao; Wang, Changliang; You, Jiabin; Wang, Pengfei; Feng, Chunmei; Xu, Guohui; Wu, Xu; Zhao, Rui; Zhang, Guohua

2017-08-01

Caspase-3 activation and apoptosis are associated with various neurodegenerative disorders. Calcium activation is an important factor in promoting apoptosis. We, therefore, assessed the role of intracellular calcium in ethanol-induced activation of caspase-3 in H4 human neuroglioma cells and the protective effect of the NMDA receptor antagonist, memantine, on ethanol-induced apoptosis in H4 cells. H4 cells were treated with 100 mM EtOH (in culture medium) for 2 days. For interaction studies, cells were treated with memantine (4 μM), EDTA (1 mM), or BAPTA-AM (10 μM) before treatment with EtOH. Knockdown of the gene encoding the NR1 subunit of the NMDA receptor was performed using RNAi. Apoptosis was detected by Annexin V-FITC/PI staining and flow cytometry. Cell viability was detected using an MTS cell proliferation kit. Fluorescence dual wavelength spectrophotometry was used to determine the intracellular calcium concentration. The levels of NR1, caspase-3, IP3R1, and SERCA1 proteins were detected by western blotting. NR1, IP3R1, and SERCA1 mRNA levels were detected by qPCR. We observed increased expression of NR1, IP3R1, SERCA1, and increased intracellular levels of calcium ions in H4 cells exposed to ethanol. In addition, the calcium chelators, EDTA and BAPTA, and RNAi disruption of the NMDA receptor reduced ethanol-induced caspase-3 activation in H4 cells. Memantine treatment reduced the ethanol-induced increase of intracellular calcium, caspase-3 activation, apoptosis, and the ethanol-induced decrease in cell viability. Our results indicate that ethanol-induced caspase-3 activation and apoptosis are likely to be dependent on cytosolic calcium levels and that they can be reduced by memantine treatment.

Inflammatory sensitization of nociceptors depends on activation of NMDA receptors in DRG satellite cells.

PubMed

Ferrari, Luiz Fernando; Lotufo, Celina Monteiro; Araldi, Dionéia; Rodrigues, Marcos A; Macedo, Larissa P; Ferreira, Sérgio H; Parada, Carlos Amilcar

2014-12-23

The present study evaluated the role of N-methyl-D-aspartate receptors (NMDARs) expressed in the dorsal root ganglia (DRG) in the inflammatory sensitization of peripheral nociceptor terminals to mechanical stimulation. Injection of NMDA into the fifth lumbar (L5)-DRG induced hyperalgesia in the rat hind paw with a profile similar to that of intraplantar injection of prostaglandin E2 (PGE2), which was significantly attenuated by injection of the NMDAR antagonist D(-)-2-amino-5-phosphonopentanoic acid (D-AP-5) in the L5-DRG. Moreover, blockade of DRG AMPA receptors by the antagonist 6,7-dinitroquinoxaline-2,3-dione had no effect in the PGE2-induced hyperalgesia in the paw, showing specific involvement of NMDARs in this modulatory effect and suggesting that activation of NMDAR in the DRG plays an important role in the peripheral inflammatory hyperalgesia. In following experiments we observed attenuation of PGE2-induced hyperalgesia in the paw by the knockdown of NMDAR subunits NR1, NR2B, NR2D, and NR3A with antisense-oligodeoxynucleotide treatment in the DRG. Also, in vitro experiments showed that the NMDA-induced sensitization of cultured DRG neurons depends on satellite cell activation and on those same NMDAR subunits, suggesting their importance for the PGE2-induced hyperalgesia. In addition, fluorescent calcium imaging experiments in cultures of DRG cells showed induction of calcium transients by glutamate or NMDA only in satellite cells, but not in neurons. Together, the present results suggest that the mechanical inflammatory nociceptor sensitization is dependent on glutamate release at the DRG and subsequent NMDAR activation in satellite glial cells, supporting the idea that the peripheral hyperalgesia is an event modulated by a glutamatergic system in the DRG.

Calcium movements during pigment aggregation in freshwater shrimp chromatophores.

PubMed

Ribeiro, Márcia; McNamara, John Campbell

2007-02-01

Pigment granule migration within crustacean chromatophores provides an excellent model with which to investigate cytoplasmic movements, given the antagonistic, neurosecretory peptide regulation of granule translocation, and the absence of innervation in these large, brightly colored cells. Red pigment-concentrating hormone (RPCH) induces pigment aggregation in shrimp chromatophores via an increase in intracellular Ca2+; however, how this increase is brought about is not known. To examine the putative Ca2+ movements leading to pigment translocation in red, ovarian chromatophores of the freshwater shrimp, Macrobrachium olfersii, this study manipulates intra- and extracellular Ca2+ employing ER Ca2+-ATPase inhibitors, ryanodine-sensitive, ER Ca2+ channel blockers, and EDTA/EGTA-buffered A23187/Ca2+-containing salines. Our findings reveal that during pigment aggregation, cytosolic Ca2+ apparently increases from an intracellular source, the abundant SER, loaded by the SERCA and released through ryanodine-sensitive receptor/channels, triggered by capacitative calcium influx and/or calcium-induced calcium release mechanisms. Aggregation also depends on external calcium, which may modulate RPCH/receptor coupling. Such calcium-regulated pigment movements form the basis of a complex system of chromatic adaptation, which confers selective advantages like camouflage and protection against ultra-violet radiation to this palaemonid shrimp.

Ultrafine particles cause cytoskeletal dysfunctions in macrophages: role of intracellular calcium

PubMed Central

Möller, Winfried; Brown, David M; Kreyling, Wolfgang G; Stone, Vicki

2005-01-01

Background Particulate air pollution is reported to cause adverse health effects in susceptible individuals. Since most of these particles are derived form combustion processes, the primary composition product is carbon with a very small diameter (ultrafine, less than 100 nm in diameter). Besides the induction of reactive oxygen species and inflammation, ultrafine particles (UFP) can cause intracellular calcium transients and suppression of defense mechanisms of alveolar macrophages, such as impaired migration or phagocytosis. Methods In this study the role of intracellular calcium transients caused by UFP was studied on cytoskeleton related functions in J774A.1 macrophages. Different types of fine and ultrafine carbon black particles (CB and ufCB, respectively), such as elemental carbon (EC90), commercial carbon (Printex 90), diesel particulate matter (DEP) and urban dust (UD), were investigated. Phagosome transport mechanisms and mechanical cytoskeletal integrity were studied by cytomagnetometry and cell viability was studied by fluorescence microscopy. Macrophages were exposed in vitro with 100 and 320 μg UFP/ml/million cells for 4 hours in serum free medium. Calcium antagonists Verapamil, BAPTA-AM and W-7 were used to block calcium channels in the membrane, to chelate intracellular calcium or to inhibit the calmodulin signaling pathways, respectively. Results Impaired phagosome transport and increased cytoskeletal stiffness occurred at EC90 and P90 concentrations of 100 μg/ml/million cells and above, but not with DEP or UD. Verapamil and W-7, but not BAPTA-AM inhibited the cytoskeletal dysfunctions caused by EC90 or P90. Additionally the presence of 5% serum or 1% bovine serum albumin (BSA) suppressed the cytoskeletal dysfunctions. Cell viability showed similar results, where co-culture of ufCB together with Verapamil, W-7, FCS or BSA produced less cell dead compared to the particles only. PMID:16202162

Selective activation of group III metabotropic glutamate receptor subtypes produces different patterns of γ-aminobutyric acid immunoreactivity and glutamate release in the retina.

PubMed

Guimarães-Souza, E M; Calaza, K C

2012-12-01

Glutamate, the major excitatory neurotransmitter in the retina, functions by activation of both ionotropic (iGluR) and metabotropic (mGluR) glutamate receptors. Group III mGluRs, except for mGluR6, are mostly found in the inner plexiform layer (IPL), and their retinal functions are not well known. Therefore, we decided to investigate the effect of mGluRIII on glutamate release and GABAergic amacrine cells in the chick retina. The nonselective mGluRIII agonist L-SOP promoted a decrease in the number of γ-aminobutyric acid (GABA)-positive cells and in the GABA immunoreactivity in all sublayers of the IPL. This effect was prevented by the antagonist MAP-4, by GAT-1 inhibitor, and by antagonists of iGluR. Under the conditions used, L-SOP did not alter endogenous glutamate release. VU0155041, an mGluR4-positive allosteric modulator, reduced GABA immunoreactivity in amacrine cells and in sublayers 2 and 4 of the IPL but evoked an increase in the glutamate released. VU0155041's effect was inhibited by the absence of calcium. AMN082, a selective mGluR7-positive allosteric modulator, also decreased GABA immunoreactivity in amacrine cells and sublayers 1, 2, and 3 and increased glutamate release, and this effect was also inhibited by calcium absence. DCPG, an mGluR8-selective agonist, did not significantly alter GABA immunoreactivity in amacrine cells or glutamate release. However, it did significantly increase GABA immunoreactivity in sublayers 4 and 5. The results suggest that mGluRIIIs are involved in the modulation of glutamate and GABA release in the retina, possibly participating in distinct visual pathways: mGluR4 might be involved with cholinergic circuitry, whereas mGluR7 and mGluR8 might participate, respectively, in the OFF and the ON pathways. Copyright © 2012 Wiley Periodicals, Inc.

Multiple target of hAmylin on rat primary hippocampal neurons.

PubMed

Zhang, Nan; Yang, Shengchang; Wang, Chang; Zhang, Jianghua; Huo, Lifang; Cheng, Yiru; Wang, Chuan; Jia, Zhanfeng; Ren, Leiming; Kang, Lin; Zhang, Wei

2017-02-01

Alzheimer's disease (AD) and type II diabetes mellitus (DM2) are the most common aging-related diseases and are characterized by β-amyloid and amylin accumulation, respectively. Multiple studies have indicated a strong correlation between these two diseases. Amylin oligomerization in the brain appears to be a novel risk factor for developing AD. Although amylin aggregation has been demonstrated to induce cytotoxicity in neurons through altering Ca 2+ homeostasis, the underlying mechanisms have not been fully explored. In this study, we investigated the effects of amylin on rat hippocampal neurons using calcium imaging and whole-cell patch clamp recordings. We demonstrated that the amylin receptor antagonist AC187 abolished the Ca 2+ response induced by low concentrations of human amylin (hAmylin). However, the Ca 2+ response induced by higher concentrations of hAmylin was independent of the amylin receptor. This effect was dependent on extracellular Ca 2+ . Additionally, blockade of L-type Ca 2+ channels partially reduced hAmylin-induced Ca 2+ response. In whole-cell recordings, hAmylin depolarized the membrane potential. Moreover, application of the transient receptor potential (TRP) channel antagonist ruthenium red (RR) attenuated the hAmylin-induced increase in Ca 2+ . Single-cell RT-PCR demonstrated that transient receptor potential vanilloid 4 (TRPV4) mRNA was expressed in most of the hAmylin-responsive neurons. In addition, selective knockdown of TRPV4 channels inhibited the hAmylin-evoked Ca 2+ response. These results indicated that different concentrations of hAmylin act through different pathways. The amylin receptor mediates the excitatory effects of low concentrations of hAmylin. In contrast, for high concentrations of hAmylin, hAmylin aggregates precipitated on the neuronal membrane, activated TRPV4 channels and subsequently triggered membrane voltage-gated calcium channel opening followed by membrane depolarization. Therefore, our data suggest that TRPV4 is a key molecular mediator for the cytotoxic effects of hAmylin on hippocampal neurons. Copyright © 2016 Elsevier Ltd. All rights reserved.

Identification of a Potent Tryptophan-based TRPM8 Antagonist With in vivo Analgesic Activity.

PubMed

Bertamino, Alessia; Iraci, Nunzio; Ostacolo, Carmine; Ambrosino, Paolo; Musella, Simona; Di Sarno, Veronica; Ciaglia, Tania; Pepe, Giacomo; Sala, Marina; Soldovieri, Maria Virginia; Mosca, Ilaria; Gonzalez-Rodriguez, Sara; Fernández-Carvajal, Asia; Ferrer-Montiel, Antonio; Novellino, Ettore; Taglialatela, Maurizio; Campiglia, Pietro; Gomez-Monterrey, Isabel M

2018-06-25

TRPM8 has been implicated in nociception and pain and is currently regarded as an attractive target for the pharmacological treatment of neuropathic pain syndromes. A series of analogues of N,N'-dibenzyl tryptamine 1, a potent TRPM8 antagonist, were prepared and screened using a fluorescence-based in vitro assay based on menthol-evoked calcium influx in TRPM8 stably-transfected HEK293 cells. The tryptophan derivative 14 was identified as a potent (IC 50 0.2±0.2 nM) and selective TRPM8 antagonist. In vivo, 14 showed significant target coverage in both an icilin-induced WDS (at 1-30 mg/kg s.c.) and oxaliplatin-induced cold allodynia (at 0.1-1 μg s.c.) mice models. Molecular modeling studies identified the putative binding mode of these antagonists, suggesting that they could influence an interaction network between the S1-4 transmembrane segments and the TRP domains of the channel subunits. The tryptophan moiety provides a new pharmacophoric scaffold for the design of highly potent modulators of TRPM8-mediated pain.

Salicylate enables cochlear arachidonic-acid-sensitive NMDA receptor responses.

PubMed

Ruel, Jérôme; Chabbert, Christian; Nouvian, Régis; Bendris, Rim; Eybalin, Michel; Leger, Claude Louis; Bourien, Jérôme; Mersel, Marcel; Puel, Jean-Luc

2008-07-16

Currently, many millions of people treated for various ailments receive high doses of salicylate. Consequently, understanding the mechanisms by which salicylate induces tinnitus is an important issue for the research community. Behavioral testing in rats have shown that tinnitus induced by salicylate or mefenamate (both cyclooxygenase blockers) are mediated by cochlear NMDA receptors. Here we report that the synapses between the sensory inner hair cells and the dendrites of the cochlear spiral ganglion neurons express NMDA receptors. Patch-clamp recordings and two-photon calcium imaging demonstrated that salicylate and arachidonate (a substrate of cyclooxygenase) enabled the calcium flux and the neural excitatory effects of NMDA on cochlear spiral ganglion neurons. Salicylate also increased the arachidonate content of the whole cochlea in vivo. Single-unit recordings of auditory nerve fibers in adult guinea pig confirmed the neural excitatory effect of salicylate and the blockade of this effect by NMDA antagonist. These results suggest that salicylate inhibits cochlear cyclooxygenase, which increased levels of arachidonate. The increased levels of arachidonate then act on NMDA receptors to enable NMDA responses to glutamate that inner hair cells spontaneously release. This new pharmacological profile of salicylate provides a molecular mechanism for the generation of tinnitus at the periphery of the auditory system.

Changes in IP3 Receptor Expression and Function in Aortic Smooth Muscle of Atherosclerotic Mice

PubMed Central

Ewart, Marie-Ann; Ugusman, Azizah; Vishwanath, Anisha; Almabrouk, Tarek A.M.; Alganga, Husam; Katwan, Omar J.; Hubanova, Pavlina; Currie, Susan; Kennedy, Simon

2017-01-01

Peroxynitrite is an endothelium-independent vasodilator that induces relaxation via membrane hyperpolarization. The activation of IP3 receptors triggers the opening of potassium channels and hyperpolarization. Previously we found that relaxation to peroxynitrite was maintained during the development of atherosclerosis due to changes in the expression of calcium-regulatory proteins. In this study we investigated: (1) the mechanism of peroxynitrite-induced relaxation in the mouse aorta, (2) the effect of atherosclerosis on relaxation to peroxynitrite and other vasodilators, and (3) the effect of atherosclerosis on the expression and function of the IP3 receptor. Aortic function was studied using wire myography, and atherosclerosis was induced by fat-feeding ApoE−/− mice. The expression of IP3 receptors was studied using Western blotting and immunohistochemistry. Relaxation to peroxynitrite was attenuated by the IP3 antagonists 2-APB and xestospongin C and also the Kv channel blocker 4-aminopyridine (4-AP). Atherosclerosis attenuated vasodilation to cromakalim and the AMPK activator A769662 but not peroxynitrite. Relaxation was attenuated to a greater extent by 2-APB in atherosclerotic aortae despite the reduced expression of IP3 receptors. 4-AP was less effective in ApoE−/− mice fat-fed for 4 months. Peroxynitrite relaxation involves an IP3-induced calcium release and KV channel activation. This mechanism becomes less important as atherosclerosis develops, and relaxation to peroxynitrite may be maintained by increased calcium extrusion. PMID:28365690

1,4-Dihydropyridine Derivatives: Dihydronicotinamide Analogues—Model Compounds Targeting Oxidative Stress

PubMed Central

Velena, Astrida; Zarkovic, Neven; Gall Troselj, Koraljka; Bisenieks, Egils; Krauze, Aivars; Poikans, Janis; Duburs, Gunars

2016-01-01

Many 1,4-dihydropyridines (DHPs) possess redox properties. In this review DHPs are surveyed as protectors against oxidative stress (OS) and related disorders, considering the DHPs as specific group of potential antioxidants with bioprotective capacities. They have several peculiarities related to antioxidant activity (AOA). Several commercially available calcium antagonist, 1,4-DHP drugs, their metabolites, and calcium agonists were shown to express AOA. Synthesis, hydrogen donor properties, AOA, and methods and approaches used to reveal biological activities of various groups of 1,4-DHPs are presented. Examples of DHPs antioxidant activities and protective effects of DHPs against OS induced damage in low density lipoproteins (LDL), mitochondria, microsomes, isolated cells, and cell cultures are highlighted. Comparison of the AOA of different DHPs and other antioxidants is also given. According to the data presented, the DHPs might be considered as bellwether among synthetic compounds targeting OS and potential pharmacological model compounds targeting oxidative stress important for medicinal chemistry. PMID:26881016

Short infrared (IR) laser pulses can induce nanoporation

NASA Astrophysics Data System (ADS)

Roth, Caleb C.; Barnes, Ronald A.; Ibey, Bennett L.; Glickman, Randolph D.; Beier, Hope T.

2016-03-01

Short infrared (IR) laser pulses on the order of hundreds of microseconds to single milliseconds with typical wavelengths of 1800-2100 nm, have shown the capability to reversibly stimulate action potentials (AP) in neuronal cells. While the IR stimulation technique has proven successful for several applications, the exact mechanism(s) underlying the AP generation has remained elusive. To better understand how IR pulses cause AP stimulation, we determined the threshold for the formation of nanopores in the plasma membrane. Using a surrogate calcium ion, thallium, which is roughly the same shape and charge, but lacks the biological functionality of calcium, we recorded the flow of thallium ions into an exposed cell in the presence of a battery of channel antagonists. The entry of thallium into the cell indicated that the ions entered via nanopores. The data presented here demonstrate a basic understanding of the fundamental effects of IR stimulation and speculates that nanopores, formed in response to the IR exposure, play an upstream role in the generation of AP.

SMOC can act as both an antagonist and an expander of BMP signaling.

PubMed

Thomas, J Terrig; Eric Dollins, D; Andrykovich, Kristin R; Chu, Tehyen; Stultz, Brian G; Hursh, Deborah A; Moos, Malcolm

2017-03-21

The matricellular protein SMOC (Secreted Modular Calcium binding protein) is conserved phylogenetically from vertebrates to arthropods. We showed previously that SMOC inhibits bone morphogenetic protein (BMP) signaling downstream of its receptor via activation of mitogen-activated protein kinase (MAPK) signaling. In contrast, the most prominent effect of the Drosophila orthologue, pentagone ( pent ), is expanding the range of BMP signaling during wing patterning. Using SMOC deletion constructs we found that SMOC-∆EC, lacking the extracellular calcium binding (EC) domain, inhibited BMP2 signaling, whereas SMOC-EC (EC domain only) enhanced BMP2 signaling. The SMOC-EC domain bound HSPGs with a similar affinity to BMP2 and could expand the range of BMP signaling in an in vitro assay by competition for HSPG-binding. Together with data from studies in vivo we propose a model to explain how these two activities contribute to the function of Pent in Drosophila wing development and SMOC in mammalian joint formation.

The role of NMDA and mGluR5 receptors in calcium mobilization and neurotoxicity of homocysteine in trigeminal and cortical neurons and glial cells.

PubMed

Abushik, Polina A; Niittykoski, Minna; Giniatullina, Raisa; Shakirzyanova, Anastasia; Bart, Genevieve; Fayuk, Dmitriy; Sibarov, Dmitry A; Antonov, Sergei M; Giniatullin, Rashid

2014-04-01

Recent studies suggested contribution of homocysteine (HCY) to neurodegenerative disorders and migraine. However, HCY effect in the nociceptive system is essentially unknown. To explore the mechanism of HCY action, we studied short- and long-term effects of this amino acid on rat peripheral and central neurons. HCY induced intracellular Ca²⁺ transients in cultured trigeminal neurons and satellite glial cells (SGC), which were blocked by the NMDA antagonist AP-5 in neurons, but not in SGCs. In contrast, 3-((2-Methyl-4-thiazolyl)ethynyl)pyridine (MTEP), the metabotropic mGluR5 (metabotropic glutamate receptor 5 subtype) antagonist, preferentially inhibited Ca²⁺ transients in SGCs. Prolonged application of HCY induced apoptotic cell death of both kinds of trigeminal cells. The apoptosis was blocked by AP-5 or by the mGluR5 antagonist MTEP. Likewise, in cortical neurons, HCY-induced cell death was inhibited by AP-5 or MTEP. Imaging with 2',7'-dichlorodihydrofluorescein diacetate or mitochondrial dye Rhodamine-123 as well as thiobarbituric acid reactive substances assay did not reveal involvement of oxidative stress in the action of HCY. Thus, elevation of intracellular Ca²⁺ by HCY in neurons is mediated by NMDA and mGluR5 receptors while SGC are activated through the mGluR5 subtype. Long-term neurotoxic effects in peripheral and central neurons involved both receptor types. Our data suggest glutamatergic mechanisms of HCY-induced sensitization and apoptosis of trigeminal nociceptors. © 2013 International Society for Neurochemistry.

Hydrostatic Pressure–Induced Release of Stored Calcium in Cultured Rat Optic Nerve Head Astrocytes

PubMed Central

Mandal, Amritlal; Delamere, Nicholas A.

2010-01-01

Purpose. Elevated intraocular pressure is associated with glaucomatous optic nerve damage. Other investigators have shown functional changes in optic nerve head astrocytes subjected to elevated hydrostatic pressure (HP) for 1 to 5 days. Recently, the authors reported ERK1/2, p90RSK and NHE1 phosphorylation after 2 hours. Here they examine calcium responses at the onset of HP to determine what precedes ERK1/2 phosphorylation. Methods. Cytoplasmic calcium concentration ([Ca2+]i) was measured in cultured rat optic nerve astrocytes loaded with fura-2. The cells were placed in a closed imaging chamber and subjected to an HP increase of 15 mm Hg. Protein phosphorylation was detected by Western blot analysis. Results. The increase of HP caused an immediate slow increase in [Ca2+]i. The response persisted in calcium-free solution and when nickel chloride (4 mM) was added to suppress channel-mediated calcium entry. Previous depletion of the ER calcium stores by cyclopiazonic acid abolished the HP-induced calcium level increase. The HP-induced increase persisted in cells exposed to xestospongin C, an inhibitor of IP3R-mediated calcium release. In contrast, ryanodine receptor (RyR) antagonist ruthenium red (10 μM) or dantrolene (25 μM) inhibited the HP-induced calcium increase. The HP-induced calcium increase was abolished when ryanodine-sensitive calcium stores were pre-depleted with caffeine (3 mM). HP caused ERK1/2 phosphorylation. The magnitude of the ERK1/2 phosphorylation response was reduced by ruthenium red and dantrolene. Conclusions. Increasing HP causes calcium release from a ryanodine-sensitive cytoplasmic store and subsequent ERK1/2 activation. Calcium store release appears to be a required early step in the initial astrocyte response to an HP increase. PMID:20071675

[Primary and secondary arterial hypertension - update 2016].

PubMed

Sanner, Bernd; Hausberg, Martin

2016-06-01

In patients with hypertension without diabetes and with an increased risk of cardiovascular complications a blood pressure of below 130 mmHg should be targeted. Hypertensive patients with an age above 80 years should be treated in the same way as younger hypertensive patients if they are otherwise healthy and functionally independent. On the other hand frail elderly patients could have an increased morbidity and mortality with intensive blood pressure control. In patients with resistant hypertension spironolactone was the most effective drug when given in addition to their baseline drugs (ACE-inhibitor/angiotensin receptor antagonist, calcium channel blocker and thiazide diuretic). © Georg Thieme Verlag KG Stuttgart · New York.

Curine inhibits eosinophil activation and airway hyper-responsiveness in a mouse model of allergic asthma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ribeiro-Filho, Jaime; Laboratório de Imunofarmacologia, Departamento de Fisiologia e Patologia, UFPB, João Pessoa, Paraíba; Calheiros, Andrea Surrage

Allergic asthma is a chronic inflammatory airway disease with increasing prevalence around the world. Current asthma therapy includes drugs that usually cause significant side effects, justifying the search for new anti-asthmatic drugs. Curine is a bisbenzylisoquinoline alkaloid that modulates calcium influx in many cell types; however, its anti-allergic and putative toxic effects remain to be elucidated. Our aim was to investigate the effects of curine on eosinophil activation and airway hyper-responsiveness (AHR) and to characterize its potential toxic effects. We used a mouse model of allergic asthma induced by sensitization and challenge with ovalbumin (OVA) to evaluate the anti-allergic effectsmore » of oral treatment with curine. The oral administration of curine significantly inhibited eosinophilic inflammation, eosinophil lipid body formation and AHR in animals challenged with OVA compared with animals in the untreated group. The curine treatment also reduced eotaxin and IL-13 production triggered by OVA. Verapamil, a calcium channel antagonist, had similar anti-allergic properties, and curine pre-treatment inhibited the calcium-induced tracheal contractile response ex-vivo, suggesting that the mechanism by which curine exerts its effects is through the inhibition of a calcium-dependent response. A toxicological evaluation showed that orally administered curine did not significantly alter the biochemical, hematological, behavioral and physical parameters measured in the experimental animals compared with saline-treated animals. In conclusion, curine showed anti-allergic activity through mechanisms that involve inhibition of IL-13 and eotaxin and of Ca{sup ++} influx, without inducing evident toxicity and as such, has the potential for the development of anti-asthmatic drugs. - Highlights: • Curine is a bisbenzylisoquinoline alkaloid from Chondrodendron platyphyllum. • Curine inhibits eosinophil influx and activation and airway hyper-responsiveness. • Curine mechanisms involve inhibition of Ca{sup 2+} influx, and IL-13 and eotaxin secretion. • No significant toxicity was observed in mice orally treated with curine for 7 days. • Curine has the potential for the development of anti-asthmatic drugs.« less

Synthesis and Structure-Activity Relationships of (-)-cis-N-Normetazocine-Based LP1 Derivatives.

PubMed

Pasquinucci, Lorella; Parenti, Carmela; Amata, Emanuele; Georgoussi, Zafiroula; Pallaki, Paschalina; Camarda, Valeria; Calò, Girolamo; Arena, Emanuela; Montenegro, Lucia; Turnaturi, Rita

2018-05-05

(−)- cis - N -Normetazocine represents a rigid scaffold able to mimic the tyramine moiety of endogenous opioid peptides, and the introduction of different N -substituents influences affinity and efficacy of respective ligands at MOR (mu opioid receptor), DOR (delta opioid receptor), and KOR (kappa opioid receptor). We have previously identified LP1, a MOR/DOR multitarget opioid ligand, with an N -phenylpropanamido substituent linked to (−)- cis - N -Normetazocine scaffold. Herein, we report the synthesis, competition binding and calcium mobilization assays of new compounds 10 ⁻ 16 that differ from LP1 by the nature of the N -substituent. In radioligand binding experiments, the compounds 10 ⁻ 13 , featured by an electron-withdrawing or electron-donating group in the para position of phenyl ring, displayed improved affinity for KOR (K i = 0.85⁻4.80 μM) in comparison to LP1 (7.5 μM). On the contrary, their MOR and DOR affinities were worse (K i = 0.18⁻0.28 μM and K i = 0.38⁻1.10 μM, respectively) with respect to LP1 values (K i = 0.049 and 0.033 μM). Analogous trends was recorded for the compounds 14 ⁻ 16 , featured by indoline, tetrahydroquinoline, and diphenylamine functionalities in the N -substituent. In calcium mobilization assays, the compound 10 with a p -fluorophenyl in the N -substituent shared the functional profile of LP1 (pEC 50 MOR = 7.01), although it was less active. Moreover, the p -methyl- ( 11 ) and p -cyano- ( 12 ) substituted compounds resulted in MOR partial agonists and DOR/KOR antagonists. By contrast, the derivatives 13 ⁻ 15 resulted as MOR antagonists, and the derivative 16 as a MOR/KOR antagonist (pK B MOR = 6.12 and pK B KOR = 6.11). Collectively, these data corroborated the critical role of the N -substituent in (−)- cis - N -Normetazocine scaffold. Thus, the new synthesized compounds could represent a template to achieve a specific agonist, antagonist, or mixed agonist/antagonist functional profile.

Cardiovascular and diuretic activity of kaurene derivatives of Xylopia aethiopica and Alepidea amatymbica.

PubMed

Somova, L I; Shode, F O; Moodley, K; Govender, Y

2001-10-01

The extractives, crude and pure, of Alepidea amatymbica (AA) and Xylopia aethiopica (XA) were subjected to bioassay-directed phytochemical examination for potential cardiovascular and diuretic activity. All extractives and derivatives (XA/O, AA/1, xylopic acid, AA/3, AA/4, AA/5, AA/6, XA/1, XA/2, XA/3) displayed low toxicity, with LC(50) 0.5-5.0 ng/ml. For the first time, diterpene kaurenoids were reported to have significant systemic hypotensive and coronary vasodilatory effect accompanied with bradycardia. These effects were attributed to calcium antagonistic mechanism. The diuretic and natriuretic effects found were similar to the effects of chlorothiazide, suggesting inhibition of Na+ and K+ reabsorption in the early portion of the distal tubule. Further experiments are needed to elaborate the exact mechanisms of the hypotensive and diuretic effects of diterpene kaurenoids.

Effects of cilnidipine, a novel dihydropyridine calcium antagonist, on autonomic function, ambulatory blood pressure and heart rate in patients with essential hypertension.

PubMed

Minami, J; Kawano, Y; Makino, Y; Matsuoka, H; Takishita, S

2000-12-01

The aim of the present study was to evaluate the effects of cilnidipine, a novel dihydropyridine calcium antagonist, on autonomic function, ambulatory blood pressure and heart rate in patients with essential hypertension. Ten inpatients with mild to moderate essential hypertension (four men and six women; age: 44-64 years) underwent a drug-free period for 7 days and a treatment period with cilnidipine 10 mg orally for another 7 days, in a randomized crossover study. On the sixth day of each period, they underwent autonomic function tests including a mental arithmetic test, a cold pressor test and a Valsalva manoeuvre. After these tests, 24 h ambulatory blood pressure, heart rate, and the electrocardiogram R-R intervals were monitored every 30 min. A power spectral analysis of R-R intervals was performed to obtain the low-and high-frequency components. Cilnidipine significantly decreased the 24 h blood pressure by 6.5 +/- 1.7 mm Hg systolic (mean +/- s.e.mean; P < 0.01) and 5.0 +/- 1.1 mmHg diastolic (P < 0.01), whereas cilnidipine did not change heart rate or any indices of power spectral components. During the cold pressor test, the maximum change in systolic blood pressure and percentage changes in both systolic and diastolic blood pressures were significantly lower during the treatment period with cilnidipine than during the drug-free period. The baroreflex sensitivity measured from the overshoot phase of the Valsalva manoeuvre did not differ significantly between the two periods. Cilnidipine is effective as a once-daily antihypertensive agent and causes little influence on heart rate and the autonomic nervous system in patients with mild to moderate essential hypertension. Moreover, it is suggested that cilnidipine has an additional clinical benefit in the inhibition of the pressor response induced by acute cold stress.

Effect of BN 52021, a specific antagonist of platelet activating factor (PAF-acether), on calcium movements and phosphatidic acid production induced by PAF-acether in human platelets

DOE Office of Scientific and Technical Information (OSTI.GOV)

Simon, M.F.; Chap, H.; Braquet, P.

1987-02-15

/sup 32/P-labelled human platelets loaded with quin 2 and pretreated with aspirin were stimulated with 1-100 nM platelet activating factor (PAF-acether or 1-0-alkyl-2-acetyl-sn-glycero-3-phosphocholine) in a medium containing the ADP-scavenging system creatine phosphate/creatine phosphokinase. Under these conditions, PAF-acether evoked a characteristic fluorescence change allowing to quantify elevations in cytoplasmic free Ca/sup 2 +/ from internal stores (Ca/sup 2 +/ mobilization) or from external medium (Ca/sup 2 +/ influx), as well as an increased production of phosphatidic acid, reflecting phospholipase C activation. These effects, which can be attributed to PAF-acether only and not to released products such as ADP or thromboxane A2,more » were strongly inhibited in a dose-dependent manner by BN 52021, a specific antagonist of PAF-acether isolated from Ginkgo biloba. As the drug remained inactive against the same effects elicited by thrombin, it is concluded that BN 52021 does not interfere directly with the mechanism of transmembrane signalling involving inositol-phospholipids or (and) some putative receptor-operated channels, but rather acts on the binding of PAF-acether to its presumed membrane receptor.« less

Adrenomedullin increases the short-circuit current in the mouse seminal vesicle: actions on chloride secretion.

PubMed

Liao, S B; Cheung, K H; O, W S; Tang, Fai

2014-08-01

Adrenomedullin (ADM) may regulate seminal vesicle fluid secretion, and this may affect sperm quality. In this study, we investigated the effect of ADM on chloride secretion in the mouse seminal vesicle. The presence of ADM in mouse seminal vesicle was confirmed using immunostaining, and the molecular species was determined using gel filtration chromatography coupled with enzyme-linked assay for ADM. The effects of ADM on chloride secretion were studied by short-circuit current technique in a whole-mount preparation of mouse seminal vesicle in an Ussing chamber. The effects of specific ADM and calcitonin gene-related peptide (CGRP) receptor antagonists were investigated. Whether the ADM effect depended on the cAMP- and/or calcium-activated chloride channel was also studied using specific chloride channel blockers. The results showed that ADM was present in seminal vesicle epithelial cells. The major molecular species was precursor in the mouse seminal vesicle. ADM increased short-circuit current through the calcium-activated chloride channel in mouse seminal vesicle, and CGRP receptor was involved. We conclude that ADM may regulate chloride and fluid secretion from the seminal vesicle, which may affect the composition of the seminal plasma bathing the sperm and, hence, fertility. © 2014 by the Society for the Study of Reproduction, Inc.

RNA-seq analysis identifies potential modulators of gravity response in spores of Ceratopteris (Parkeriaceae): evidence for modulation by calcium pumps and apyrase activity.

PubMed

Bushart, Thomas J; Cannon, Ashley E; Ul Haque, Aeraj; San Miguel, Phillip; Mostajeran, Kathy; Clark, Gregory B; Porterfield, D Marshall; Roux, Stanley J

2013-01-01

Gravity regulates the magnitude and direction of a trans-cell calcium current in germinating spores of Ceratopteris richardii. Blocking this current with nifedipine blocks the spore's downward polarity alignment, a polarization that is fixed by gravity ∼10 h after light induces the spores to germinate. RNA-seq analysis at 10 h was used to identify genes potentially important for the gravity response. The data set will be valuable for other developmental and phylogenetic studies. De novo Newbler assembly of 958 527 reads from Roche 454 sequencing was executed. The sequences were identified and analyzed using in silico methods. The roles of endomembrane Ca(2+)-ATPase pumps and apyrases in the gravity response were further tested using pharmacological agents. Transcripts related to calcium signaling and ethylene biosynthesis were identified as notable constituents of the transcriptome. Inhibiting the activity of endomembrane Ca(2+)-ATPase pumps with 2,5-di-(t-butyl)-1,4-hydroquinone diminished the trans-cell current, but increased the orientation of the polar axis to gravity. The effects of applied nucleotides and purinoceptor antagonists gave novel evidence implicating extracellular nucleotides as regulators of the gravity response in these fern spores. In addition to revealing general features of the transcriptome of germinating spores, the results highlight a number of calcium-responsive and light-receptive transcripts. Pharmacologic assays indicate endomembrane Ca(2+)-ATPases and extracellular nucleotides may play regulatory roles in the gravity response of Ceratopteris spores.

Extracellular cyclic ADP-ribose potentiates ACh-induced contraction in bovine tracheal smooth muscle.

PubMed

Franco, L; Bruzzone, S; Song, P; Guida, L; Zocchi, E; Walseth, T F; Crimi, E; Usai, C; De Flora, A; Brusasco, V

2001-01-01

Cyclic ADP-ribose (cADPR), a universal calcium releaser, is generated from NAD(+) by an ADP-ribosyl cyclase and is degraded to ADP-ribose by a cADPR hydrolase. In mammals, both activities are expressed as ectoenzymes by the transmembrane glycoprotein CD38. CD38 was identified in both epithelial cells and smooth myocytes isolated from bovine trachea. Intact tracheal smooth myocytes (TSMs) responded to extracellular cADPR (100 microM) with an increase in intracellular calcium concentration ([Ca(2+)](i)) both at baseline and after acetylcholine (ACh) stimulation. The nonhydrolyzable analog 3-deaza-cADPR (10 nM) elicited the same effects as cADPR, whereas the cADPR antagonist 8-NH(2)-cADPR (10 microM) inhibited both basal and ACh-stimulated [Ca(2+)](i) levels. Extracellular cADPR or 3-deaza-cADPR caused a significant increase of ACh-induced contraction in tracheal smooth muscle strips, whereas 8-NH(2)-cADPR decreased it. Tracheal mucosa strips, by releasing NAD(+), enhanced [Ca(2+)](i) in isolated TSMs, and this increase was abrogated by either NAD(+)-ase or 8-NH(2)-cADPR. These data suggest the existence of a paracrine mechanism whereby mucosa-released extracellular NAD(+) plays a hormonelike function and cADPR behaves as second messenger regulating calcium-related contractility in TSMs.

State-of-the-art treatment of hypertension: established and new drugs.

PubMed

Burnier, Michel; Vuignier, Yann; Wuerzner, Gregoire

2014-03-01

The treatment of essential hypertension is based essentially on the prescription of four major classes of antihypertensive drugs, i.e. blockers of the renin--angiotensin system, calcium channel blockers, diuretics and beta-blockers. In recent years, very few new drug therapies of hypertension have become available. Therefore, it is crucial for physicians to optimize their antihypertensive therapies with the drugs available on the market. In each of the classes of antihypertensive drugs, questions have recently been raised: are angiotensin-converting enzyme (ACE) inhibitors superior to angiotensin II receptor blockers (ARB)? Is it possible to reduce the incidence of peripheral oedema with calcium antagonists? Is hydrochlorothiazide really the good diuretic to use in combination therapies? The purpose of this review is to discuss these various questions in the light of the most recent clinical studies and meta-analyses. These latter suggest that ACE inhibitors and ARB are equivalent except for a better tolerability profile of ARB. Third generation calcium channel blockers enable to reduce the incidence of peripheral oedema and chlorthalidone is certainly more effective than hydrochlorothiazide in preventing cardiovascular events in hypertension. At last, studies suggest that drug adherence and long-term persistence under therapy is one of the major issues in the actual management of essential hypertension.

In vitro and in vivo characterization of a calcium modulator of the diphenylalkylamine type with selective coronary dilatory properties.

PubMed

Caldirola, P; Monteil, A; Zandberg, P; Mannhold, R; Timmerman, H

1997-11-01

VUF 8929 (N-¿2-[bis(p-fluorophenyl)methoxy]ethyl¿-(2-phenyl)ethylamine maleate, CAS 140890-71-7) is a diphenylalkylamine derivative structurally related to prenylamine. The calcium antagonistic properties of this compound have been studied in in vitro and in vivo systems. VUF 8929 has affinity for the voltage-operated calcium channel. Its pKD for the displacement of [3H]nitrendipine bound to cerebral rat cortex is 6.27 (+/- 0.17). The compound influences the [3H]nitrendipine binding through an allosteric interaction with a site adjacent to the dihydropyridine binding site. Competitive experiments with the additional presence of the phenylalkylamine gallopamil showed that this allosteric site is a property common to diphenyl- and phenylalkylamines. It was further observed that VUF 8929 has a high affinity for calmodulin as it shows high potency in inhibiting the calmodulin mediated activation of PDE. The inhibition of K+ (IC50 0.5 mumol/l)- and noradrenaline (IC50 1.3 mumol/l)-induced contractions of rabbit aorta rings was in the same concentration range as found for the calmodulin inhibitory activity. In vitro platelet aggregation was also inhibited in the same concentration range when washed platelets were used. Thus calmodulin antagonism may contribute to the observed effects on aorta ring contractions and platelets aggregation. Platelet aggregation, however, in media in which albumin was added or in platelet rich plasma was not affected. It is assumed that due to the high lipophilicity, common to many diphenylalkylamines, VUF 8929 has a strong binding to plasma proteins. This may also explain why orally administered VUF 8929 did not affect the alpha 2-induced pressor response in pithed rats and the ex vivo collagen induced aggregation response. The haemodynamic profile in anaesthetized dogs showed that intravenous injected VUF 8929 reduced the workload of the heart while coronary blood flow increases at a dose of 0.3 mg/kg. Reversible occlusion of the coronary artery, which leads to S-T segment elevation and local venous acidosis, were reduced by VUF 8929 indicating that the compound has anti-ischaemic properties. VUF 8929 is a calcium antagonist which has anti-ischaemic properties, reduces the workload of the heart and increases coronary flow. Due to these properties VUF 8929 is a potential cardioprotective agent.

Synthesis, characterization, and in vitro evaluation of the selective P2Y2 receptor antagonist AR-C118925.

PubMed

Rafehi, Muhammad; Burbiel, Joachim C; Attah, Isaac Y; Abdelrahman, Aliaa; Müller, Christa E

2017-03-01

The G q protein-coupled, ATP- and UTP-activated P2Y 2 receptor is a potential drug target for a range of different disorders, including tumor metastasis, inflammation, atherosclerosis, kidney disorders, and osteoporosis, but pharmacological studies are impeded by the limited availability of suitable antagonists. One of the most potent and selective antagonists is the thiouracil derivative AR-C118925. However, this compound was until recently not commercially available and little is known about its properties. We therefore developed an improved procedure for the synthesis of AR-C118925 and two derivatives to allow up-scaling and assessed their potency in calcium mobilization assays on the human and rat P2Y 2 receptors recombinantly expressed in 1321N1 astrocytoma cells. The compound was further evaluated for inhibition of P2Y 2 receptor-induced β-arrestin translocation. AR-C118925 behaved as a competitive antagonist with pA 2 values of 37.2 nM (calcium assay) and 51.3 nM (β-arrestin assay). Selectivity was assessed vs. related receptors including P2X, P2Y, and adenosine receptor subtypes, as well as ectonucleotidases. AR-C118925 showed at least 50-fold selectivity against the other investigated targets, except for the P2X1 and P2X3 receptors which were blocked by AR-C118925 at concentrations of about 1 μM. AR-C118925 is soluble in buffer at pH 7.4 (124 μM) and was found to be metabolically highly stable in human and mouse liver microsomes. In Caco2 cell experiments, the compound displayed moderate permeability indicating that it may show limited peroral bioavailability. AR-C118925 appears to be a useful pharmacological tool for in vitro and in vivo studies.

Contrast media-induced nephrotoxicity--questions and answers.

PubMed

Morcos, S K

1998-04-01

The intravascular administration of contrast media (CM) can produce acute haemodynamic changes in the kidney characterized by an increase in renal vascular resistance and a decrease in the glomerular filtration rate (GFR). These changes may lead to clinically significant reduction in renal function in patients with pre-existing risk factors such as diabetic nephropathy, congestive heart failure and dehydration. The pathophysiology of the renal haemodynamic effects of CM involves activation of the tubuloglomerular feedback (TGF) mechanism and the modulation of the intrarenal production of vasoactive mediators such as prostaglandins, nitric oxide, endothelin and adenosine. The TGF response is osmolality-dependent and accounts for about 50% of the acute functional effects of high osmolar CM on the kidney. Reduction in the synthesis of the endogenous vasodilators nitric oxide and prostaglandins increases the nephrotoxicity of CM. Endothelin and adenosine play a crucial role in mediating the acute functional effects of CM. Antagonists of these mediators attenuate the reduction in renal function induced by contrast agents. Vacuolization of the cells of the proximal tubules and necrosis of those of the medullary ascending limbs of loops of Henle are the main structural effects of CM in the kidney. The reduction in renal function induced by CM could be minimized by the use of low osmolar CM and adequate hydration. The prophylactic administration of calcium channel blockers and adenosine antagonists such as theophylline may also offer some protective effect.

Pharmacologic modification of the cytotoxic effects of cadmium in LLC-PK sub 1 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Battaglia, D.R.; Kahan, B.S.; Niewenhuis, R.J.

1989-02-09

Recent results from our laboratories have shown that exposure to cadmium causes LLC-PK{sub 1} cells to shrink, detach and assume a spherical shape. The purpose of the present studies was to determine whether various pharmacologic agents can reduce or prevent these cytotoxic effects of Cd{sup 2+}. Confluent monolayers of LLC-PK{sub 1} cells were incubated with the drugs of interest (50 microM final concentration) for 2 hours. CadCl{sub 2} (final concentration = 75 microM) was then added and the cells were incubated for another 20 hours. Morphologic changes were assessed qualitatively by viewing the cells with a phase contrast microscope. Themore » extent of Cd{sup 2+}-induced cellular damage was also quantified by staining the cells that remained on the growing surface with methylene blue, solubilizing the stained cells, and determining the absorbance at 660 nm. The results showed that several drugs, particularly the calmodulin antagonists trifluoperazine chlorpromazine, and the calcium channel blocker verapamil, significant reduced the severity of Cd{sup 2+}-induced cytotoxicity. By contrast, a variety of other agents, such as chlorpromazine sulfoxide, trifluoperazine sulfoxide, phenytoin and zinc, had no such protective effect. These findings indicate that Ca{sup 2+} antagonists can attenuate the cytotoxic effects of Cd{sup 2+} and that Cd{sup 2+} may produce some of its effects by activating Ca{sup 2+} -dependent systems.« less

DDPH ameliorated oxygen and glucose deprivation-induced injury in rat hippocampal neurons via interrupting Ca2+ overload and glutamate release.

PubMed

He, Zhi; Lu, Qing; Xu, Xulin; Huang, Lin; Chen, Jianguo; Guo, Lianjun

2009-01-28

Our previous work has demonstrated that DDPH (1-(2, 6-dimethylphenoxy)-2-(3, 4-dimethoxyphenylethylamino) propane hydrochloride), a competitive alpha(1)-adrenoceptor antagonist, could improve cognitive deficits, reduce histopathological damage and facilitate synaptic plasticity in vivo possibly via increasing NR2B (NMDA receptor 2B) expression and antioxidation of DDPH itself. The present study further evaluated effects of DDPH on OGD (Oxygen and glucose deprivation)-induced neuronal damage in rat primary hippocampal cells. The addition of DDPH to the cultured cells 12 h before OGD for 4 h significantly reduced neuronal damage as determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and LDH (lactate dehydrogenase) release experiments. The effects of DDPH on intracellular calcium concentration were explored by Fura-2 based calcium imaging techniques and results showed that DDPH at the dosages of 5 microM and 10 microM suppressed the increase of intracellular calcium ([Ca(2+)](i)) stimulated by 50 mM KCl in Ca(2+)-containing extracellular solutions. However, DDPH couldn't suppress the increase of [Ca(2+)](i) induced by both 50 microM glutamate in Ca(2+)-containing extracellular solutions and 20 microM ATP (Adenosine Triphosphate) in Ca(2+)-free solution. These results indicated that DDPH prevented [Ca(2+)](i) overload in hippocampal neurons by blocking Ca(2+) influx (voltage-dependent calcium channel) but not Ca(2+) mobilization from the intracellular Ca(2+) store in endoplasm reticulum (ER). We also demonstrated that DDPH could decrease glutamate release when hippocampal cells were subjected to OGD. These observations demonstrated that DDPH protected hippocampal neurons against OGD-induced damage by preventing the Ca(2+) influx and decreasing glutamate release.

Induction of oxidative stress causes functional alterations in mouse urothelium via a TRPM8-mediated mechanism: implications for aging.

PubMed

Nocchi, Linda; Daly, Donna M; Chapple, Christopher; Grundy, David

2014-06-01

The incidence of bladder conditions such as overactive bladder syndrome and its associated urinary incontinence is highly prevalent in the elderly. However, the mechanisms underlying these disorders are unclear. Studies suggest that the urothelium forms a 'sensory network' with the underlying innervation, alterations in which, could compromise bladder function. As the accumulation of reactive oxygen species can cause functional alterations with age, the aim of this study was to investigate whether oxidative stress alters urothelial sensory signalling and whether the mechanism underlying the effect of oxidative stress on the urothelium plays a role in aging. Five-month-old(young) and 24-month-old (aged) mice were used. H2O2 , used to induce oxidative stress, resulted in an increase in bladder afferent nerve activity and urothelial intracellular calcium in preparations from young mice. These functional changes were concurrent with upregulation of TRPM8 in the urothelium. Moreover, application of a TRPM8 antagonist significantly attenuated the H2O2 -induced calcium responses. Interestingly, an upregulation of TRPM8 was also found in the urothelium from aged mice, where high oxidative stress levels were observed, together with a greater calcium response to the TRPM8 agonist WS12. Furthermore, these calcium responses were attenuated by pretreatment with the antioxidant N-acetyl-cysteine. This study shows that oxidative stress affects urothelial function involving a TRPM8-mediated mechanism and these effects may have important implications for aging. These data provide an insight into the possible mechanisms by which oxidative stress causes physiological alterations in the bladder, which may also occur in other organs susceptible to aging. © 2014 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

Induction of oxidative stress causes functional alterations in mouse urothelium via a TRPM8-mediated mechanism: implications for aging

PubMed Central

Nocchi, Linda; Daly, Donna M; Chapple, Christopher; Grundy, David

2014-01-01

The incidence of bladder conditions such as overactive bladder syndrome and its associated urinary incontinence is highly prevalent in the elderly. However, the mechanisms underlying these disorders are unclear. Studies suggest that the urothelium forms a ‘sensory network’ with the underlying innervation, alterations in which, could compromise bladder function. As the accumulation of reactive oxygen species can cause functional alterations with age, the aim of this study was to investigate whether oxidative stress alters urothelial sensory signalling and whether the mechanism underlying the effect of oxidative stress on the urothelium plays a role in aging. Five-month-old(young) and 24-month-old (aged) mice were used. H2O2, used to induce oxidative stress, resulted in an increase in bladder afferent nerve activity and urothelial intracellular calcium in preparations from young mice. These functional changes were concurrent with upregulation of TRPM8 in the urothelium. Moreover, application of a TRPM8 antagonist significantly attenuated the H2O2-induced calcium responses. Interestingly, an upregulation of TRPM8 was also found in the urothelium from aged mice, where high oxidative stress levels were observed, together with a greater calcium response to the TRPM8 agonist WS12. Furthermore, these calcium responses were attenuated by pretreatment with the antioxidant N-acetyl-cysteine. This study shows that oxidative stress affects urothelial function involving a TRPM8-mediated mechanism and these effects may have important implications for aging. These data provide an insight into the possible mechanisms by which oxidative stress causes physiological alterations in the bladder, which may also occur in other organs susceptible to aging. PMID:24593692

Homology-guided mutational analysis reveals the functional requirements for antinociceptive specificity of collapsin response mediator protein 2-derived peptides.

PubMed

Moutal, Aubin; Li, Wennan; Wang, Yue; Ju, Weina; Luo, Shizhen; Cai, Song; François-Moutal, Liberty; Perez-Miller, Samantha; Hu, Jackie; Dustrude, Erik T; Vanderah, Todd W; Gokhale, Vijay; Khanna, May; Khanna, Rajesh

2017-02-05

N-type voltage-gated calcium (Ca v 2.2) channels are critical determinants of increased neuronal excitability and neurotransmission accompanying persistent neuropathic pain. Although Ca v 2.2 channel antagonists are recommended as first-line treatment for neuropathic pain, calcium-current blocking gabapentinoids inadequately alleviate chronic pain symptoms and often exhibit numerous side effects. Collapsin response mediator protein 2 (CRMP2) targets Ca v 2.2 channels to the sensory neuron membrane and allosterically modulates their function. A 15-amino-acid peptide (CBD3), derived from CRMP2, disrupts the functional protein-protein interaction between CRMP2 and Ca v 2.2 channels to inhibit calcium influx, transmitter release and acute, inflammatory and neuropathic pain. Here, we have mapped the minimal domain of CBD3 necessary for its antinociceptive potential. Truncated as well as homology-guided mutant versions of CBD3 were generated and assessed using depolarization-evoked calcium influx in rat dorsal root ganglion neurons, binding between CRMP2 and Ca v 2.2 channels, whole-cell voltage clamp electrophysiology and behavioural effects in two models of experimental pain: post-surgical pain and HIV-induced sensory neuropathy induced by the viral glycoprotein 120. The first six amino acids within CBD3 accounted for all in vitro activity and antinociception. Spinal administration of a prototypical peptide (TAT-CBD3-L5M) reversed pain behaviours. Homology-guided mutational analyses of these six amino acids identified at least two residues, Ala1 and Arg4, as being critical for antinociception in two pain models. These results identify an antinociceptive scaffold core in CBD3 that can be used for development of low MW mimetics of CBD3. © 2017 The British Pharmacological Society.

Neurotransmitter modulation of extracellular H+ fluxes from isolated retinal horizontal cells of the skate

PubMed Central

Molina, Anthony J A; Verzi, Michael P; Birnbaum, Andrea D; Yamoah, Ebenezer N; Hammar, Katherine; Smith, Peter J S; Malchow, Robert Paul

2004-01-01

Self-referencing H+-selective microelectrodes were used to measure extracellular H+ fluxes from horizontal cells isolated from the skate retina. A standing H+ flux was detected from quiescent cells, indicating a higher concentration of free hydrogen ions near the extracellular surface of the cell as compared to the surrounding solution. The standing H+ flux was reduced by removal of extracellular sodium or application of 5-(N-ethyl-N-isopropyl) amiloride (EIPA), suggesting activity of a Na+–H+ exchanger. Glutamate decreased H+ flux, lowering the concentration of free hydrogen ions around the cell. AMPA/kainate receptor agonists mimicked the response, and the AMPA/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) eliminated the effects of glutamate and kainate. Metabotropic glutamate agonists were without effect. Glutamate-induced alterations in H+ flux required extracellular calcium, and were abolished when cells were bathed in an alkaline Ringer solution. Increasing intracellular calcium by photolysis of the caged calcium compound NP-EGTA also altered extracellular H+ flux. Immunocytochemical localization of the plasmalemma Ca2+–H+-ATPase (PMCA pump) revealed intense labelling within the outer plexiform layer and on isolated horizontal cells. Our results suggest that glutamate modulation of H+ flux arises from calcium entry into cells with subsequent activation of the plasmalemma Ca2+–H+-ATPase. These neurotransmitter-induced changes in extracellular pH have the potential to play a modulatory role in synaptic processing in the outer retina. However, our findings argue against the hypothesis that hydrogen ions released by horizontal cells normally act as the inhibitory feedback neurotransmitter onto photoreceptor synaptic terminals to create the surround portion of the centre-surround receptive fields of retinal neurones. PMID:15272044

Modulation of intracellular Ca2+ via L-type calcium channels in heart cells by the autoantibody directed against the second extracellular loop of the alpha1-adrenoceptors.

PubMed

Bkaily, Ghassan; El-Bizri, Nesrine; Bui, Michel; Sukarieh, Rami; Jacques, Danielle; Fu, Michael L X

2003-03-01

The effects of methoxamine, a selective alpha1-adrenergic receptor agonist, and the autoantibody directed against the second extracellular loop of alpha1-adrenoceptors were studied on intracellular free Ca2+ levels using confocal microscopy and ionic currents using the whole-cell patch clamp technique in single cells of 10-day-old embryonic chick and 20-week-old fetal human hearts. We observed that like methoxamine, the autoantibody directed against the second extracellular loop of alpha1-adrenoreceptors significantly increased the L-type calcium current (I(Ca(L))) but had no effect on the T-type calcium current (I(Ca(T))), the delayed outward potassium current, or the fast sodium current. This effect of the autoantibody was prevented by a prestimulation of the receptors with methoxamine and vice versa. Moreover, treating the cells with prazosin, a selective alpha1-adrenergic receptor antagonist blocked the methoxamine and the autoantibody-induced increase in I(Ca(L)), respectively. In absence of prazosin, both methoxamine and the autoantibody showed a substantial enhancement in the frequency of cell contraction and that of the concomitant cytosolic and nuclear free Ca2+ variations. The subsequent addition of nifedipine, a specific L-type Ca2+ channel blocker, reversed not only the methoxamine or the autoantibody-induced effect but also completely abolished cell contraction. These results demonstrated that functional alpha1-adrenoceptors exist in both 10-day-old embryonic chick and 20-week-old human fetal hearts and that the autoantibody directed against the second extracellular loop of this type of receptors plays an important role in stimulating their activity via activation of L-type calcium channels. This loop seems to have a functional significance by being the target of alpha1-receptor agonists like methoxamine.

A second mechanism of increase of cerebellar hypermetria in humans

PubMed Central

Manto, Mario-Ubaldo; Bosse, Pierre

2003-01-01

So far, there is only one procedure known to increase hypermetria in cerebellar patients. Facing an increased inertia of the moving limb, patients presenting a lesion of the lateral cerebellum are able to increase appropriately the intensity of the agonist electromyographic (EMG) activity (the launching force), but are unable to adapt the intensity of the antagonist activity (the braking force). As a result, hypermetria is larger when the inertial load is artificially increased. Recent studies have demonstrated that hyperventilation increases hypermetria in patients presenting a spinocerebellar ataxia type 6 (SCA 6), a disorder associated with polyglutamine expansions in the α1A-voltage-dependent calcium channel. The mechanism of this increase of hypermetria has not been identified so far. In the present work, we combined kinematic, EMG and transcranial Doppler studies to understand the effects of hyperventilation on fast goal-directed movements in patients presenting a SCA 6. Both in the normal mechanical state and after increasing the inertial load of the moving hand, hyperventilation induced an increase of hypermetria. Hyperventilation increased the delay of the onset latency of the antagonist EMG activity and decreased the rate of rise of both the agonist and the antagonist EMG activities. Hyperventilation induced a marked decrease in cerebral blood flow velocities. The mechanism of this provocative test is original and is distinct from the mechanism of the load-induced increase of hypermetria. PMID:12588903

Comparative risk-benefit assessment of drugs used in the management of hypertension in pregnancy.

PubMed

Kyle, P M; Redman, C W

1992-01-01

Antihypertensive treatment in pregnancy is needed to protect the mother from the dangers of severe hypertension (greater than or equal to 170/110mm Hg), particularly cerebral haemorrhage in the context of preeclampsia. There is no evidence that treatment of the hypertension confers any other benefit; the onset and progression of preeclampsia is neither prevented nor ameliorated. Therefore, there are no indications for treating mild-to-moderate hypertension (140 to 169/90 to 109mm Hg). Intravenous hydralazine and oral nifedipine are effective drugs to treat severe hypertension acutely, the latter having the advantage of ease of administration. For long term therapy, methyldopa is the only drug which has been fully assessed and shown to be safe for the neonate and infant. beta-Adrenoceptor antagonists are safe to use in the third trimester but cause significant intrauterine growth retardation when used for longer periods. ACE inhibitors are contraindicated and diuretics should be avoided. Although calcium antagonists appear to have much potential they require further assessment of their use in pregnancy.

Proteomic analysis of human bladder epithelial cells by 2D blue native SDS-PAGE reveals TCDD-induced alterations of calcium and iron homeostasis possibly mediated by nitric oxide.

PubMed

Verma, Nisha; Pink, Mario; Petrat, Frank; Rettenmeier, Albert W; Schmitz-Spanke, Simone

2015-01-02

A proteomic analysis of the interaction among multiprotein complexes involved in 2,3,7,8-dibenzo-p-dioxin (TCDD)-mediated toxicity in urinary bladder epithelial RT4 cells was performed using two-dimensional blue native SDS-PAGE (2D BN/SDS-PAGE). To enrich the protein complexes, unexposed and TCDD-exposed cells were fractionated. BN/SDS-PAGE of the resulting fractions led to an effective separation of proteins and protein complexes of various origins, including cell membrane, mitochondria, and other intracellular compartments. Major differences between the proteome of control and exposed cells involved the alteration of many calcium-regulated proteins (calmodulin, protein S100-A2, annexin A5, annexin A10, gelsolin isoform b) and iron-regulated proteins (ferritin, heme-binding protein 2, transferrin). On the basis of these findings, the intracellular calcium concentration was determined, revealing a significant increase after 24 h of exposure to TCDD. Moreover, the concentration of the labile iron pool (LIP) was also significantly elevated in TCDD-exposed cells. This increase was strongly inhibited by the calmodulin (CaM) antagonist W-7, which pointed toward a possible interaction between iron and calcium signaling. Because nitric oxide (NO) production was significantly enhanced in TCDD-exposed cells and was also inhibited by W-7, we hypothesize that alterations in calcium and iron homeostasis upon exposure to TCDD may be linked through NO generated by CaM-activated nitric oxide synthase. In our model, we propose that NO produced upon TCDD exposure interacts with the iron centers of iron-regulatory proteins (IRPs) that modulate the alteration of ferritin and transferrin, resulting in an augmented cellular LIP and, hence, increased toxicity.

Properties of Lactobacillus reuteri chitosan-calcium-alginate encapsulation under simulated gastrointestinal conditions.

PubMed

Huang, Hui-Ying; Tang, Yi-Ju; King, V An-Erl; Chou, Jen-Wei; Tsen, Jen-Horng

2015-03-01

The protective effects of encapsulation on the survival of Lactobacillus reuteri and the retention of the bacterium's probiotic properties under simulated gastrointestinal conditions were investigated. Viable counts and the remaining probiotic properties of calcium (Ca)-alginate encapsulated (A group), chitosan-Ca-alginate encapsulated (CA group), and unencapsulated, free L. reuteri (F group) were determined. Encapsulation improved the survival of L. reuteri subjected to simulated gastrointestinal conditions, with the greatest protective effect achieved in the CA group. The degree of cell membrane injury increased with increasing bile salt concentrations at constant pH, but the extent of injury was less in the encapsulated than in the free cells. Adherence rates were, in descending order: CA (0.524%)>A (0.360%)>F (0.275%). Lactobacillus reuteri cells retained their antagonistic activity toward Listeria monocytogenes even after incubation of the lactobacilli under simulated gastrointestinal conditions. Displacement of the pathogen by cells released from either of the encapsulation matrices was higher than that by free cells. The safety of L. reuteri was demonstrated in an in vitro invasion assay. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

Prostaglandin D2 regulates human colonic ion transport via the DP1 receptor.

PubMed

Medani, M; Collins, D; Mohan, H M; Walsh, E; Winter, D C; Baird, A W

2015-02-01

Prostaglandin D2 is released by mast cells and is important in allergies. Its role in gastrointestinal function is not clearly defined. This study aimed to determine the effect of exogenous PGD2 on ion transport in ex vivo normal human colonic mucosa. Mucosal sheets were mounted in Ussing chambers and voltage clamped to zero electric potential. Ion transport was quantified as changes in short-circuit current. In separate experiments epithelial monolayers or colonic crypts, isolated by calcium chelation, were treated with PGD2 and cAMP levels determined by ELISA or calcium levels were determined by fluorimetry. PGD2 caused a sustained, concentration-dependent rise in short-circuit current by increasing chloride secretion (EC50=376nM). This effect of PGD2 is mediated by the DP1 receptor, as the selective DP1 receptor antagonist BW A686C inhibited PGD2-induced but not PGE2-induced rise in short-circuit current. PGD2 also increased intracellular cAMP in isolated colonic crypts with no measurable influence on cytosolic calcium. PGD2 induces chloride secretion in isolated human colonic mucosa in a concentration-dependent manner with concomitant elevation of cytoplasmic cAMP in epithelial cells. The involvement of DP2 receptor subtypes has not previously been considered in regulation of ion transport in human intestine. Since inflammatory stimuli may induce production of eicosanoids, selective regulation of these pathways may be pivotal in determining therapeutic strategies and in understanding disease. Copyright © 2014. Published by Elsevier Inc.

Design, synthesis and biological evaluation of non-peptide PAR1 thrombin receptor antagonists based on small bifunctional templates: arginine and phenylalanine side chain groups are keys for receptor activity.

PubMed

Androutsou, Maria-Eleni; Saifeddine, Mahmoud; Hollenberg, Morley D; Matsoukas, John; Agelis, George

2010-04-01

In the present study, we report the synthesis and biological evaluation of a series of new non-peptide PAR(1) mimetic receptor antagonists, based on conformational analysis of the S(42)FLLR(46) tethered ligand (TL) sequence of PAR(1). These compounds incorporate the key pharmacophore groups in the TL sequence, guanidyl, amino and phenyl, which are essential for triggering receptor activity. Compounds 5 and 15 (50-100 microM) inhibited both TFLLR-amide (10 microM) and thrombin-mediated (0.5 and 1 U/ml; 5 and 10 microM) calcium signaling in a cultured human HEK cell assay.

DREAM Mediated Regulation of GCM1 in the Human Placental Trophoblast

PubMed Central

Baczyk, Dora; Kibschull, Mark; Mellstrom, Britt; Levytska, Khrystyna; Rivas, Marcos; Drewlo, Sascha; Lye, Stephen J.; Naranjo, Jose R.; Kingdom, John C. P.

2013-01-01

The trophoblast transcription factor glial cell missing-1 (GCM1) regulates differentiation of placental cytotrophoblasts into the syncytiotrophoblast layer in contact with maternal blood. Reduced placental expression of GCM1 and abnormal syncytiotrophoblast structure are features of hypertensive disorder of pregnancy – preeclampsia. In-silico techniques identified the calcium-regulated transcriptional repressor – DREAM (Downstream Regulatory Element Antagonist Modulator) - as a candidate for GCM1 gene expression. Our objective was to determine if DREAM represses GCM1 regulated syncytiotrophoblast formation. EMSA and ChIP assays revealed a direct interaction between DREAM and the GCM1 promoter. siRNA-mediated DREAM silencing in cell culture and placental explant models significantly up-regulated GCM1 expression and reduced cytotrophoblast proliferation. DREAM calcium dependency was verified using ionomycin. Furthermore, the increased DREAM protein expression in preeclamptic placental villi was predominantly nuclear, coinciding with an overall increase in sumolylated DREAM and correlating inversely with GCM1 levels. In conclusion, our data reveal a calcium-regulated pathway whereby GCM1-directed villous trophoblast differentiation is repressed by DREAM. This pathway may be relevant to disease prevention via calcium-supplementation. PMID:23300953

Hydrogen-Deuterium Exchange Mass Spectrometry Reveals Calcium Binding Properties and Allosteric Regulation of Downstream Regulatory Element Antagonist Modulator (DREAM).

PubMed

Zhang, Jun; Li, Jing; Craig, Theodore A; Kumar, Rajiv; Gross, Michael L

2017-07-18

Downstream regulatory element antagonist modulator (DREAM) is an EF-hand Ca 2+ -binding protein that also binds to a specific DNA sequence, downstream regulatory elements (DRE), and thereby regulates transcription in a calcium-dependent fashion. DREAM binds to DRE in the absence of Ca 2+ but detaches from DRE under Ca 2+ stimulation, allowing gene expression. The Ca 2+ binding properties of DREAM and the consequences of the binding on protein structure are key to understanding the function of DREAM. Here we describe the application of hydrogen-deuterium exchange mass spectrometry (HDX-MS) and site-directed mutagenesis to investigate the Ca 2+ binding properties and the subsequent conformational changes of full-length DREAM. We demonstrate that all EF-hands undergo large conformation changes upon calcium binding even though the EF-1 hand is not capable of binding to Ca 2+ . Moreover, EF-2 is a lower-affinity site compared to EF-3 and -4 hands. Comparison of HDX profiles between wild-type DREAM and two EF-1 mutated constructs illustrates that the conformational changes in the EF-1 hand are induced by long-range structural interactions. HDX analyses also reveal a conformational change in an N-terminal leucine-charged residue-rich domain (LCD) remote from Ca 2+ -binding EF-hands. This LCD domain is responsible for the direct interaction between DREAM and cAMP response element-binding protein (CREB) and regulates the recruitment of the co-activator, CREB-binding protein. These long-range interactions strongly suggest how conformational changes transmit the Ca 2+ signal to CREB-mediated gene transcription.

The association between drugs frequently used by the elderly and vitamin D blood levels: a review of observational and experimental studies.

PubMed

van Orten-Luiten, Anne Claire B; Janse, André; Dhonukshe-Rutten, Rosalie A M; Witkamp, Renger F

2014-02-01

The risk of adverse drug reactions (ADRs) rises with increasing age. In the field of ADRs, drug-nutrient interactions (DNIs) are a relatively unexplored area. More knowledge will contribute to the simple prevention of this type of ADR. As the prevalence of vitamin D deficiency in the elderly is high, the primary objective of this review is to evaluate the literature on the relationship between drug use and vitamin D status, focusing on medicines commonly used by the elderly. PubMed was searched for human epidemiological and clinical studies published until early 2013, investigating the relationship between vitamin D blood levels and use of drugs from one of the following groups: proton pump inhibitors (PPIs), biguanides, vitamin K antagonists, platelet aggregation inhibitors, thiazide diuretics, loop diuretics, beta-blocking agents, calcium channel blockers, angiotensin-converting enzyme (ACE) inhibitors, angiotensin-II antagonists, statins, benzodiazepines, and antidepressants. A total of 63 publications were identified. Thiazide diuretics, statins, and calcium channel blocking agents were the most frequently studied drug groups. Associations between thiazides and vitamin D were mixed (n = 22), statins had no or positive associations (n = 16) and calcium blockers were not associated or were negatively associated with vitamin D (n = 10). In conclusion, several knowledge gaps exist on the relationship between drug use and vitamin D blood levels. Available data are scarce (particularly for the aged), study characteristics are highly variable, and found associations may be confounded by, amongst other things, the underlying disease. Nonetheless, this review provides a basis for future research on ADRs that contribute to nutrient deficiencies.

Distribution of L-type calcium channels in rat thalamic neurones.

PubMed

Budde, T; Munsch, T; Pape, H C

1998-02-01

One major pathway for calcium entry into neurones is through voltage-activated calcium channels. The distribution of calcium channels over the membrane surface is important for their contribution to neuronal function. Electrophysiological recordings from thalamic cells in situ and after acute isolation demonstrated the presence of high-voltage activated calcium currents. The use of specific L-type calcium channel agonists and antagonists of the dihydropyridine type revealed an about 40% contribution of L-type channels to the total high-voltage-activated calcium current. In order to localize L-type calcium channels in thalamic neurones, fluorescent dihydropyridines were used. They were combined with the fluorescent dye RH414, which allowed the use of a ratio technique and thereby the determination of channel density. The distribution of L-type channels was analysed in the three main thalamic cell types: thalamocortical relay cells, local interneurones and reticular thalamic neurones. While channel density was highest in the soma and decreased significantly in the dendritic region, channels appeared to be clustered differentially in the three types of cells. In thalamocortical cells, L-type channels were clustered in high density around the base of dendrites, while they were more evenly distributed on the soma of interneurones. Reticular thalamic neurones exhibited high density of L-type channels in more central somatic regions. The differential localization of L-type calcium channels found in this study implies their predominate involvement in the regulation of somatic and proximal dendritic calcium-dependent processes, which may be of importance for specific thalamic functions, such as those mediating the transition from rhythmic burst activity during sleep to single spike activity during wakefulness or regulating the relay of visual information.

Spatial temperature gradients guide axonal outgrowth

PubMed Central

Black, Bryan; Vishwakarma, Vivek; Dhakal, Kamal; Bhattarai, Samik; Pradhan, Prabhakar; Jain, Ankur; Kim, Young-tae; Mohanty, Samarendra

2016-01-01

Formation of neural networks during development and regeneration after injury depends on accuracy of axonal pathfinding, which is primarily believed to be influenced by chemical cues. Recently, there is growing evidence that physical cues can play crucial role in axonal guidance. However, detailed mechanism involved in such guidance cues is lacking. By using weakly-focused near-infrared continuous wave (CW) laser microbeam in the path of an advancing axon, we discovered that the beam acts as a repulsive guidance cue. Here, we report that this highly-effective at-a-distance guidance is the result of a temperature field produced by the near-infrared laser light absorption. Since light absorption by extracellular medium increases when the laser wavelength was red shifted, the threshold laser power for reliable guidance was significantly lower in the near-infrared as compared to the visible spectrum. The spatial temperature gradient caused by the near-infrared laser beam at-a-distance was found to activate temperature-sensitive membrane receptors, resulting in an influx of calcium. The repulsive guidance effect was significantly reduced when extracellular calcium was depleted or in the presence of TRPV1-antagonist. Further, direct heating using micro-heater confirmed that the axonal guidance is caused by shallow temperature-gradient, eliminating the role of any non-photothermal effects. PMID:27460512

Roles of cyclic AMP and Ca in epithelial ion transport across corneal epithelium: a review.

PubMed

Reinach, P S

1985-04-01

The messenger roles of cyclic AMP and the calcium ion in stimulus-secretion coupling are considered in the frog and bovine corneal epithelium, respectively. In the frog cornea, epinephrine stimulates net C1 transport by increasing cyclic AMP content. This stimulation is associated with a larger apical membrane C1 conductance and basolateral membrane ionic conductance. The response of the apical membrane conductance is thought to result from an increase in cyclic AMP content whereas the basolateral membrane ionic conductance increase is unrelated based on measurements of the effects of the calcium channel antagonist, diltiazem, and the beta agonist, isoproterenol, on the electrical parameters and cyclic AMP content. The basolateral membrane is essentially K permselective since the K channel blocker, Ba, depolarized the intracellular potential difference and increased the basolateral membrane resistance. Diltiazem had even larger effects on these parameters suggesting that this compound is a more effective inhibitor of K channel activity than barium. In broken cell preparations of bovine corneal epithelium, a high affinity form of Ca + Mg activated ATPase is present (Km = .06 microM for Ca) and is essentially of plasma membrane origin. This ATPase activation is at a Ca activity similar to the expected intracellular value and suggests that this activity is the enzymatic basis for net Ca transport.

Spatial temperature gradients guide axonal outgrowth

NASA Astrophysics Data System (ADS)

Black, Bryan; Vishwakarma, Vivek; Dhakal, Kamal; Bhattarai, Samik; Pradhan, Prabhakar; Jain, Ankur; Kim, Young-Tae; Mohanty, Samarendra

2016-07-01

Formation of neural networks during development and regeneration after injury depends on accuracy of axonal pathfinding, which is primarily believed to be influenced by chemical cues. Recently, there is growing evidence that physical cues can play crucial role in axonal guidance. However, detailed mechanism involved in such guidance cues is lacking. By using weakly-focused near-infrared continuous wave (CW) laser microbeam in the path of an advancing axon, we discovered that the beam acts as a repulsive guidance cue. Here, we report that this highly-effective at-a-distance guidance is the result of a temperature field produced by the near-infrared laser light absorption. Since light absorption by extracellular medium increases when the laser wavelength was red shifted, the threshold laser power for reliable guidance was significantly lower in the near-infrared as compared to the visible spectrum. The spatial temperature gradient caused by the near-infrared laser beam at-a-distance was found to activate temperature-sensitive membrane receptors, resulting in an influx of calcium. The repulsive guidance effect was significantly reduced when extracellular calcium was depleted or in the presence of TRPV1-antagonist. Further, direct heating using micro-heater confirmed that the axonal guidance is caused by shallow temperature-gradient, eliminating the role of any non-photothermal effects.

Antiapolipoprotein A-1 IgG chronotropic effects require nongenomic action of aldosterone on L-type calcium channels.

PubMed

Rossier, Michel F; Pagano, Sabrina; Python, Magaly; Maturana, Andres D; James, Richard W; Mach, François; Roux-Lombard, Pascale; Vuilleumier, Nicolas

2012-03-01

Autoantibodies to apolipoprotein A-1 (antiapoA-1 IgG) have been shown to be associated with higher resting heart rate and morbidity in myocardial infarction patients and to behave as a chronotropic agent in the presence of aldosterone on isolated neonatal rat ventricular cardiomyocytes (NRVC). We aimed at identifying the pathways accounting for this aldosterone-dependent antiapoA-1 IgG-positive chronotropic effect on NRVC. The rate of regular spontaneous contractions was determined on NRVC in the presence of different steroid hormones and antagonists. AntiapoA-1 IgG chronotropic response was maximal within 20 min and observed only in aldosterone-pretreated cells but not in those exposed to other steroids. The positive antiapoA-1 IgG chronotropic effect was already significant after 5 min aldosterone preincubation, was dependent on 3-kinase and protein kinase A activities, was not inhibited by actinomycin D, and was fully abrogated by eplerenone (but not by spironolactone), demonstrating the dependence on a nongenomic action of aldosterone elicited through the mineralocorticoid receptor (MR). Under oxidative conditions (but not under normal redox state), corticosterone mimicked the permissive action of aldosterone on the antiapoA-1 IgG chronotropic response. Pharmacological and patch-clamp studies identified L-type calcium channels as crucial effectors of antiapoA-1 IgG chronotropic action, involving two converging pathways that increase the channel activity. The first one involves the rapid, nongenomic activation of the phosphatidylinositol 3-kinase enzyme by MR, and the second one requires a constitutive basal protein kinase A activity. In conclusion, our results indicate that, on NRVC, the aldosterone-dependent chronotropic effects of antiapoA-1 IgG involve the nongenomic activation of L-type calcium channels.

Activation of the sigma receptor 1 modulates AMPA receptor-mediated light-evoked excitatory postsynaptic currents in rat retinal ganglion cells.

PubMed

Liu, Lei-Lei; Deng, Qin-Qin; Weng, Shi-Jun; Yang, Xiong-Li; Zhong, Yong-Mei

2016-09-22

Sigma receptor (σR), a unique receptor family, is classified into three subtypes: σR1, σR2 and σR3. It was previously shown that σR1 activation induced by 1μM SKF10047 (SKF) suppressed N-methyl-d-aspartate (NMDA) receptor-mediated responses of rat retinal ganglion cells (GCs) and the suppression was mediated by a distinct Ca(2+)-dependent phospholipase C (PLC)-protein kinase C (PKC) pathway. In the present work, using whole-cell patch-clamp techniques in rat retinal slice preparations, we further demonstrate that SKF of higher dosage (50μM) significantly suppressed AMPA receptor (AMPAR)-mediated light-evoked excitatory postsynaptic currents (L-EPSCs) of retinal ON-type GCs (ON GCs), and the effect was reversed by the σR1 antagonist BD1047, suggesting the involvement of σR1. The SKF (50μM) effect was unlikely due to a change in glutamate release from bipolar cells, as suggested by the unaltered paired-pulse ratio (PPR) of AMPAR-mediated EPSCs of ON GCs. SKF (50μM) did not change L-EPSCs of ON GCs when the G protein inhibitor GDP-β-S or the protein kinase G (PKG) inhibitor KT5823 was intracellularly infused. Calcium imaging further revealed that SKF (50μM) did not change intracellular calcium concentration in GCs and persisted to suppress L-EPSCs when intracellular calcium was chelated by BAPTA. The SKF (50μM) effect was intact when protein kinase A (PKA) and phosphatidylinostiol (PI)-PLC signaling pathways were both blocked. We conclude that the SKF (50μM) effect is Ca(2+)-independent, PKG-dependent, but not involving PKA, PI-PLC pathways. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

PPARγ antagonist attenuates mouse immune-mediated bone marrow failure by inhibition of T cell function

PubMed Central

Sato, Kazuya; Feng, Xingmin; Chen, Jichun; Li, Jungang; Muranski, Pawel; Desierto, Marie J.; Keyvanfar, Keyvan; Malide, Daniela; Kajigaya, Sachiko; Young, Neal S.

2016-01-01

Acquired aplastic anemia is an immune-mediated disease, in which T cells target hematopoietic cells; at presentation, the bone marrow is replaced by fat. It was reported that bone marrow adipocytes were negative regulators of hematopoietic microenvironment. To examine the role of adipocytes in bone marrow failure, we investigated peroxisomal proliferator-activated receptor gamma, a key transcription factor in adipogenesis, utilizing an antagonist of this factor called bisphenol-A-diglycidyl-ether. While bisphenol-A-diglycidyl-ether inhibited adipogenesis as expected, it also suppressed T cell infiltration of bone marrow, reduced plasma inflammatory cytokines, decreased expression of multiple inflammasome genes, and ameliorated marrow failure. In vitro, bisphenol-A-diglycidyl-ether suppressed activation and proliferation, and reduced phospholipase C gamma 1 and nuclear factor of activated T-cells 1 expression, as well as inhibiting calcium flux in T cells. The in vivo effect of bisphenol-A-diglycidyl-ether on T cells was confirmed in a second immune-mediated bone marrow failure model, using different strains and non-major histocompatibility antigen mismatched: bisphenol-A-diglycidyl-ether ameliorated marrow failure by inhibition of T cell infiltration of bone marrow. Our data indicate that peroxisomal proliferator-activated receptor gamma antagonists may attenuate murine immune-mediated bone marrow failure, at least in part, by suppression of T cell activation, which might hold implications in the application of peroxisomal proliferator-activated receptor gamma antagonists in immune-mediated pathophysiologies, both in the laboratory and in the clinic. Genetically “fatless” mice developed bone marrow failure with accumulation of marrow adipocytes in our model, even in the absence of body fat, suggesting different mechanisms of systematic and marrow adipogenesis and physiologic versus pathophysiologic fat accumulation. PMID:26589913

Implication of the ryanodine receptor in TRPV4-induced calcium response in pulmonary arterial smooth muscle cells from normoxic and chronically hypoxic rats.

PubMed

Dahan, Diana; Ducret, Thomas; Quignard, Jean-François; Marthan, Roger; Savineau, Jean-Pierre; Estève, Eric

2012-11-01

There is a growing body of evidence indicating that transient receptor potential (TRP) channels are implicated in calcium signaling and various cellular functions in the pulmonary vasculature. The aim of this study was to investigate the expression, functional role, and coupling to reticulum calcium channels of the type 4 vanilloid TRP subfamily (TRPV4) in the pulmonary artery from both normoxic (Nx) and chronically hypoxic (CH) rats. Activation of TRPV4 with the specific agonist 4α-phorbol-12,13-didecanoate (4α-PDD, 5 μM) increased the intracellular calcium concentration ([Ca(2+)](i)). This effect was significantly reduced by a high concentration of ryanodine (100 μM) or chronic caffeine (5 mM) that blocked ryanodine receptor (RyR) but was insensitive to xestospongin C (10 μM), an inositol trisphosphate receptor antagonist. Inhibition of RyR1 and RyR3 only with 10 μM of dantrolene did not attenuate the 4α-PDD-induced [Ca(2+)](i) increase. Western blotting experiments revealed the expression of TRPV4 and RyR2 with an increase in both receptors in pulmonary arteries from CH rats vs. Nx rats. Accordingly, the 4α-PDD-activated current, measured with patch-clamp technique, was increased in pulmonary artery smooth muscle cells (PASMC) from CH rats vs. Nx rats. 4α-PDD increased isometric tension in artery rings, and this response was also potentiated under chronic hypoxia conditions. 4α-PDD-induced calcium response, current, and contraction were all inhibited by the selective TRPV4 blocker HC-067047. Collectively, our findings provide evidence of the interplay between TRPV4 and RyR2 in the Ca(2+) release mechanism and contraction in PASMC. This study provides new insights onto the complex calcium signaling in PASMC and point out the importance of the TRPV4-RyR2 signaling pathway under hypoxic conditions that may lead to pulmonary hypertension.

P/Q-type and T-type voltage-gated calcium channels are involved in the contraction of mammary and brain blood vessels from hypertensive patients.

PubMed

Thuesen, A D; Lyngsø, K S; Rasmussen, L; Stubbe, J; Skøtt, O; Poulsen, F R; Pedersen, C B; Rasmussen, L M; Hansen, P B L

2017-03-01

Calcium channel blockers are widely used in cardiovascular diseases. Besides L-type channels, T- and P/Q-type calcium channels are involved in the contraction of human renal blood vessels. It was hypothesized that T- and P/Q-type channels are involved in the contraction of human brain and mammary blood vessels. Internal mammary arteries from bypass surgery patients and cerebral arterioles from patients with brain tumours with and without hypertension were tested in a myograph and perfusion set-up. PCR and immunohistochemistry were performed on isolated blood vessels. The P/Q-type antagonist ω-agatoxin IVA (10 -8  mol L -1 ) and the T-type calcium blocker mibefradil (10 -7  mol L -1 ) inhibited KCl depolarization-induced contraction in mammary arteries from hypertensive patients with no effect on blood vessels from normotensive patients. ω-Agatoxin IVA decreased contraction in cerebral arterioles from hypertensive patients. L-type blocker nifedipine abolished the contraction in mammary arteries. PCR analysis showed expression of P/Q-type (Ca v 2.1), T-type (Ca v 3.1 and Ca v 3.2) and L-type (Ca v 1.2) calcium channels in mammary and cerebral arteries. Immunohistochemical labelling of mammary and cerebral arteries revealed the presence of Ca v 2.1 in endothelial and smooth muscle cells. Ca v 3.1 was also detected in mammary arteries. P/Q- and T-type Ca v are present in human internal mammary arteries and in cerebral penetrating arterioles. P/Q- and T-type calcium channels are involved in the contraction of mammary arteries from hypertensive patients but not from normotensive patients. Furthermore, in cerebral arterioles P/Q-type channels importance was restricted to hypertensive patients might lead to that T- and P/Q-type channels could be a new target in hypertensive patients. © 2016 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

Implementation of a Fluorescence-Based Screening Assay Identifies Histamine H3 Receptor Antagonists Clobenpropit and Iodophenpropit as Subunit-Selective N-Methyl-d-Aspartate Receptor Antagonists

PubMed Central

Hansen, Kasper B.; Mullasseril, Praseeda; Dawit, Sara; Kurtkaya, Natalie L.; Yuan, Hongjie; Vance, Katie M.; Orr, Anna G.; Kvist, Trine; Ogden, Kevin K.; Le, Phuong; Vellano, Kimberly M.; Lewis, Iestyn; Kurtkaya, Serdar; Du, Yuhong; Qui, Min; Murphy, T. J.; Snyder, James P.; Bräuner-Osborne, Hans

2010-01-01

N-Methyl-d-aspartate (NMDA) receptors are ligand-gated ion channels that mediate a slow, Ca2+-permeable component of excitatory synaptic transmission in the central nervous system and play a pivotal role in synaptic plasticity, neuronal development, and several neurological diseases. We describe a fluorescence-based assay that measures NMDA receptor-mediated changes in intracellular calcium in a BHK-21 cell line stably expressing NMDA receptor NR2D with NR1 under the control of a tetracycline-inducible promoter (Tet-On). The assay selectively identifies allosteric modulators by using supramaximal concentrations of glutamate and glycine to minimize detection of competitive antagonists. The assay is validated by successfully identifying known noncompetitive, but not competitive NMDA receptor antagonists among 1800 screened compounds from two small focused libraries, including the commercially available library of pharmacologically active compounds. Hits from the primary screen are validated through a secondary screen that used two-electrode voltage-clamp recordings on recombinant NMDA receptors expressed in Xenopus laevis oocytes. This strategy identified several novel modulators of NMDA receptor function, including the histamine H3 receptor antagonists clobenpropit and iodophenpropit, as well as the vanilloid receptor transient receptor potential cation channel, subfamily V, member 1 (TRPV1) antagonist capsazepine. These compounds are noncompetitive antagonists and the histamine H3 receptor ligand showed submicromolar potency at NR1/NR2B NMDA receptors, which raises the possibility that compounds can be developed that act with high potency on both glutamate and histamine receptor systems simultaneously. Furthermore, it is possible that some actions attributed to histamine H3 receptor inhibition in vivo may also involve NMDA receptor antagonism. PMID:20197375

Membrane depolarization inhibits spiral ganglion neurite growth via activation of multiple types of voltage sensitive calcium channels and calpain

PubMed Central

Roehm, Pamela C.; Xu, Ningyong; Woodson, Erika A.; Green, Steven H.; Hansen, Marlan R.

2008-01-01

The effect of membrane electrical activity on spiral ganglion neuron (SGN) neurite growth remains unknown despite its relevance to cochlear implant technology. We demonstrate that membrane depolarization delays the initial formation and inhibits the subsequent extension of cultured SGN neurites. This inhibition depends directly on the level of depolarization with higher levels of depolarization causing retraction of existing neurites. Cultured SGNs express subunits for L-type, N-type, and P/Q type voltage-gated calcium channels (VGCCs) and removal of extracellular Ca2+ or treatment with a combination of L-type, N-type, P/Q-type VGCC antagonists rescues SGN neurite growth under depolarizing conditions. By measuring the fluorescence intensity of SGNs loaded with the fluorogenic calpain substrate t-butoxy carbonyl-Leu-Met-chloromethylaminocoumarin (20 μM), we demonstrate that depolarization activates calpains. Calpeptin (15 μM), a calpain inhibitor, prevents calpain activation by depolarization and rescues neurite growth in depolarized SGNs suggesting that calpain activation contributes to the inhibition of neurite growth by depolarization. PMID:18055215

Diarylureas as allosteric modulators of the cannabinoid CB1 receptor: structure-activity relationship studies on 1-(4-chlorophenyl)-3-{3-[6-(pyrrolidin-1-yl)pyridin-2-yl]phenyl}urea (PSNCBAM-1).

PubMed

German, Nadezhda; Decker, Ann M; Gilmour, Brian P; Gay, Elaine A; Wiley, Jenny L; Thomas, Brian F; Zhang, Yanan

2014-09-25

The recent discovery of allosteric modulators of the CB1 receptor including PSNCBAM-1 (4) has generated significant interest in CB1 receptor allosteric modulation. Here in the first SAR study on 4, we have designed and synthesized a series of analogs focusing on modifications at two positions. Pharmacological evaluation in calcium mobilization and binding assays revealed the importance of alkyl substitution at the 2-aminopyridine moiety and electron deficient aromatic groups at the 4-chlorophenyl position for activity at the CB1 receptor, resulting in several analogs with comparable potency to 4. These compounds increased the specific binding of [(3)H]CP55,940, in agreement with previous reports. Importantly, 4 and two analogs dose-dependently reduced the Emax of the agonist curve in the CB1 calcium mobilization assays, confirming their negative allosteric modulator characteristics. Given the side effects associated with CB1 receptor orthosteric antagonists, negative allosteric modulators provide an alternative approach to modulate the pharmacologically important CB1 receptor.

Antioxidant, Anti-Nephrolithe Activities and in Vitro Digestibility Studies of Three Different Cyanobacterial Pigment Extracts

PubMed Central

Paliwal, Chetan; Ghosh, Tonmoy; Bhayani, Khushbu; Maurya, Rahulkumar; Mishra, Sandhya

2015-01-01

Phycobiliprotein-containing water and carotenoid-containing methanolic extracts of three different cyanobacteria, Pseudanabaena sp., Spirulina sp. and Lyngbya sp., were studied for their DPPH scavenging, iso-bolographic studies, and anti-nephrolithe activities. The best EC50 values for DPPH scavenging were in Lyngbya water (LW, 18.78 ± 1.57 mg·mg−1 DPPH) and Lyngbya methanol (LM, 59.56 ± 37.38 mg·mg−1 DPPH) extracts. Iso-bolographic analysis revealed most of the combinations of extracts were antagonistic to each other, although LM—Spirulina methanol (SM) 1:1 had the highest synergistic rate of 86.65%. In vitro digestion studies showed that DPPH scavenging activity was considerably decreased in all extracts except for Pseudanabaena methanol (PM) and LM after the simulated digestion. All of the extracts were effective in reducing the calcium oxalate crystal size by nearly 60%–65% compared to negative control, while PM and Spirulina water (SW) extracts could inhibit both nucleation and aggregation of calcium oxalate by nearly 60%–80%. PMID:26308007

Tryptase potentiates enteric nerve activation by histamine and serotonin: Relevance for the effects of mucosal biopsy supernatants from irritable bowel syndrome patients.

PubMed

Ostertag, D; Annahazi, A; Krueger, D; Michel, K; Demir, I E; Ceyhan, G O; Zeller, F; Schemann, M

2017-09-01

We previously showed that mucosal biopsy supernatants from irritable bowel syndrome patients activated neurons despite low concentrations of tryptase, histamine, and serotonin which individually would not cause spike discharge. We studied the potentiating responses between these mediators on excitability of enteric neurons. Calcium-imaging was performed using the calcium-sensitive dye Fluo-4 AM in human submucous plexus preparations from 45 individuals. Histamine, serotonin, and tryptase were applied alone and in combinations to evaluate nerve activation which was assessed by analyzing increase in intracellular Ca 2+ ([Ca 2+ ] i ), the proportion of responding neurons and the product of both defined as Ca-neuroindex (NI). Protease activated receptor (PAR) 2 activating peptide, PAR2 antagonist and the serine protease-inhibitor FUT-175 were used to particularly investigate the role of proteases. Histamine or serotonin (1 μmol/L each) evoked only few small responses (median NI [25%/75%]: 0 [0/148]; 85 [0/705] respectively). Their combined application evoked statistically similar responses (216 [21/651]). Addition of the PAR2 activator tryptase induced a significantly higher Ca-NI (1401 [867/4075]) compared to individual application of tryptase or to coapplied histamine and serotonin. This synergistic potentiation was neither mimicked by PAR2 activating peptide nor reversed by the PAR2 antagonist GB83, but abolished by FUT-175. We observed synergistic potentiation between histamine, serotonin, and tryptase in enteric neurons, which is mediated by proteolytic activity rather than PAR2 activation. This explained neuronal activation by a cocktail of these mediators despite their low concentrations and despite a relatively small PAR2-mediated response in human submucous neurons. © 2017 John Wiley & Sons Ltd.

Exploration of the Ca2+ interaction modes of the nifedipine calcium channel antagonist.

PubMed

Liu, Huichun; Zhang, Liang; Li, Ping; Cukier, Robert I; Bu, Yuxiang

2007-02-02

A comprehensive study is carried out using quantum chemical computation and molecular dynamics (MD) simulations to gain insight into the interaction between Ca(2+) ions and the most important class of calcium channel antagonists--nifedipine. First, the chelating structures and energetic characters of nifedipine-Ca(2+) in the gas phase are explored, and 25 isomers are found. The most favorable chelating mode is a tridentate one, that is, Ca(2+) binds to two carbonyl O atoms and one nitryl O atom, where Ca(2+) is above the plane of the three O atoms to form a pyramidal structure. Accurate geometric structures, relative stabilities, vertical and adiabatic binding energies, and charge distributions are discussed. The differences in the geometries and energies among these isomers are analyzed from the contributions of chelating sites, electrostatics and polarizations, steric repulsions, and charge distributions. The interconversions among isomers with similar geometries and energies are also investigated because of the importance of the geometric transformation in the biological system. Furthermore, certain numbers of water molecules are added to the nifedipine-Ca(2+) system to probe the effect of water. A detailed study is performed on the hydrated geometries on the basis of the most stable isomer 1. Stepwise hydration can weaken the nifedipine-Ca(2+) interaction, and the chelating sites of nifedipine are gradually replaced by the added water molecules. Hexacoordination is found to be the most favorable geometry no matter how many water molecules were added, which can be verified by the MD simulations. The transfer of water molecules from the inner shell to the outer shell is also supported by MD simulations of the hexahydrated complexes.

Spontaneous calcium transients in human neural progenitor cells mediated by transient receptor potential channels.

PubMed

Morgan, Peter J; Hübner, Rayk; Rolfs, Arndt; Frech, Moritz J

2013-09-15

Calcium signals affect many developmental processes, including proliferation, migration, survival, and apoptosis, processes that are of particular importance in stem cells intended for cell replacement therapies. The mechanisms underlying Ca(2+) signals, therefore, have a role in determining how stem cells respond to their environment, and how these responses might be controlled in vitro. In this study, we examined the spontaneous Ca(2+) activity in human neural progenitor cells during proliferation and differentiation. Pharmacological characterization indicates that in proliferating cells, most activity is the result of transient receptor potential (TRP) channels that are sensitive to Gd(3+) and La(3+), with the more subtype selective antagonist Ruthenium red also reducing activity, suggesting the involvement of transient receptor potential vanilloid (TRPV) channels. In differentiating cells, Gd(3+) and La(3+)-sensitive TRP channels also appear to underlie the spontaneous activity; however, no sub-type-specific antagonists had any effect. Protein levels of TRPV2 and TRPV3 decreased in differentiated cells, which is demonstrated by western blot. Thus, it appears that TRP channels represent the main route of Ca(2+) entry in human neural progenitor cells (hNPCs), but the responsible channel types are subject to substitution under differentiating conditions. The level of spontaneous activity could be increased and decreased by lowering and raising the extracellular K(+) concentration. Proliferating cells in low K(+) slowed the cell cycle, with a disproportionate increased percentage of cells in G1 phase and a reduction in S phase. Taken together, these results suggest a link between external K(+) concentration, spontaneous Ca(2+) transients, and cell cycle distribution, which is able to influence the fate of stem and progenitor cells.

Calcium channel subtypes differ at two types of cholinergic synapse in lumbar sympathetic neurones of guinea-pigs.

PubMed

Ireland, D R; Davies, P J; McLachlan, E M

1999-01-01

1. The involvement of different presynaptic Ca2+ channels in transmission at 'weak' (subthreshold) and 'strong' (suprathreshold) synapses was investigated in guinea-pig paravertebral ganglia isolated in vitro. Selective Ca2+ channel antagonists were used to block excitatory synaptic currents evoked by stimulating single preganglionic axons. 2. The N-type Ca2+ channel blocker, omega-conotoxin GVIA (100 nM), reduced peak synaptic conductance by similar amounts at weak synapses (by 39 +/- 6 %) and strong synapses (34 +/- 6 %). 3. The P-type Ca2+ channel blocker, omega-agatoxin IVA (40 nM), significantly reduced transmitter release at weak synapses (by 42 +/- 6 %) but had only a small effect at strong synapses (reduced by 6 +/- 2 %). 4. Blockers of Q-, L- or T-type Ca2+ channels had no significant effects on peak synaptic conductance at either type of synapse. 5. We conclude that the two functionally distinct types of preganglionic terminal in sympathetic ganglia which synapse on the same neurone differ in their expression of particular types of voltage-dependent Ca2+ channels. Both types utilize N-type channels and channels resistant to blockade by specific antagonists, but Ca2+ entry through P-type channels makes a substantial contribution to acetylcholine release only at weak synapses.

Calcium channel subtypes differ at two types of cholinergic synapse in lumbar sympathetic neurones of guinea-pigs

PubMed Central

Ireland, David R; Davies, Philip J; McLachlan, Elspeth M

1999-01-01

The involvement of different presynaptic Ca2+ channels in transmission at ‘weak’ (subthreshold) and ‘strong’ (suprathreshold) synapses was investigated in guinea-pig paravertebral ganglia isolated in vitro. Selective Ca2+ channel antagonists were used to block excitatory synaptic currents evoked by stimulating single preganglionic axons.The N-type Ca2+ channel blocker, ω-conotoxin GVIA (100 nm), reduced peak synaptic conductance by similar amounts at weak synapses (by 39 ± 6%) and strong synapses (34 ± 6%).The P-type Ca2+ channel blocker, ω-agatoxin IVA (40 nm), significantly reduced transmitter release at weak synapses (by 42 ± 6%) but had only a small effect at strong synapses (reduced by 6 ± 2%).Blockers of Q-, L- or T-type Ca2+ channels had no significant effects on peak synaptic conductance at either type of synapse.We conclude that the two functionally distinct types of preganglionic terminal in sympathetic ganglia which synapse on the same neurone differ in their expression of particular types of voltage-dependent Ca2+ channels. Both types utilize N-type channels and channels resistant to blockade by specific antagonists, but Ca2+ entry through P-type channels makes a substantial contribution to acetylcholine release only at weak synapses. PMID:9831716

alpha-Adrenergic-mediated activation of human reconstituted fibrinogen receptor (integrin alphaIIbbeta3) in Chinese hamster ovary cells.

PubMed

Butta, Nora; Larrucea, Susana; Gonzalez-Manchon, Consuelo; Alonso, Sonia; Parrilla, Roberto

2004-12-01

This work reports the functional studies of CHO cells coexpressing alpha-adrenergic (alphaAR) and human fibrinogen (Fg) receptors (integrin alphaIIbbeta3). Stimulation of these cells with alpha-agonists produced a transient rise in the free cytosolic calcium (Ca(++)) accompanied by enhanced binding to soluble Fg, and these effects were prevented by specific alphaAR antagonists. The alpha-adrenergic-induced activation of alphaIIbbeta3 in CHO-alphaIIbbeta3-alphaAR increased the rate of adhesion and extension of cells onto Fg coated plates, and also induced a soluble Fg- and alphaIIbbeta3-dependent formation of cell aggregates, whereas no effects were observed by the stimulation of CHO-alphaIIbbeta3 cells. alpha-Adrenergic antagonists, the ligand mimetic peptide RGDS, pertussis toxin (PTX), or EDTA, they all prevented the alpha-adrenergic stimulation of adhesion and aggregation. However, inhibition of PKC prevented the alpha-adrenergic stimulation of cell adherence, whereas blocking the intracellular Ca(++) mobilization impeded the stimulation of cell aggregation. The alpha-adrenergic activation was associated with phosphorylation of a protein of approximately 100 kDa and proteins of the MAPK family. The former was selectively phosphorylated by alpha-adrenergic stimulation whereas the latter were phosphorylated by the binding of cells to Fg and markedly intensified by alpha-adrenergic stimulation.

Impact of ranolazine on clinical outcomes and healthcare resource utilization in patients with refractory angina pectoris.

PubMed

Ling, Hua; Packard, Kathleen A; Burns, Tammy L; Hilleman, Daniel E

2013-12-01

Ranolazine is a novel antianginal medication approved for the treatment of chronic angina. There are only limited data concerning the efficacy of ranolazine in reducing healthcare resource utilization in patients with refractory angina pectoris. The primary objective of this analysis was to evaluate the efficacy and safety of ranolazine in refractory angina pectoris. In addition, the impact of ranolazine on healthcare resource utilization was assessed. Consecutive patients with refractory angina pectoris treated with ranolazine at two cardiology practices in the state of Nebraska were included in this analysis. The Canadian Cardiovascular Society (CCS) angina class and frequency and type of healthcare resource consumption were determined during the 12 months prior to and the 12 months after initiation of ranolazine. A total of 150 pts (64 % men) with a mean age of 66 ± 12 years were included in this analysis. All patients had previously undergone coronary revascularization. Nitrates, β-adrenoceptor antagonists (β-blockers), and calcium antagonists (calcium channel blockers) were being used in 83, 97, and 75 % of patients, respectively. During ranolazine treatment, a significant improvement in CCS angina class was observed, with 23 patients improving by one class and no patient experiencing a deterioration in functional class (p = 0.025). A total of 53 side effects occurred in 28 (19 %) patients receiving ranolazine. Of those patients with side effects, four required dose reduction and seven required drug discontinuation. The frequency of clinic visits and emergency room visits was lower during ranolazine treatment, but the differences in frequency were not significant. The number of patients hospitalized and the number of hospitalizations were significantly lower during ranolazine therapy than in the pre-ranolazine study period (p = 0.002). Ranolazine improved the CCS angina class and reduced hospitalizations over a 12-month follow-up period in a group of patients with difficult-to-treat refractory angina pectoris.

Comparison of mibefradil and derivative NNC 55-0396 effects on behavior, cytochrome P450 activity, and tremor in mouse models of essential tremor

PubMed Central

Quesada, Arnulfo; Bui, Peter H.; Homanics, Gregg E.; Hankinson, Oliver; Handforth, Adrian

2014-01-01

NNC 55-0396 [(1S,2S)-2-(2-(N-[(3-benzimidazol-2-yl)propyl]-N-methylamino)ethyl)-6-fluoro-1,2, 3,4-tetrahydro-1-isopropyl-2-naphtyl cyclopropanecarboxylate dihydrochloride], is a mibefradil derivative that retains potent in vitro T-type calcium channel antagonist efficacy. We compared the two compounds for behavioral toxicity, effects on cytochrome P450 activity, and efficacy against tremor in the γ-aminobutyric acid type A (GABAA) receptor subunit α1-null mouse, and the harmaline tremor model of essential tremor in wild-type mice. NNC 55-0396 was better tolerated than mibefradil in the horizontal wire test of sedation/motor function, with 3/6 failing at 300 and 30 mg/kg respectively. To assess for a potential interaction with harmaline, mice were given the drugs, followed by harmaline or vehicle, and tested 30 min later in the inverted wire grid test. Mibefradil exacerbated, whereas NNC 55-0396 ameliorated harmaline-induced test deficits. In mouse liver microsomes, NNC 55-0396 was a less potent inhibitor of harmaline O-demethylation than mibefradil (Ki: 0.95 and 0.29 µM respectively), and also less potent at inhibiting testosterone 6-β-hydroxylation (Ki: 0.71 and 0.12 µM respectively). In the GABAA α1-null model, NNC 55-0396 but not mibefradil, (each at 20 mg/kg), suppressed tremor while NNC 55-0396 at 12.5 mg/kg suppressed harmaline-induced tremor by half by 20–100 min, whereas mibefradil at the same dose did not significantly affect tremor. In contrast to mibefradil, NNC 55-0396 is well tolerated and suppresses tremor, and exerts less cytochrome P450 inhibition. These results suggest potential clinical utility for NNC 55-0396 or similar derivatives as a T-type calcium antagonist. PMID:21256842

Electrophysiological evidence for voltage-gated calcium channel 2 (Cav2) modulation of mechano- and thermosensitive spinal neuronal responses in a rat model of osteoarthritis.

PubMed

Rahman, W; Patel, R; Dickenson, A H

2015-10-01

Osteoarthritis (OA) remains one of the greatest healthcare burdens in western society, with chronic debilitating pain-dominating clinical presentation yet therapeutic strategies are inadequate in many patients. Development of better analgesics is contingent on improved understanding of the molecular mechanisms mediating OA pain. Voltage-gated calcium channels 2.2 (Cav2.2) play a critical role in spinal nociceptive transmission, therefore blocking Cav2.2 activity represents an attractive opportunity for OA pain treatment, but the only available licensed Cav2.2 antagonist ziconitide (PrilatTM) is of limited use. TROX-1 is an orally available, use dependent and state-selective Cav2 antagonist, exerting its analgesic effect primarily via Cav2.2 blockade, with an improved therapeutic window compared with ziconitide. Using a rat model of monosodium iodoacetate (MIA), 2 mg, induced OA we used in vivo electrophysiology to assess the effects of spinal or systemic administration of TROX-1 on the evoked activity of wide dynamic range spinal dorsal horn neurons in response to electrical, natural mechanical (dynamic brush and von Frey 2, 8, 26 and 6 g) and thermal (40, 45 and 45 °C) stimuli applied to the peripheral receptive field. MIA injection into the knee joint resulted in mechanical hypersensitivity of the ipsilateral hind paw and weight-bearing asymmetry. Spinal administration of TROX-1 (0.1 and 1 μg/50 μl) produced a significant dose-related inhibition of dynamic brush, mechanical (von Frey filament (vF) 8, 26 and 60 g) and noxious thermal-(45 and 48 °C) evoked neuronal responses in MIA rats only. Systemic administration of TROX-1 produced a significant inhibition of the mechanical-(vF 8, 26 and 60 g) evoked neuronal responses in MIA rats. TROX-1 did not produce any significant effect on any neuronal measure in Sham controls. Our in vivo electrophysiological results demonstrate a pathological state-dependent effect of TROX-1, which suggests an increased functional role of Cav2, likely Cav2.2, channels in mediating OA pain. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Electrophysiological evidence for voltage-gated calcium channel 2 (Cav2) modulation of mechano- and thermosensitive spinal neuronal responses in a rat model of osteoarthritis

PubMed Central

Rahman, W.; Patel, R.; Dickenson, A.H.

2015-01-01

Osteoarthritis (OA) remains one of the greatest healthcare burdens in western society, with chronic debilitating pain-dominating clinical presentation yet therapeutic strategies are inadequate in many patients. Development of better analgesics is contingent on improved understanding of the molecular mechanisms mediating OA pain. Voltage-gated calcium channels 2.2 (Cav2.2) play a critical role in spinal nociceptive transmission, therefore blocking Cav2.2 activity represents an attractive opportunity for OA pain treatment, but the only available licensed Cav2.2 antagonist ziconitide (PrilatTM) is of limited use. TROX-1 is an orally available, use dependent and state-selective Cav2 antagonist, exerting its analgesic effect primarily via Cav2.2 blockade, with an improved therapeutic window compared with ziconitide. Using a rat model of monosodium iodoacetate (MIA), 2 mg, induced OA we used in vivo electrophysiology to assess the effects of spinal or systemic administration of TROX-1 on the evoked activity of wide dynamic range spinal dorsal horn neurons in response to electrical, natural mechanical (dynamic brush and von Frey 2, 8, 26 and 6 g) and thermal (40, 45 and 45 °C) stimuli applied to the peripheral receptive field. MIA injection into the knee joint resulted in mechanical hypersensitivity of the ipsilateral hind paw and weight-bearing asymmetry. Spinal administration of TROX-1 (0.1 and 1 μg/50 μl) produced a significant dose-related inhibition of dynamic brush, mechanical (von Frey filament (vF) 8, 26 and 60 g) and noxious thermal-(45 and 48 °C) evoked neuronal responses in MIA rats only. Systemic administration of TROX-1 produced a significant inhibition of the mechanical-(vF 8, 26 and 60 g) evoked neuronal responses in MIA rats. TROX-1 did not produce any significant effect on any neuronal measure in Sham controls. Our in vivo electrophysiological results demonstrate a pathological state-dependent effect of TROX-1, which suggests an increased functional role of Cav2, likely Cav2.2, channels in mediating OA pain. PMID:26247695

Role of calmodulin and calcineurin in regulating flagellar motility and wave polarity in Leishmania.

PubMed

Mukhopadhyay, Aakash Gautam; Dey, Chinmoy Sankar

2017-11-01

We have previously reported the involvement of cyclic AMP in regulating flagellar waveforms in Leishmania. Here, we investigated the roles of calcium, calmodulin, and calcineurin in flagellar motility regulation in L. donovani. Using high-speed videomicroscopy, we show that calcium-independent calmodulin and calcineurin activity is necessary for motility in Leishmania. Inhibition of calmodulin and calcineurin induced ciliary beats interrupting flagellar beating in both live (in vivo) and ATP-reactivated (in vitro) parasites. Our results indicate that signaling mediated by calmodulin and calcineurin operates antagonistically to cAMP signaling in regulating the waveforms of Leishmania flagellum. These two pathways are possibly involved in maintaining the balance between the two waveforms, essential for responding to environmental cues, survival, and infectivity.

Adrenergic signaling mediates mechanical hyperalgesia through activation of P2X3 receptors in primary sensory neurons of rats with chronic pancreatitis.

PubMed

Wang, Shusheng; Zhu, Hong-Yan; Jin, Yi; Zhou, Youlang; Hu, Shufen; Liu, Tong; Jiang, Xinghong; Xu, Guang-Yin

2015-04-15

The mechanism of pain in chronic pancreatitis (CP) is poorly understood. The aim of this study was designed to investigate roles of norepinephrine (NE) and P2X receptor (P2XR) signaling pathway in the pathogenesis of hyperalgesia in a rat model of CP. CP was induced in male adult rats by intraductal injection of trinitrobenzene sulfonic acid (TNBS). Mechanical hyperalgesia was assessed by referred somatic behaviors to mechanical stimulation of rat abdomen. P2XR-mediated responses of pancreatic dorsal root ganglion (DRG) neurons were measured utilizing calcium imaging and whole cell patch-clamp-recording techniques. Western blot analysis and immunofluorescence were performed to examine protein expression. TNBS injection produced a significant upregulation of P2X3R expression and an increase in ATP-evoked responses of pancreatic DRG neurons. The sensitization of P2X3Rs was reversed by administration of β-adrenergic receptor antagonist propranolol. Incubation of DRG neurons with NE significantly enhanced ATP-induced intracellular calcium signals, which were abolished by propranolol, and partially blocked by protein kinase A inhibitor H-89. Interestingly, TNBS injection led to a significant elevation of NE concentration in DRGs and the pancreas, an upregulation of β2-adrenergic receptor expression in DRGs, and amplification of the NE-induced potentiation of ATP responses. Importantly, pancreatic hyperalgesia was markedly attenuated by administration of purinergic receptor antagonist suramin or A317491 or β2-adrenergic receptor antagonist butoxamine. Sensitization of P2X3Rs, which was likely mediated by adrenergic signaling in primary sensory neurons, contributes to pancreatic pain, thus identifying a potential target for treating pancreatic pain caused by inflammation. Copyright © 2015 the American Physiological Society.

Slow synaptic transmission mediated by TRPV1 channels in CA3 interneurons of the hippocampus.

PubMed

Eguchi, Noriomi; Hishimoto, Akitoyo; Sora, Ichiro; Mori, Masahiro

2016-03-11

Metabotropic glutamate receptors (mGluRs) modulate various neuronal functions in the central nervous system. Many studies reported that mGluRs have linkages to neuronal disorders such as schizophrenia and autism related disorders, indicating that mGluRs are involved in critical functions of the neuronal circuits. To study this possibility further, we recorded mGluR-induced synaptic responses in the interneurons of the CA3 stratum radiatum using rat hippocampal organotypic slice cultures. Electrical stimulation in the CA3 pyramidal cell layer evoked a slow inward current in the interneurons at a holding potential of -70mV in the presence of antagonists for AMPA/kainate receptors, NMDA receptors, GABAA receptors and GABAB receptors. The slow inward current was blocked in the absence of extracellular calcium, suggesting that this was a synaptic response. The slow excitatory postsynaptic current (EPSC) reversed near 0mV, reflecting an increase in a non-selective cationic conductance. The slow EPSC is mediated by group I mGluRs, as it was blocked by AP3, a group I mGluR antagonist. Neither a calcium chelator BAPTA nor a phospholipase C (PLC) inhibitor U73122 affected the slow EPSC. La(3+), a general TRP channel blocker or capsazepine, a selective TRPV1 channel antagonist significantly suppressed the slow EPSC. DHPG, a selective group I mGluRs agonist induced an inward current, which was suppressed by capsazepine. These results indicate that in the interneurons of the hippocampal CA3 stratum radiatum group I mGluRs activate TRPV1 channels independently of PLC and intracellular Ca(2+), resulting in the slow EPSC in the interneurons. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Zinc release in the lateral nucleus of the amygdala by stimulation of the entorhinal cortex.

PubMed

Takeda, Atsushi; Imano, Sachie; Itoh, Hiromasa; Oku, Naoto

2006-11-06

Zinc release in the lateral nucleus of the amygdala was examined using rat brain slices. The lateral and basolateral nuclei in the amygdala were evidently stained by Timm's sulfide-silver staining method. When the amygdala including both the nuclei was stimulated with 100 mM KCl by means of in vivo microdialysis, extracellular zinc concentration was increased significantly. Zinc release in the lateral nucleus of the amygdala innervated by the entorhinal cortex was next examined in brain slices double-stained with zinc and calcium indicators. Extracellular zinc signal (ZnAF-2) in the lateral nucleus was increased with intracellular calcium signal (calcium orange) during delivery of tetanic stimuli to the entorhinal cortex. Both the increases were completely inhibited by addition of 1 micro M tetrodotoxin, a sodium channel blocker. Furthermore, calcium signal in the lateral nucleus during delivery of tetanic stimuli to the entorhinal cortex was increased in the presence of 10 micro M CNQX, an AMPA/KA receptor antagonist, and this increase was facilitated by addition of 1 mM CaEDTA, a membrane-impermeable zinc chelator. The present study suggested that zinc is released in the lateral nucleus of the amygdala by depolarization of the entorhinal neurons. In the lateral nucleus, zinc released may suppress the increase in presynaptic calcium signal.

Influence of Prostanoids in the Diuretic and Natriuretic Effects of Extracts and Kaempferitrin from Bauhinia forficata Link Leaves in Rats.

PubMed

de Souza, Priscila; da Silva, Luisa Mota; Boeing, Thaise; Somensi, Lincon Bordignon; Cechinel-Zanchett, Camile Cecconi; Campos, Adriana; Krueger, Clarissa de Medeiros Amorim; Bastos, Jairo Kenupp; Cechinel-Filho, Valdir; Andrade, Sérgio Faloni de

2017-10-01

Although Bauhinia forficata Link is popularly used in Brazil to induce diuresis, no scientific investigation has focused on demonstrating its efficacy in preclinical trials. For that, normotensive male Wistar and spontaneously hypertensive rats were used to test the effect of extracts and kaempferitrin obtained from Bauhinia forficata leaves in the experimental model of diuresis. Cumulative urine volume, Na + and K + excretion, calcium, creatinine, prostaglandin E 2 , pH, density, and conductivity were measured at the end of the experiment (after 8 or 24 h). The treatment with aqueous infusion, methanolic extract, trichloromethane, or ethyl acetate-butanolic fractions significantly increase urinary volume and electrolytes levels when orally given to rats, without altering the pH or density parameters. Kaempferitrin induced diuretic, natriuretic, but not kaliuretic effects in both normotensive and hypertensive rats. In addition, kaempferitrin enhanced urinary creatinine and prostaglandin E 2 excretion, without modifying calcium levels. Kaempferitrin-induced diuresis was unaffected by previous treatment with a nonselective inhibitor of nitric oxide synthase and neither with a nonselective muscarinic receptor antagonist. On the other hand, a cyclooxygenase inhibitor was able to decrease its effect when compared with vehicle-treated rats, suggesting that the diuretic and natriuretic properties from kaempferitrin are associated with endogenous prostanoids generation. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Potential New Agents for the Management of Hyperkalemia.

PubMed

Packham, David K; Kosiborod, Mikhail

2016-02-01

Hyperkalemia is a common electrolyte disturbance with multiple potential etiologies. It is usually observed in the setting of reduced renal function. Mild to moderate hyperkalemia is usually asymptomatic, but is associated with poor prognosis. When severe, hyperkalemia may cause serious acute cardiac arrhythmias and conduction abnormalities, and may result in sudden death. The rising prevalence of conditions associated with hyperkalemia (heart failure, chronic kidney disease, and diabetes) and broad use of renin-angiotensin-aldosterone system (RAAS) inhibitors and mineralocorticoid receptor antagonists (MRAs), which improve patient outcomes but increase the risk of hyperkalemia, have led to a significant rise in hyperkalemia-related hospitalizations and deaths. Current non-invasive therapies for hyperkalemia either do not remove excess potassium or have poor efficacy and tolerability. There is a clear need for safer, more effective potassium-lowering therapies suitable for both acute and chronic settings. Patiromer sorbitex calcium and sodium zirconium cyclosilicate (ZS-9) are two new potassium-lowering compounds currently in development. Although they have not yet been approved by the US FDA, both have demonstrated efficacy and safety in recent trials. Patiromer sorbitex calcium is a polymer resin and sorbitol complex that binds potassium in exchange for calcium; ZS-9, a non-absorbed, highly selective inorganic cation exchanger, traps potassium in exchange for sodium and hydrogen. This review discusses the merits of both novel drugs and how they may help optimize the future management of patients with hyperkalemia.

Zinc Modulates Nanosilver-Induced Toxicity in Primary Neuronal Cultures.

PubMed

Ziemińska, Elżbieta; Strużyńska, Lidia

2016-02-01

Silver nanoparticles (NAg) have recently become one of the most commonly used nanomaterials. Since the ability of nanosilver to enter the brain has been confirmed, there has been a need to investigate mechanisms of its neurotoxicity. We previously showed that primary neuronal cultures treated with nanosilver undergo destabilization of calcium homeostasis via a mechanism involving glutamatergic NMDA receptors. Considering the fact that zinc interacts with these receptors, the aim of the present study was to examine the role of zinc in mechanisms of neuronal cell death in primary cultures. In cells treated with nanosilver, we noted an imbalance between extracellular and intracellular zinc levels. Thus, the influence of zinc deficiency and supplementation on nanosilver-evoked cytotoxicity was investigated by treatment with TPEN (a chelator of zinc ions), or ZnCl(2), respectively. Elimination of zinc leads to complete death of nanosilver-treated CGCs. In contrast, supplementation with ZnCl(2) increases viability of CGCs in a dose-dependent manner. Addition of zinc provided protection against the extra/intracellular calcium imbalance in a manner similar to MK-801, an antagonist of NMDA receptors. Zinc chelation by TPEN decreases the mitochondrial potential and dramatically increases the rate of production of reactive oxygen species. Our results indicate that zinc supplementation positively influences nanosilver-evoked changes in CGCs. This is presumed to be due to an inhibitory effect on NMDA-sensitive calcium channels.

Neurobiology of anxiety disorders and implications for treatment.

PubMed

Garakani, Amir; Mathew, Sanjay J; Charney, Dennis S

2006-11-01

The neurobiology of the anxiety disorders, which include panic disorder, post-traumatic stress disorder (PTSD), and specific phobias, among others, has been clarified by advances in the field of classical or Pavlovian conditioning, and in our understanding of basic mechanisms of memory and learning. Fear conditioning occurs when a neutral conditioned stimulus (such as a tone) is paired with an aversive, or unconditioned stimulus (such as a footshock), and then in the absence of the unconditioned stimulus, causes a conditioned fear response. Preclinical studies have shown that the amygdala plays a key role in fear circuitry, and that abnormalities in amygdala pathways can affect the acquisition and expression of fear conditioning. Drugs such as glutamate N-methyl-D-aspartate (NMDA) antagonists, and blockers of voltage-gated calcium channels, in the amygdala, may block these effects. There is also preliminary evidence for the use of centrally acting beta-adrenergic antagonists, like propranolol, to inhibit consolidation of traumatic memories in PTSD. Finally, fear extinction, which entails new learning of fear inhibition, is central to the mechanism of effective anti-anxiety treatments. Several pharmacological manipulations, such as D-cycloserine, a partial NMDA agonist, have been found to facilitate extinction. Combining these medication approaches with psychotherapies that promote extinction, such as cognitive behavioral therapy (CBT), may offer patients with anxiety disorders a rapid and robust treatment with good durability of effect.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kikuno, Shota; Taguchi, Keiko; Iwamoto, Noriko

1,2-Naphthoquinone (1,2-NQ) has recently been identified as an environmental quinone in diesel exhaust particles (DEP) and atmospheric PM{sub 2.5}. We have found that this quinone is capable of causing a concentration-dependent contraction of tracheal smooth muscle in guinea pigs with EC{sub 5} value of 18.7 {mu}M. The contraction required extracellular calcium and was suppressed by L-type calcium channel blockers nifedipine and diltiazem. It was found that 1,2-NQ activated phospholipase A2 (PLA2)/lipoxygenase (LO)/vanilloid receptor (VR1) signaling. Additionally, 1,2-NQ was capable of transactivating protein tyrosine kinases (PTKs) such as epidermal growth factor receptor (EGFR) in guinea pig trachea, suggesting that phosphorylation ofmore » PTKs contributes to 1,2-NQ-induced tracheal contraction. Consistent with this notion, this action was blocked by the PTKs inhibitor genistein and the EGFR antagonist PD153035, indicating that contraction was, at least in part, attributable to PTKs phosphorylation that activates VR1, resulting in increased intracellular calcium content in the smooth muscle cells.« less

Study of the mechanism of the relaxant action of (+)-glaucine in rat vas deferens.

PubMed Central

Orallo, F.; Fernández Alzueta, A.; Loza, M. I.; Vivas, N.; Badía, A.; Campos, M.; Honrubia, M. A.; Cadavid, M. I.

1993-01-01

1. Effects of the aporphinoid alkaloid, (+)-glaucine, on rat vas deferens were investigated. 2. (+)-Glaucine (2-18 microM) competitively inhibited contractions induced by noradrenaline and methoxamine with a pA2 value of about 6. 3. (+)-Glaucine (2 and 18 microM) did not change the accumulation of tritium during incubation of the vas deferens with [3H]-noradrenaline. 4. (+)-Glaucine (0.3 nM-0.1 mM) inhibited specific [3H]-prazosin binding to membranes from rat vas deferens with a pKi value of 6.63, which is close to the pA2 value obtained against noradrenaline and methoxamine in functional studies. 5. In electrically-stimulated rat vas deferens, (+)-glaucine (0.3-10 microM) enhanced twitch contractions and competitively antagonized the inhibitory effect of clonidine with a pA2 value of 5.91. 6. In tissues incubated in depolarizing calcium-free high-potassium medium, (+)-glaucine (30-80 microM) inhibited Ca(2+)-induced contractions with depression of the maximal response at higher doses and with a pD'2 value of 3.65. Furthermore, (+)-glaucine (50 microM) did not modify basal 45Ca uptake but strongly inhibited the influx of 45Ca induced by K+. 7. These results suggest that (+)-glaucine has non-selective alpha 1- and alpha 2-adrenoceptor blocking properties. At higher doses, (+)-glaucine shows calcium antagonist activity which may be responsible, at least in part, for the inhibition of the contractions induced by Ca2+ in calcium-free high-potassium medium. PMID:8298818

N-methyl-D-aspartate receptor antagonist MK-801 prevents apoptosis in rats that have undergone fetal spinal cord transplantation following spinal hemisection.

PubMed

Zhang, Qiang; Shao, Yang; Zhao, Changsong; Cai, Juan; Sun, Sheng

2014-12-01

Spinal cord injury is the main cause of paraplegia, but effective therapies for it are lacking. Embryonic spinal cord transplantation is able to repair spinal cord injury, albeit with a large amount of neuronal apoptosis remaining in the spinal cord. MK-801, an N-methyl-D-aspartate (NMDA) receptor antagonist, is able to reduce cell death by decreasing the concentration of excitatory amino acids and preventing extracellular calcium ion influx. In this study, the effect of MK-801 on the apoptosis of spinal cord neurons in rats that have received a fetal spinal cord (FSC) transplant following spinal hemisection was investigated. Wistar rats were divided into three groups: Spinal cord hemisection injury with a combination of FSC transplantation and MK-801 treatment (group A); spinal cord hemisection injury with FSC transplantation (group B); and spinal cord injury with insertion of a Gelfoam pledget (group C). The rats were sacrificed 1, 3, 7 and 14 days after the surgery. Apoptosis in spinal slices from the injured spinal cord was examined by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling reaction, and the expression of B-cell lymphoma-2 (Bcl-2) was measured by immunohistochemistry. The positive cells were quantitatively analyzed using a computer image analysis system. The rate of apoptosis and the positive expression of Bcl-2 protein in the spinal cord neurons in the three groups decreased in the following order: C>B>A (P<0.05) and A>B>C (P<0.05), respectively. This indicates that treatment with the NMDA receptor antagonist MK-801 prevents apoptosis in the spinal cord neurons of rats that have undergone FSC transplantation following spinal hemisection.

N-methyl-D-aspartate receptor antagonist MK-801 prevents apoptosis in rats that have undergone fetal spinal cord transplantation following spinal hemisection

PubMed Central

ZHANG, QIANG; SHAO, YANG; ZHAO, CHANGSONG; CAI, JUAN; SUN, SHENG

2014-01-01

Spinal cord injury is the main cause of paraplegia, but effective therapies for it are lacking. Embryonic spinal cord transplantation is able to repair spinal cord injury, albeit with a large amount of neuronal apoptosis remaining in the spinal cord. MK-801, an N-methyl-D-aspartate (NMDA) receptor antagonist, is able to reduce cell death by decreasing the concentration of excitatory amino acids and preventing extracellular calcium ion influx. In this study, the effect of MK-801 on the apoptosis of spinal cord neurons in rats that have received a fetal spinal cord (FSC) transplant following spinal hemisection was investigated. Wistar rats were divided into three groups: Spinal cord hemisection injury with a combination of FSC transplantation and MK-801 treatment (group A); spinal cord hemisection injury with FSC transplantation (group B); and spinal cord injury with insertion of a Gelfoam pledget (group C). The rats were sacrificed 1, 3, 7 and 14 days after the surgery. Apoptosis in spinal slices from the injured spinal cord was examined by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling reaction, and the expression of B-cell lymphoma-2 (Bcl-2) was measured by immunohistochemistry. The positive cells were quantitatively analyzed using a computer image analysis system. The rate of apoptosis and the positive expression of Bcl-2 protein in the spinal cord neurons in the three groups decreased in the following order: C>B>A (P<0.05) and A>B>C (P<0.05), respectively. This indicates that treatment with the NMDA receptor antagonist MK-801 prevents apoptosis in the spinal cord neurons of rats that have undergone FSC transplantation following spinal hemisection. PMID:25371724

Studies on the mechanism of salicylate-induced increase of insulin secretion in man.

PubMed

Giugliano, D; Cozzolino, D; Ceriello, A; Cerciello, T; Varano, R; Saccomanno, F; Torella, R

1988-01-01

Salicylate compounds are known to increase basal and stimulated insulin secretion in man. In our studies, infusion of lysine acetylsalicylate (72 mg/min) increased basal insulin levels and amplified insulin responses to glucose (5 g i.v.), arginine (5 g i.v.) and tolbutamide (1 g i.v.). Verapamil, an organic calcium antagonist, did not modify LAS-induced increase of basal insulin levels, but reduced the effect of LAS on glucose-induced insulin secretion. Calcitonin and somatostatin, two agents that inhibit basal and glucose-stimulated insulin secretion, inhibited the insulin response to glucose in presence of LAS infusion. The ability of salicylate compounds to augment insulin secretion might be due to multiple sites of action in the Beta-cells.

Calcium Sensing Receptors Mediate Local Inhibitory Reflexes Evoked by L-Phenylalanine in Guinea Pig Jejunum.

PubMed

Gwynne, Rachel M; Ly, Kenny D K N; Parry, Laura J; Bornstein, Joel C

2017-01-01

Amino acids applied to the mucosa evoke inhibitory reflexes in guinea-pig jejunum, but the receptors involved in sensory transduction are still unclear. One promising candidate is the extracellular calcium sensing receptor (CaSR), which is expressed by mucosal enteroendocrine cells and is preferentially activated by aromatic L-amino acids. We tested this by applying various amino acids to the mucosa and recording the resulting inhibitory junction potentials (IJPs) in nearby circular smooth muscle via intracellular recording. The CaSR is stereospecific and L-Phenylalanine evoked a significantly larger response than D-Phenylalanine when both were applied to the same site. The same pattern was seen with L- and D-Tryptophan, another aromatic amino acid. The CaSR is preferentially activated by aromatic amino acids and responses to L-Leucine and L-Lysine were significantly lower than those to L-Phenylalanine applied to the same site. Responses to L-Phenylalanine were dose-dependently suppressed by blockade of the CaSR with NPS2143, a CaSR antagonist, and mimicked by mucosal application of cinacalcet, a CaSR agonist. Responses to cinacalcet had similar pharmacology to that of responses to L-Phenylalanine, in that each requires both P2 purinoreceptors and 5-HT receptors. L-Glutamate evoked IJPs similar to those produced by L-Phenylalanine and these were depressed by blockade of P2 receptors and 5-HT 3 plus 5-HT 4 receptors, but NPS2143 was ineffective. The AMPA receptor antagonists DNQX (10 μM) and CNQX (10 μM) reduced IJPs evoked by L-Glutamate by 88 and 79% respectively, but neither BAY367260 (mGluR5 antagonist) nor 2APV (NMDA antagonist) affected IJPs evoked by L-Glutamate. We conclude that local inhibitory reflexes evoked by aromatic L-amino acids in guinea pig jejunum involve activation of CaSRs which triggers release of ATP and 5-HT from the mucosa. L-Glutamate also evokes inhibitory reflexes, via a pathway that does not involve CaSRs. It is likely there are multiple receptors acting as sensory transducers for different luminal amino acids.

Response of wheat seedlings to ni stress: effects of supplemental calcium.

PubMed

Ouzounidou, G; Moustakas, M; Symeonidis, L; Karataglis, S

2006-04-01

The effect of excess Ni (1 mM Ni) on wheat plants as well as the role of Ca (1 mM Ni+5000 microM Ca) for amelioration of toxicity and recovery of growth and photosynthesis in Ni-stressed wheat was evaluated. Growth, nutrient status (Ca, Mg, Fe, K, Na), and photosynthesis showed a distinct decrease strictly related to the period of treatment. Calcium ameliorated to a certain extent toxic symptoms of Ni, due to antagonistic action between Ni and Ca ions. Since chlorophyll content and variable fluorescence (Fv) decreased significantly, but Fo did not particularly change, the decrease of t1/2 with increasing duration of Ni exposure indicates negative changes on the acceptor side of PSII, which also may result from diminution of Calvin cycle. The maximum quantum yield for energy trapping was also suppressed. Plant transfer to Hoagland solution+5000 microM Ca caused recovery to plant morphology and physiology. Even in control plants, during recovery period an increased Ca concentration in plant tissues with concomitant increased rates of growth and morphology was observed. Ni concentration in plants exposed to 1 mM Ni+5000 microM Ca was lower than in plants exposed to 1 mM Ni. In all treatments a certain increase of plant nutrients was observed during recovery.

The effect of nimodipine on memory impairment during spontaneous morphine withdrawal in mice: Corticosterone interaction.

PubMed

Vaseghi, Golnaz; Rabbani, Mohammed; Hajhashemi, Valiollah

2012-11-15

Effects of the nimodipine, L-type calcium channel antagonist, has been studied on memory loss caused by spontaneous morphine withdrawal in mice. Mice were made dependent by increasing doses of morphine over three days. Memory was evaluated using object recognition task, which is based on tendency of rodents to exploration of new objects. The test was comprised of three sections: 15 min habitation, 12 min first trial and 5 min test trial. Recognition index was evaluated 4h after the last dose of morphine. Nimodipine was administrated either in chronic form (1, 5 and 10mg/kg) with daily doses of morphine or it was given as a single injection (5 and 10mg/kg) on the last day. Nimodipine in both treatment forms prevented the memory impairment following spontaneous morphine withdrawal. Corticosterone concentration was increased in brain and blood of mice during abstinence phase and pretreatment with nimodipine prevented the increase in brain and blood corticosterone concentration. The results show that blockade of L-type calcium channels improves memory deficits caused by morphine withdrawal. This indicates that some kind of treatments, such as nimodipine, administrated over the acute withdrawal phase, can prevent memory deficit during withdrawal. Copyright © 2012 Elsevier B.V. All rights reserved.

Calcium current in isolated neonatal rat ventricular myocytes.

PubMed Central

Cohen, N M; Lederer, W J

1987-01-01

1. Calcium currents (ICa) from neonatal rat ventricular heart muscle cells grown in primary culture were examined using the 'whole-cell' voltage-clamp technique (Hamill, Marty, Neher, Sakmann & Sigworth, 1981). Examination of ICa was limited to one calcium channel type, 'L' type (Nilius, Hess, Lansman & Tsien, 1985), by appropriate voltage protocols. 2. We measured transient and steady-state components of ICa, and could generally describe ICa in terms of the steady-state activation (d infinity) and inactivation (f infinity) parameters. 3. We observed that the reduction of ICa by the calcium channel antagonist D600 can be explained by both a shift of d infinity to more positive potentials as well as a slight reduction of ICa conductance. D600 did not significantly alter either the rate of inactivation of ICa or the voltage dependence of f infinity. 4. The calcium channel modulator BAY K8644 shifted both d infinity and f infinity to more negative potentials. Additionally, BAY K8644 increased the rate of inactivation at potentials between +5 and +55 mV. Furthermore, BAY K8644 also increased ICa conductance, a change consistent with a promotion of 'mode 2' calcium channel activity (Hess, Lansman & Tsien, 1984). 5. We conclude that, as predicted by d infinity and f infinity, there is a significant steady-state component of ICa ('window current') at plateau potentials in neonatal rat heart cells. Modulation of the steady-state and transient components of ICa by various agents can be attributed both to specific alterations in d infinity and f infinity and to more complicated alterations in the mode of calcium channel activity. PMID:2451004

Metabotropic glutamate receptors activate dendritic calcium waves and TRPM channels which drive rhythmic respiratory patterns in mice

PubMed Central

Mironov, S L

2008-01-01

Respiration in vertebrates is generated by a compact network which is located in the lower brainstem but cellular mechanisms which underlie persistent oscillatory activity of the respiratory network are yet unknown. Using two-photon imaging and patch-clamp recordings in functional brainstem preparations of mice containing pre-Bötzinger complex (preBötC), we examined the actions of metabotropic glutamate receptors (mGluR1/5) on the respiratory patterns. The agonist DHPG potentiated and antagonist LY367385 depressed respiration-related activities. In the inspiratory neurons, we observed rhythmic activation of non-selective channels which had a conductance of 24 pS. Their activity was enhanced with membrane depolarization and after elevation of calcium from the cytoplasmic side of the membrane. They were activated by a non-hydrolysable PIP2 analogue and blocked by flufenamate, ATP4− and Gd3+. All these properties correspond well to those of TRPM4 channels. Calcium imaging of functional slices revealed rhythmic transients in small clusters of neurons present in a network. Calcium transients in the soma were preceded by the waves in dendrites which were dependent on mGluR activation. Initiation and propagation of waves required calcium influx and calcium release from internal stores. Calcium waves activated TPRM4-like channels in the soma and promoted generation of inspiratory bursts. Simulations of activity of neurons communicated via dendritic calcium waves showed emerging activity within neuronal clusters and its synchronization between the clusters. The experimental and theoretical data provide a subcellular basis for a recently proposed group-pacemaker hypothesis and describe a novel mechanism of rhythm generation in neuronal networks. PMID:18308826

Doxazosin in the current treatment of hypertension.

PubMed

Wykretowicz, Andrzej; Guzik, Przemyslaw; Wysocki, Henryk

2008-03-01

Evidence from various studies has shown that even a very small reduction in the blood pressure explains the majority of benefits in cardiovascular outcomes. Although doxazosin is a relatively old drug, it is a very effective add-on therapeutic agent. It has been used in a variety of clinical trials, including different groups of hypertensive patients such as diabetics, the elderly, patients with benign prostatic hyperplasia or hypercholesterolaemia, the obese or Afro-Americans and in combination with all major groups of antihypertensive drugs such as: calcium channel antagonists, diuretics, beta-adrenoreceptor antagonists, angiotensin-converting enzyme inhibitors and angiotensin-2 receptor blockers. The purpose of this article is to review the role of doxazosin in the current treatment of hypertension. English-language reports published in 1986 - 2007 were retrieved. Data extraction was based on articles reporting at least 15 or more patients treated with combination of doxazosin with other antihypertensive drugs. In these studies doxazosin in either standard or extended-release formulation was added as a second- or third-line agent. The two formulations of doxazosin, although chemically identical, appear to differ in some other effects. A large proportion of patients demonstrated a favourable blood pressure response with relatively few treatment-associated side effects showing that this drug appears to be a valuable add-on antihypertensive treatment option. Doxazosin reduces blood pressure and additionally influences other risk factors of coronary heart disease. However, it should be clearly stated that the clinical role of 'beyond blood pressure' effects of doxazosin are largely undetermined.

David J. Triggle: Medicinal chemistry, to pharmacology, calcium channels, and beyond.

PubMed

Walker, Michael J A

2015-11-15

David Triggle's scientific career began as a chemist, went through medicinal chemistry into pharmacology, and finally on to somewhat more philosophical interests in later years. It was a career marked by many contributions to all of those aspects of science. Chief amongst his many contributions, in addition to those in medicinal chemistry, was his work on the drugs known as calcium ion channel blockers or (calcium antagonists). In the calcium ion channel field he was a particularly instrumental figure in sorting out the mechanisms, actions and roles of the class of calcium channel blockers, known chemical and pharmacologically as the dihydropyridines (DHPs) in particular, as well as other calcium blockers of diverse structures. During the course of a long career, and extensive journeys into medicinal chemistry and pharmacology, he published voluminously in terms of papers, reviews, conference proceedings and books. Notably, many of his papers often had limited authorship where, as senior author it reflected his deep involvement in all aspects of the reported work. His work always helped clarify the field while his incisive reviews, together with his role in coordinating and running scientific meetings, were a great help in clarifying and organizing various fields of study. He has had a long and illustrious career, and is wellknown in the world of biomedical science; his contributions are appreciated, and well recognized everywhere. The following article attempts to chart a path through his work and contributions to medicinal chemistry, pharmacology, science, academia and students. Copyright © 2015 Elsevier Inc. All rights reserved.

Angiotensin II receptor blocker-based therapy in Japanese elderly, high-risk, hypertensive patients.

PubMed

Ogawa, Hisao; Kim-Mitsuyama, Shokei; Matsui, Kunihiko; Jinnouchi, Tomio; Jinnouchi, Hideaki; Arakawa, Kikuo

2012-10-01

It is unknown whether high-dose angiotensin II receptor blocker therapy or angiotensin II receptor blocker + calcium channel blocker combination therapy is better in elderly hypertensive patients with high cardiovascular risk. The objective of the study was to compare the efficacy of these treatments in elderly, high-risk Japanese hypertensive patients. The OlmeSartan and Calcium Antagonists Randomized (OSCAR) study was a multicenter, prospective, randomized, open-label, blinded-end point study of 1164 hypertensive patients aged 65 to 84 years with type 2 diabetes or cardiovascular disease. Patients with uncontrolled hypertension during treatment with olmesartan 20 mg/d were randomly assigned to receive 40 mg/d olmesartan (high-dose angiotensin II receptor blocker) or a calcium channel blocker + 20 mg/d olmesartan (angiotensin II receptor blocker + calcium channel blocker). The primary end point was a composite of cardiovascular events and noncardiovascular death. During a 3-year follow-up, blood pressure was significantly lower in the angiotensin II receptor blocker + calcium channel blocker group than in the high-dose angiotensin II receptor blocker group. Mean blood pressure at 36 months was 135.0/74.3 mm Hg in the high-dose angiotensin II receptor blocker group and 132.6/72.6 mm Hg in the angiotensin II receptor blocker + calcium channel blocker group. More primary end points occurred in the high-dose angiotensin II receptor blocker group than in the angiotensin II receptor blocker + calcium channel blocker group (58 vs 48 events, hazard ratio [HR], 1.31, 95% confidence interval, 0.89-1.92; P=.17). In patients with cardiovascular disease at baseline, more primary events occurred in the high-dose angiotensin II receptor blocker group (HR, 1.63, P=.03); in contrast, fewer events were observed in the subgroup without cardiovascular disease (HR, 0.52, P=.14). This treatment-by-subgroup interaction was significant (P=.02). The angiotensin II receptor blocker and calcium channel blocker combination lowered blood pressure more than the high-dose angiotensin II receptor blocker and reduced the incidence of primary end points more than the high-dose angiotensin II receptor blocker in patients with cardiovascular disease. The addition of a second antihypertensive agent is more effective at lowering blood pressure than simply doubling the dose of an existing agent. Copyright © 2012 Elsevier Inc. All rights reserved.

Houttuynia cordata Improves Cognitive Deficits in Cholinergic Dysfunction Alzheimer's Disease-Like Models.

PubMed

Huh, Eugene; Kim, Hyo Geun; Park, Hanbyeol; Kang, Min Seo; Lee, Bongyong; Oh, Myung Sook

2014-05-01

Cognitive impairment is a result of dementia of diverse causes, such as cholinergic dysfunction and Alzheimer's disease (AD). Houttuynia cordata Thunb. (Saururaceae) has long been used as a traditional herbal medicine. It has biological activities including protective effects against amyloid beta (Aβ) toxicity, via regulation of calcium homeostasis, in rat hippocampal cells. To extend previous reports, we investigated the effects of water extracts of H. cordata herb (HCW) on tauopathies, also involving calcium influx. We then confirmed the effects of HCW in improving memory impairment and neuronal damage in mice with Aβ-induced neurotoxicity. We also investigated the effects of HCW against scopolamine-induced cholinergic dysfunction in mice. In primary neuronal cells, HCW inhibited the phosphorylation of tau by regulating p25/p35 expression in Aβ-induced neurotoxicity. In mice with Aβ-induced neurotoxicity, HCW improved cognitive impairment, as assessed with behavioral tasks, such as novel object recognition, Y-maze, and passive avoidance tasks. HCW also inhibited the degeneration of neurons in the CA3 region of the hippocampus in Aβ-induced neurotoxicity. Moreover, HCW, which had an IC50 value of 79.7 μg/ml for acetylcholinesterase inhibition, ameliorated scopolamine-induced cognitive impairment significantly in Y-maze and passive avoidance tasks. These results indicate that HCW improved cognitive impairment, due to cholinergic dysfunction, with inhibitory effects against tauopathies and cholinergic antagonists, suggesting that HCW may be an interesting candidate to investigate for the treatment of AD.

Houttuynia cordata Improves Cognitive Deficits in Cholinergic Dysfunction Alzheimer’s Disease-Like Models

PubMed Central

Huh, Eugene; Kim, Hyo Geun; Park, Hanbyeol; Kang, Min Seo; Lee, Bongyong; Oh, Myung Sook

2014-01-01

Cognitive impairment is a result of dementia of diverse causes, such as cholinergic dysfunction and Alzheimer’s disease (AD). Houttuynia cordata Thunb. (Saururaceae) has long been used as a traditional herbal medicine. It has biological activities including protective effects against amyloid beta (Aβ) toxicity, via regulation of calcium homeostasis, in rat hippocampal cells. To extend previous reports, we investigated the effects of water extracts of H. cordata herb (HCW) on tauopathies, also involving calcium influx. We then confirmed the effects of HCW in improving memory impairment and neuronal damage in mice with Aβ-induced neurotoxicity. We also investigated the effects of HCW against scopolamine-induced cholinergic dysfunction in mice. In primary neuronal cells, HCW inhibited the phosphorylation of tau by regulating p25/p35 expression in Aβ-induced neurotoxicity. In mice with Aβ-induced neurotoxicity, HCW improved cognitive impairment, as assessed with behavioral tasks, such as novel object recognition, Y-maze, and passive avoidance tasks. HCW also inhibited the degeneration of neurons in the CA3 region of the hippocampus in Aβ-induced neurotoxicity. Moreover, HCW, which had an IC50 value of 79.7 μg/ml for acetylcholinesterase inhibition, ameliorated scopolamine-induced cognitive impairment significantly in Y-maze and passive avoidance tasks. These results indicate that HCW improved cognitive impairment, due to cholinergic dysfunction, with inhibitory effects against tauopathies and cholinergic antagonists, suggesting that HCW may be an interesting candidate to investigate for the treatment of AD. PMID:25009697

Calcilytic Ameliorates Abnormalities of Mutant Calcium-Sensing Receptor (CaSR) Knock-In Mice Mimicking Autosomal Dominant Hypocalcemia (ADH).

PubMed

Dong, Bingzi; Endo, Itsuro; Ohnishi, Yukiyo; Kondo, Takeshi; Hasegawa, Tomoka; Amizuka, Norio; Kiyonari, Hiroshi; Shioi, Go; Abe, Masahiro; Fukumoto, Seiji; Matsumoto, Toshio

2015-11-01

Activating mutations of calcium-sensing receptor (CaSR) cause autosomal dominant hypocalcemia (ADH). ADH patients develop hypocalcemia, hyperphosphatemia, and hypercalciuria, similar to the clinical features of hypoparathyroidism. The current treatment of ADH is similar to the other forms of hypoparathyroidism, using active vitamin D3 or parathyroid hormone (PTH). However, these treatments aggravate hypercalciuria and renal calcification. Thus, new therapeutic strategies for ADH are needed. Calcilytics are allosteric antagonists of CaSR, and may be effective for the treatment of ADH caused by activating mutations of CaSR. In order to examine the effect of calcilytic JTT-305/MK-5442 on CaSR harboring activating mutations in the extracellular and transmembrane domains in vitro, we first transfected a mutated CaSR gene into HEK cells. JTT-305/MK-5442 suppressed the hypersensitivity to extracellular Ca(2+) of HEK cells transfected with the CaSR gene with activating mutations in the extracellular and transmembrane domains. We then selected two activating mutations locating in the extracellular (C129S) and transmembrane (A843E) domains, and generated two strains of CaSR knock-in mice to build an ADH mouse model. Both mutant mice mimicked almost all the clinical features of human ADH. JTT-305/MK-5442 treatment in vivo increased urinary cAMP excretion, improved serum and urinary calcium and phosphate levels by stimulating endogenous PTH secretion, and prevented renal calcification. In contrast, PTH(1-34) treatment normalized serum calcium and phosphate but could not reduce hypercalciuria or renal calcification. CaSR knock-in mice exhibited low bone turnover due to the deficiency of PTH, and JTT-305/MK-5442 as well as PTH(1-34) increased bone turnover and bone mineral density (BMD) in these mice. These results demonstrate that calcilytics can reverse almost all the phenotypes of ADH including hypercalciuria and renal calcification, and suggest that calcilytics can become a novel therapeutic agent for ADH. © 2015 American Society for Bone and Mineral Research.

Microfluorimetric analysis of a purinergic receptor (P2X7) in GH4C1 rat pituitary cells: effects of a bioactive substance produced by Pfiesteria piscicida.

PubMed Central

Melo, A C; Moeller, P D; Glasgow, H; Burkholder, J M; Ramsdell, J S

2001-01-01

Pfiesteria piscicida Steidinger & Burkholder is a toxic dinoflagellate that leads to fish and human toxicity. It produces a bioactive substance that leads to cytotoxicity of GH4C1 rat pituitary cells. Extracellular adenosine 5'-triphosphate (ATP) acting on P2X7 purinergic receptors induces the formation of a nonselective cation channel, causing elevation of the cytosolic free calcium followed by a characteristic permeabilization of the cell to progressively larger ions and subsequent cell lysis. We investigated whether GH4C1 rat pituitary cells express functional P2X7 receptors, and if so, are they activated by a bioactive substance isolated from toxic P. piscicida cultures. We tested the selective agonist 2'-3'-O-(benzoyl-4-benzoyl)-ATP (BzATP) and antagonists piridoxalphosphate-6-azophenyl-2'-4'-disulfonic acid (PPADS) and oxidized-ATP (oxATP) using elevated cytosolic free calcium in Fura-2 loaded cells, and induced permeability of these cells to the fluorescent dye YO-PRO-1 as end points. We demonstrated that in GH4C1 cells, BzATP induces both the elevation of cytosolic free calcium and the permeabilization of the cell membrane. ATP-induced membrane permeabilization was inhibited by PPADS reversibly and by oxATP irreversibly. The putative Pfiesteria toxin (pPfTx) also elevated cytosolic free calcium in Fura-2 in GH4C1 cells and increased the permeability to YO-PRO-1 in a manner inhibited fully by oxATP. This study indicates that GH4C1 cells express a purinoceptor with characteristics consistent with the P2X7 subtype, and that pPfTx mimics the kinetics of cell permeabilization by ATP. PMID:11677182

Endothelin receptors and activity differ in human, dog, and rabbit lung.

PubMed

McKay, K O; Armour, C L; Black, J L

1996-01-01

In this study, we have examined dog and rabbit airways as potential models for human airways in regard to the activity of endothelin. The receptors involved in the response to endothelin-1 (ET-1) in airway tissue from human, rabbit, and dog lung were investigated, as was the mechanism responsible for the contraction to ET-1 in tissue from the three species. By using specific endothelin receptor agonists and antagonists, we have demonstrated that ETB receptors predominate in rabbit and human airways and ETA receptors in dog airways. The contraction to ET-1 is not dependent on cyclooxygenase products of arachidonic acid, as indomethacin had no effect on the response to ET-1. Extracellular calcium influx via voltage-dependent channels is necessary for contraction to ET-1 in rabbit and dog airways. These results are in contrast to our previously reported results in human airways, in which neither removal of extracellular calcium nor verapamil affected the ET-1 response. The sustained phase of the contraction to ET-1 in all three species may be mediated in part by activation of protein kinase C (PKC), as the inhibitor staurosporine significantly altered the time course of the response to endothelin. We therefore conclude that in rabbit airways ET-1 activates ETB receptors, triggers the influx of extracellular calcium through voltage-dependent channels, and induces a contractile response that is, in part, dependent upon stimulation of PKC. The same mechanism is triggered in dog bronchus; however, the receptors involved in this species are of the ETA type. Finally, in human airways, the contractile response to ET-1, while independent of extracellular calcium influx, is dependent upon PKC activation after binding of the peptide to ETB receptors.

Pinoresinol-4,4'-di-O-beta-D-glucoside from Valeriana officinalis root stimulates calcium mobilization and chemotactic migration of mouse embryo fibroblasts.

PubMed

Do, Kee Hun; Choi, Young Whan; Kim, Eun Kyoung; Yun, Sung Ji; Kim, Min Sung; Lee, Sun Young; Ha, Jung Min; Kim, Jae Ho; Kim, Chi Dae; Son, Beung Gu; Kang, Jum Soon; Khan, Ikhlas A; Bae, Sun Sik

2009-06-01

Lignans are major constituents of plant extracts and have important pharmacological effects on mammalian cells. Here we showed that pinoresinol-4,4'-di-O-beta-D-glucoside (PDG) from Valeriana officinalis induced calcium mobilization and cell migration through the activation of lysophosphatidic acid (LPA) receptor subtypes. Stimulation of mouse embryo fibroblast (MEF) cells with 10 microM PDG resulted in strong stimulation of MEF cell migration and the EC(50) was about 2 microM. Pretreatment with pertussis toxin (PTX), an inhibitor of G(i) protein, completely blocked PDG-induced cell migration demonstrating that PDG evokes MEF cell migration through the activation of the G(i)-coupled receptor. Furthermore, pretreatment of MEF cells with Ki16425 (10 microM), which is a selective antagonist for LPA(1) and LPA(3) receptors, completely blocked PDG-induced cell migration. Likewise, PDG strongly induced calcium mobilization, which was also blocked by Ki16425 in a dose-dependent manner. Prior occupation of the LPA receptor with LPA itself completely blocked PDG-induced calcium mobilization. Finally, PDG-induced MEF cell migration was attenuated by pretreatment with a phosphatidylinositol 3-kinase (PI3K) inhibitor such as LY294002. Cells lacking downstream mediator of PI3K such as Akt1 and Akt2 (DKO cells) showed loss of PDG-induced migration. Re-expression of Akt1 (but not Akt2) completely restored PDG-induced DKO cell migration. Given these results, we conclude that PDG is a strong inducer of cell migration. We suggest that the pharmacological action of PDG may occur through the activation of an LPA receptor whereby activation of PI3K/Akt signaling pathway mediates PDG-induced MEF cell migration.

[Effects of 2-(p-dimethylaminostyryl) pyridine methycholide (DSPM-Ci) on ECG, left atrium contractivity and on papillary muscle action potentials].

PubMed

Jiang, X Y; Zhou, C M; Li, D M; Zhang, K J

1996-01-01

The effects of DSPM-Cl on ECG in rats, on the dose-effect curve in guinea pig left atria and on the fast action potential (AP), high-K+ depolarized slow action potential (SAP) in guinea pigs papillary muscle were examined electrophysiologically. DSPM-Cl (2 mg.kg-1) showed significant nagative frequency, negative conductivity effect, and prolonged the PP and PR interval. DSPM-CI (30-50 mumol.L-1) was shown to inhibit left atria contractility and shift the concentration-response curve of Iso and CaCl2 to the right with PD2' values of 4.60 and 4.13, respectively. In addition, DSPM-Cl was found to prolong the duration of action potential 90 (APD90) and effective refractory period (ERP), and decrease the maximal upstroke velocity (Vmax) in K(+)-depolarized guinea pigs papillary muscles. The results suggest that, like verpamil, DSPM-Cl might be a calcium antagonist.

Hereditary vacuolar internal anal sphincter myopathy causing proctalgia fugax and constipation: a new case contribution.

PubMed

de la Portilla, Fernando; Borrero, Juan José; Rafel, Enrique

2005-03-01

Hereditary anal sphincter myopathy is rare. We present a family with one affected member with proctalgia fugax, constipation and internal anal sphincter hypertrophy. Ultrastructural findings show vacuolization of smooth muscle cells without the characteristic polyglucosan inclusion. Further relief of symptoms was obtained using an oral calcium antagonist. Based on clinical presentation, endosonography and morphological findings, we consider our case is a histological variant of the vacuolar myopathy originally described.

Diffusion of dihydropyridine calcium channel antagonists in cardiac sarcolemmal lipid multibilayers.

PubMed Central

Chester, D W; Herbette, L G; Mason, R P; Joslyn, A F; Triggle, D J; Koppel, D E

1987-01-01

A membrane bilayer pathway model has been proposed for the interaction of dihydropyridine (DHP) calcium channel antagonists with receptors in cardiac sarcolemma (Rhodes, D.G., J.G. Sarmiento, and L.G. Herbette. 1985. Mol. Pharmacol. 27:612-623) involving drug partition into the bilayer with subsequent receptor binding mediated (though probably not rate-limited) by diffusion within the bilayer. Recently, we have characterized the partition step, demonstrating that DHPs reside, on a time-average basis, near the bilayer hydrocarbon core/water interface. Drug distribution about this interface may define a plane of local concentration for lateral diffusion within the membrane. The studies presented herein examine the diffusional dynamics of an active rhodamine-labeled DHP and a fluorescent phospholipid analogue (DiIC16) in pure cardiac sarcolemmal lipid multibilayer preparations as a function of bilayer hydration. At maximal bilayer hydration, the drug diffuses over macroscopic distances within the bilayer at a rate identical to that of DiI (D = 3.8 X 10(-8) cm2/s), demonstrating the overall feasibility of the membrane diffusion model. The diffusion coefficients for both drug and lipid decreased substantially as the bilayers were dehydrated. While identical at maximal hydration, drug diffusion was significantly slower than that of DiIC16 in partially dehydrated bilayers, probably reflecting differences in mass distribution of these probes in the bilayer. Images FIGURE 1 FIGURE 2 FIGURE 3 FIGURE 4 PMID:2447967

Formulation and optimisation of raft-forming chewable tablets containing H2 antagonist

PubMed Central

Prajapati, Shailesh T; Mehta, Anant P; Modhia, Ishan P; Patel, Chhagan N

2012-01-01

Purpose: The purpose of this research work was to formulate raft-forming chewable tablets of H2 antagonist (Famotidine) using a raft-forming agent along with an antacid- and gas-generating agent. Materials and Methods: Tablets were prepared by wet granulation and evaluated for raft strength, acid neutralisation capacity, weight variation, % drug content, thickness, hardness, friability and in vitro drug release. Various raft-forming agents were used in preliminary screening. A 23 full-factorial design was used in the present study for optimisation. The amount of sodium alginate, amount of calcium carbonate and amount sodium bicarbonate were selected as independent variables. Raft strength, acid neutralisation capacity and drug release at 30 min were selected as responses. Results: Tablets containing sodium alginate were having maximum raft strength as compared with other raft-forming agents. Acid neutralisation capacity and in vitro drug release of all factorial batches were found to be satisfactory. The F5 batch was optimised based on maximum raft strength and good acid neutralisation capacity. Drug–excipient compatibility study showed no interaction between the drug and excipients. Stability study of the optimised formulation showed that the tablets were stable at accelerated environmental conditions. Conclusion: It was concluded that raft-forming chewable tablets prepared using an optimum amount of sodium alginate, calcium carbonate and sodium bicarbonate could be an efficient dosage form in the treatment of gastro oesophageal reflux disease. PMID:23580933

Formulation and optimisation of raft-forming chewable tablets containing H2 antagonist.

PubMed

Prajapati, Shailesh T; Mehta, Anant P; Modhia, Ishan P; Patel, Chhagan N

2012-10-01

The purpose of this research work was to formulate raft-forming chewable tablets of H2 antagonist (Famotidine) using a raft-forming agent along with an antacid- and gas-generating agent. Tablets were prepared by wet granulation and evaluated for raft strength, acid neutralisation capacity, weight variation, % drug content, thickness, hardness, friability and in vitro drug release. Various raft-forming agents were used in preliminary screening. A 2(3) full-factorial design was used in the present study for optimisation. The amount of sodium alginate, amount of calcium carbonate and amount sodium bicarbonate were selected as independent variables. Raft strength, acid neutralisation capacity and drug release at 30 min were selected as responses. Tablets containing sodium alginate were having maximum raft strength as compared with other raft-forming agents. Acid neutralisation capacity and in vitro drug release of all factorial batches were found to be satisfactory. The F5 batch was optimised based on maximum raft strength and good acid neutralisation capacity. Drug-excipient compatibility study showed no interaction between the drug and excipients. Stability study of the optimised formulation showed that the tablets were stable at accelerated environmental conditions. It was concluded that raft-forming chewable tablets prepared using an optimum amount of sodium alginate, calcium carbonate and sodium bicarbonate could be an efficient dosage form in the treatment of gastro oesophageal reflux disease.

Calcium dynamics in tomato pollen tubes using the Yellow Cameleon 3.6 sensor.

PubMed

Barberini, María Laura; Sigaut, Lorena; Huang, Weijie; Mangano, Silvina; Juarez, Silvina Paola Denita; Marzol, Eliana; Estevez, José; Obertello, Mariana; Pietrasanta, Lía; Tang, Weihua; Muschietti, Jorge

2018-06-01

In vitro tomato pollen tubes show a cytoplasmic calcium gradient that oscillates with the same period as growth. Pollen tube growth requires coordination between the tip-focused cytoplasmic calcium concentration ([Ca 2+ ] cyt ) gradient and the actin cytoskeleton. This [Ca 2+ ] cyt gradient is necessary for exocytosis of small vesicles, which contributes to the delivery of new membrane and cell wall at the pollen tube tip. The mechanisms that generate and maintain this [Ca 2+ ] cyt gradient are not completely understood. Here, we studied calcium dynamics in tomato (Solanum lycopersicum) pollen tubes using transgenic tomato plants expressing the Yellow Cameleon 3.6 gene under the pollen-specific promoter LAT52. We use tomato as an experimental model because tomato is a Solanaceous plant that is easy to transform, and has an excellent genomic database and genetic stock center, and unlike Arabidopsis, tomato pollen is a good system to do biochemistry. We found that tomato pollen tubes showed an oscillating tip-focused [Ca 2+ ] cyt gradient with the same period as growth. Then, we used a pharmacological approach to disturb the intracellular Ca 2+ homeostasis, evaluating how the [Ca 2+ ] cyt gradient, pollen germination and in vitro pollen tube growth were affected. We found that cyclopiazonic acid (CPA), a drug that inhibits plant P IIA -type Ca 2+ -ATPases, increased [Ca 2+ ] cyt in the subapical zone, leading to the disappearance of the Ca 2+ oscillations and inhibition of pollen tube growth. In contrast, 2-aminoethoxydiphenyl borate (2-APB), an inhibitor of Ca 2+ released from the endoplasmic reticulum to the cytoplasm in animals cells, completely reduced [Ca 2+ ] cyt at the tip of the tube, blocked the gradient and arrested pollen tube growth. Although both drugs have antagonistic effects on [Ca 2+ ] cyt , both inhibited pollen tube growth triggering the disappearance of the [Ca 2+ ] cyt gradient. When CPA and 2-APB were combined, their individual inhibitory effects on pollen tube growth were partially compensated. Finally, we found that GsMTx-4, a peptide from spider venom that blocks stretch-activated Ca 2+ channels, inhibited tomato pollen germination and had a heterogeneous effect on pollen tube growth, suggesting that these channels are also involved in the maintenance of the [Ca 2+ ] cyt gradient. All these results indicate that tomato pollen tube is an excellent model to study calcium dynamics.

Vasodilators and α-adrenoceptor antagonists in hypertension and heart failure

PubMed Central

Taylor, S. H.

1981-01-01

1 The mechanism of the increase in arteriolar resistance in hypertension and heart failure is differently derived. In hypertension, venous compliance is normal and the concentric narrowing of the arteriolar resistance vessels is `anatomical'; it is not due to increased stimulation or enhanced sensitivity of the vascular smooth muscle. In heart failure narrowing of both the arteriolar resistance and venous capacitance vessels derives predominantly from increased sympathoadrenal stimulation of α1-adrenoceptors in the vascular smooth muscle. 2 Vasodilator drugs which relax vascular smooth muscle differ widely in their site of activity. None are entirely specific for arteries, arterioles or veins, but they may be grouped for therapeutic convenience into those predominantly acting on arterioles (for example hydralazine) and those acting on veins (for example nitrates). 3 Control of the resting blood pressure in stable essential hypertension appears to be equally well achieved with non-specific arteriolar dilators (for example hydralazine, minoxidil, calcium antagonists) as those with specific α1-adrenoceptor blocking properties (for example prazosin, indoramin). Pressure surges due to dynamic exercise and mental stress are little influenced by either category of drug. In contrast, α-adrenoceptor antagonists appear to be capable of partly suppressing increase in ambulatory pressure and the pressor responses to isometric exercise and cold, particularly in patients pre-treated with β-blocking drugs. 4 In acute heart failure, non-selective α-blocking drugs (for example phentolamine) produce an equal reduction in left ventricular filling pressure but greater increase in cardiac output than vasodilator drugs with a more balanced relaxing effect on arterioles and venules. 5 In chronic heart failure, the little information available indicates that non-selective arteriolar dilatation is probably associated with a greater increase in cardiac output lesser reduction in left ventricular filling pressure than with specific α1-adrenoceptor blocking drugs. Attenuation of the initial haemodynamic benefits during extended treatment is common to all vasodilators, including α-adrenoceptor antagonists. 6 α-Adrenoceptor antagonists have yet to be convincingly shown to be haemodynamically superior to drugs with less specific vasodilator activity in the treatment of hypertension and heart failure.

The relevance of kalikrein-kinin system via activation of B2 receptor in LPS-induced fever in rats.

PubMed

Soares, Denis de Melo; Santos, Danielle R; Rummel, Christoph; Ott, Daniela; Melo, Míriam C C; Roth, Joachim; Calixto, João B; Souza, Glória E P

2017-11-01

This study evaluated the involvement of endogenous kallikrein-kinin system and the bradykinin (BK) B 1 and B 2 receptors on LPS- induced fever and the POA cells involved in this response. Male Wistar rats received either i.v. (1 mg/kg), i.c.v. (20 nmol) or i.h. (2 nmol) injections of icatibant (B 2 receptor antagonist) 30 or 60 min, respectively, before the stimuli. DALBK (B 1 receptor antagonist) was given either 15min before BK (i.c.v.) or 30 min before LPS (i.v.). Captopril (5 mg/kg, sc.,) was given 1 h prior LPS or BK. Concentrations of BK and total kininogenon CSF, plasma and tissue kallikrein were evaluated. Rectal temperatures (rT) were assessed by telethermometry. Ca ++ signaling in POA cells was performed in rat pup brain tissue microcultures. Icatibant reduced LPS fever while, captopril exacerbated that response, an effect abolished by icatibant. Icatibant (i.h.) reduced fever to BK (i.h.) but not that induced by LPS (i.v.). BK increased intracellular calcium concentration in neurons and astrocytes. LPS increased levels of bradykinin, tissue kallikrein and total kininogen. BK (i.c.v.) increased rT and decreased tail skin temperature. Captopril potentiated BK-induced fever an effect abolished by icatibant. DALBK reduced the fever induced by BK. BK (i.c.v.) increased the CSF PGE 2 concentration. Effect abolished by indomethacin (i.p.). LPS activates endogenous kalikrein-kinin system leading to production of BK, which by acting on B 2 -receptors of POA cells causes prostaglandin synthesis that in turn produces fever. Thus, a kinin B 2 -receptor antagonist that enters into the brain could constitute a new and interesting strategy to treat fever. Copyright © 2017 Elsevier Ltd. All rights reserved.

Preterm labour: an overview of current and emerging therapeutics.

PubMed

Schwarz, Matthias K; Page, Patrick

2003-08-01

Preterm labour is a major cause of perinatal mortality and morbidity. However, during the past 40 years of clinical studies and despite the use of multiple therapeutic agents, the rate of preterm birth has not drastically declined. In 1991, it was estimated that in the US approximately 116,000 women admitted with acute episodes of preterm labour were treated each year with ritodrine, which is the first drug approved by the US FDA and still remains the standard therapy for treating preterm labour. Ritodrine (Yutopar( trade mark )) stimulates the beta(2)-adrenergic receptor throughout the body, causing an inhibitory action in different tissues that, among other side effects, also leads to an attenuation of uterine contractility. More recently, a new therapeutic agent, atosiban (Tractocile( trade mark )), a peptidic oxytocin receptor antagonist, has been introduced to the market. However, the use of the various pharmacological agents to treat preterm labour remains restricted, due to lack of uterine selectivity, low efficacy and potentially serious side effects for the mother or the foetus. Therefore, there is an urgent need to develop drugs with myometrial selectivity that would allow long-lasting inhibition of labour and prolong pregnancy up to a stage when good foetal maturation raises the chances of survival. One of the major obstacles hampering the development of new therapeutic agents is the marked inter-species difference in terms of preterm labour physiology, which complicates the preclinical evaluation of new candidate molecules in animal models of disease. In this review, the authors will provide a comprehensive update of past, current and new approaches for the management of preterm labour, including beta(2)-adrenergic agonists, calcium channel blockers, oxytocin antagonists, prostaglandin antagonists and other potential therapeutics. For each of the therapies used today, the review will cover the mechanism of action, benefit and adverse effects, and discuss the promise and potential benefits of new emerging therapeutic agents.

6-[N,S-dimethyl-N'-cyanothioureidomethyl]-6,11-dihydro-5H- dibenz[b,e]azepine hydrochloride (Fran 12): a histamine and 5-hydroxytryptamine antagonist with pressor properties.

PubMed

Law, S C; Guyett, F J; King, R G; Boura, A L; Jackson, W R; Hodgson, W C

1992-01-01

We have synthesized and examined some of the pharmacological properties of 6-[N,S-dimethyl-N'-cyanoisothioureidomethyl]-6,11-dihydro-5H- dibenz(b,e)azepine hydrochloride (Fran 12), a derivative of 6-methylaminomethyl-6,11-dihydro-5H- dibenz[b,e,]azepine. In the guinea-pig isolated ileum, Fran 12 (10(-7)-10(-5) M) caused parallel rightward shifts of the concentration-response curves to histamine. A Schild plot gave a pA2 of 7.48, with a slope not significantly different from -1.0. In the rat stomach fundus strip and in endothelium-denuded aortic rings, Fran 12 inhibited contractile responses to 5-hydroxytryptamine in a non-competitive manner. In both chloralose-anaesthetized and pithed rats, it inhibited pressor responses to 5-hydroxytryptamine. It had no effect on depressor responses to 5-hydroxytryptamine in anaesthetized rats. In pithed rats, Fran 12 (0.25-2 mg/kg, i.v.) produced dose-dependent increases in blood pressure. These were not inhibited by i.v. phentolamine, prazosin, yohimbine, propranolol, methysergide, pentolinium or atropine but were inhibited by verapamil. These results indicate that Fran 12 is a histamine and 5-hydroxytryptamine antagonist which also exerts pressor effects via a peripheral action. The pressor action does not appear to be mediated via effects on alpha 1- or alpha 2-adrenoceptors, muscarinic or nicotinic cholinoceptors or 5-hydroxytryptamine receptors, although calcium channel activation may play a role.

A plant Kunitz-type inhibitor mimics bradykinin-induced cytosolic calcium increase and intestinal smooth muscle contraction.

PubMed

Andrade, Sheila Siqueira; Smaili, Soraya Soubhi; Monteforte, Priscila Totarelli; Miranda, Antônio; Kouyoumdjian, Maria; Sampaio, Misako Uemura; Lopes, Guiomar Silva; Oliva, Maria Luiza V

2012-09-01

BbKI is a kallikrein inhibitor with a reactive site sequence similar to that of kinins, the vasoactive peptides inserted in kininogen moieties. This structural similarity probably contributes to the strong interaction with plasma kallikrein, the enzyme that releases, from high-molecular weight kininogen (HMWK), the proinflammatory peptide bradykinin, which acts on B(2) receptors (B(2)R). BbKI was examined on smooth muscle contraction and Ca(2+) mobilization, in which the kallikrein-kinin system is involved. Contrary to expectations, BbKI (1.8 μm) increased [Ca(2+)](c) and contraction, as observed with BK (2.0 μm). Not blocked by B(1) receptors (B(1)R), the BbKI agonistic effect was blocked by the B(2)R antagonist, HOE-140 (6 μm), and the involvement of B(2)R was confirmed in B(2)R-knockout mice intestine. The same tissue response was obtained using a synthetic peptide derived from the BbKI reactive site structure, more resistant than BK to angiotensin I-converting enzyme (ACE) hydrolysis. Depending on the concentration, BbKI has a dual effect. At a low concentration, BbKI acts as a potent kallikrein inhibitor; however, due to the similarity to BK, in high concentrations, BbKI greatly increases Ca(2+) release from internal storages, as a consequence of its interaction with B(2)R. Therefore, the antagonistic and agonistic effects of BbKI may be considered in conditions of B(2)R involvement.

[Exercise in arterial hypertension].

PubMed

Predel, Hans-Georg; Schramm, Thomas

2006-09-01

Regular endurance training has established itself as a major therapeutic principle in the specter of nonpharmacological measures in arterial hypertension. An initial medical check as well as an adequate technique, dosage and intensity of the prescribed exercise training are mandatory. With respect to the concomitant pharmacological treatment, it should be considered that the beneficial effects of lifestyle modification will not be counteracted by the chosen antihypertensive drug but, ideally, synergistically supported. Based on the individual clinical situation, principally all antihypertensive drugs recommended by the current European guidelines, may be prescribed as mono- or combination therapy.beta-receptor blockers are especially capable of controlling excessive exercise-induced blood pressure increase; however, they have metabolic and exercise physiological limitations. The neutrality concerning metabolic and exercise physiological parameters as well as the positive profile of side effects favor ACE inhibitors, long-acting calcium channel blockers and especially AT(1) antagonists in physically active hypertensive patients with concomitant metabolic syndrome.

[The effect of nimodipine on cochlear blood flow in the guinea pig].

PubMed

Meyer, P; Werner, E; Schmidt, R; Grützmacher, W; Gehrig, W; Seuter, F

1994-10-01

The influence of nimodipine (Nimotop, CAS 66085-59-4), a selectively cerebrovascularly acting 1,4-dihydropyridine calcium antagonist, on the cochlear blood flow (CBF) was studied in 19 guinea pigs (6 controls). The hydrogen clearance measurements were carried out under alpha-chloralose-ethylurethane anaesthesia, artificial respiration with simultaneous control of electrocardiogram, blood pressure, body temperature and arterial pH (hourly). The indirect measurement of CBF was carried out by means of hydrogen clearance in the perilymphatic space (basal turn) before and after intravenous application of 1 microgram nimodipine/kg/min. The mean arterial blood pressure remained within the +/- 5% range of the initial value during the experiment. Under treatment with nimodipine the CBF showed a non-significant average increase of 4.69% and under placebo (20% ethanol, 17% polyethylenglycol 400, citrate buffer), a non-significant average decrease of 6.16%. The influence of nimodipine on CBF was underlined by the overcompensation of the placebo effect.

A Hypothalamic Switch for REM and Non-REM Sleep.

PubMed

Chen, Kai-Siang; Xu, Min; Zhang, Zhe; Chang, Wei-Cheng; Gaj, Thomas; Schaffer, David V; Dan, Yang

2018-03-07

Rapid eye movement (REM) and non-REM (NREM) sleep are controlled by specific neuronal circuits. Here we show that galanin-expressing GABAergic neurons in the dorsomedial hypothalamus (DMH) comprise separate subpopulations with opposing effects on REM versus NREM sleep. Microendoscopic calcium imaging revealed diverse sleep-wake activity of DMH GABAergic neurons, but the galanin-expressing subset falls into two distinct groups, either selectively activated (REM-on) or suppressed (REM-off) during REM sleep. Retrogradely labeled, preoptic area (POA)-projecting galaninergic neurons are REM-off, whereas the raphe pallidus (RPA)-projecting neurons are primarily REM-on. Bidirectional optogenetic manipulations showed that the POA-projecting neurons promote NREM sleep and suppress REM sleep, while the RPA-projecting neurons have the opposite effects. Thus, REM/NREM switch is regulated antagonistically by DMH galaninergic neurons with intermingled cell bodies but distinct axon projections. Copyright © 2018 Elsevier Inc. All rights reserved.

Effect of water treatment sludge on growth and elemental composition of tomato (Lycopersicon esculentum) shoots

DOE Office of Scientific and Technical Information (OSTI.GOV)

Elliott, H.A.; Singer, L.M.

The impact of a water treatment sludge on the fertility of a silt loam soil was assessed by monitoring the yield and elemental composition of tomato (Lycopersicon esculentum) shoots in a greenhouse study. Application of sludge at rates from 2-10% (air dry weight basis) raised the soil pH from 5.3 to 8.0 which enhanced plant growth. A substantial reduction in metal (Cd, Zn, Cu, Ni) uptake was observed with sludge amendments, even at the highest rates. The alkaline nature of this sludge (pH=9.3, calcium carbonate equivalence=53%) suggest its potential use as a liming material for agricultural soils. Overly alkaline conditionsmore » should be avoided however, as high application rates combined with ammonia fertilization had an antagonistic effect on plant growth, possibly from P deficiency induced by struvite (MgNH{sub 4}PO{sub 4}) formation.« less

[Menopause: Hypertension and vascular disease].

PubMed

Zilberman, J M

Hypertension is the main cardiovascular risk factor affecting 25% of women. Hormone changes and hypertension after menopause may lead to higher target organ damage and cardiovascular disease such as increased arterial stiffness, coronary diseases, chronic heart failure and stroke. The physiopathological mechanisms involved in the development of hypertension and cardiovascular diseases in menopausal women are controversial. There are pharmacokinetic and pharmacodynamic differences in both sexes, the women have more coughing when using the converting-enzyme inhibitors, more cramps when using thiazide diuretics and more oedema in the inferior limbs when using calcium antagonists. The aim of this review is to analyse possible physiopathological mechanisms involved in hypertension after menopause and to gain a better understanding of the biological effects mediated by vascular ageing in women when the level of oestrogen protective effect decreases over the vascular system. Copyright © 2017 SEH-LELHA. Publicado por Elsevier España, S.L.U. All rights reserved.

Caffeine and adenosine.

PubMed

Ribeiro, Joaquim A; Sebastião, Ana M

2010-01-01

Caffeine causes most of its biological effects via antagonizing all types of adenosine receptors (ARs): A1, A2A, A3, and A2B and, as does adenosine, exerts effects on neurons and glial cells of all brain areas. In consequence, caffeine, when acting as an AR antagonist, is doing the opposite of activation of adenosine receptors due to removal of endogenous adenosinergic tonus. Besides AR antagonism, xanthines, including caffeine, have other biological actions: they inhibit phosphodiesterases (PDEs) (e.g., PDE1, PDE4, PDE5), promote calcium release from intracellular stores, and interfere with GABA-A receptors. Caffeine, through antagonism of ARs, affects brain functions such as sleep, cognition, learning, and memory, and modifies brain dysfunctions and diseases: Alzheimer's disease, Parkinson's disease, Huntington's disease, Epilepsy, Pain/Migraine, Depression, Schizophrenia. In conclusion, targeting approaches that involve ARs will enhance the possibilities to correct brain dysfunctions, via the universally consumed substance that is caffeine.

Nuclear envelope breakdown and mitosis in sand dollar embryos is inhibited by microinjection of calcium buffers in a calcium-reversible fashion, and by antagonists of intracellular Ca2+ channels.

PubMed

Silver, R B

1989-01-01

Transient elevations in intracellular free Ca2+ are believed to signal the initiation of mitosis. This model predicts that mitosis might be arrested prior to nuclear envelope breakdown (NEB) or anaphase onset if intracellular Ca2+ concentration is buffered or dampened. Microinjection of a discrete dose of Ca2+ into the cell might then release the cell to resume mitotic cycling. Experimentally, one blastomere of two cell sand dollar (Echinaracnius parma) embryos was microinjected with Ca2+ buffers, Ca2+ solutions, or Ca2+ channel antagonists; the uninjected blastomere was the control. Cells were loaded with 10 pl doses of the Ca2+ buffer antipyrylazo III (ApIII) at specific times in the cell cycle to attempt a competitive inhibition of Ca2+-dependent steps in NEB and initiation of mitosis. Injection of 50 microM ApIII 6 min prior to NEB blocked NEB and further cell cycling. Injections of solutions between 0 and 30 microM ApIII were without observable effect. Control injections had no observable effect on the injected cell. Cells injected with 50 microM ApIII 2 min prior to the onset of anaphase in control cells were blocked in metaphase. Cells were sensitive to Ca2+ buffer injections 6 min prior to NEB (with a 40- to 45-sec duration), and 2 min prior to anaphase onset (with a 10- to 20-sec duration). Vital staining of these cells with H33342 demonstrated that they contained only one nucleus that had the same fluorescence intensity as seen prior to microinjection, and thus did not undergo DNA synthesis following the imposition of the Ca2+ buffer block to mitosis. Cells arrested in this fashion did not spontaneously resume mitotic cycling. This Ca2+ buffer-induced mitotic arrest was, however, experimentally reversible. Cells arrested with 50 microM ApIII 6 min prior to NEB could be returned to mitotic activity by injecting 300 microM CaCl2 5 min after the ApIII injection. The double injected cells resumed cycling, NEB, and mitosis after a delay of one cell cycle period, and remained one cell cycle out of phase with the sister (control) cell. Microinjection of antagonists of endomembrane Ca2+ channels inhibited NEB and anaphase onset in a concentration- and time-dependent fashion. The effective doses of compounds tested were 7 micrograms/ml ryanodine and 500 micrograms/ml TMB-8. These results indicate that a transient elevation of intracellular Ca2+ from endomembrane stores is required to initiate mitotic events, namely NEB and anaphase onset.(ABSTRACT TRUNCATED AT 400 WORDS)

Copper-induced activation of TRP channels promotes extracellular calcium entry and activation of CaMK, PKA, PKC, PKG and CBLPK leading to increased expression of antioxidant enzymes in Ectocarpus siliculosus.

PubMed

González, Alberto; Sáez, Claudio A; Morales, Bernardo; Moenne, Alejandra

2018-05-01

The existence of functional Transient Receptor Potential (TRP) channels was analyzed in Ectocarpus siliculosus using agonists of human TRPs and specific antagonists of TRPA1, TRPC5, TRPM8 and TRPV; intracellular calcium was detected for 60 min. Increases in intracellular calcium were observed at 13, 29, 39 and 50-52 min, which appeared to be mediated by the activation of TRPM8/V1 at 13 min, TRPV1 at 29 min, TRPA1/V1 at 39 min and TRPA1/C5 at 50-52 min. In addition, intracellular calcium increases appear to be due to extracellular calcium entry, not requiring protein kinase activation. On the other hand, 2.5 μM copper exposure induced increased intracellular calcium at 13, 29, 39 and 51 min, likely due to the activation of a TRPA1/V1 at 13 min, TRPA1/C5/M8 at 29 min, TRPC5/M8 at 39 min, and a TRPC5/V1 at 51 min. The increases in intracellular calcium induced by copper were due to extracellular calcium entry and required protein kinase activation. Furthermore, from 3 to 24 h, copper exposure induced an increase in the level of transcripts encoding antioxidant enzymes such as superoxide dismutase, ascorbate peroxidase, glutathione reductase and peroxiredoxin. The described upregulation decreased with inhibitors of CaMK, PKA, PKC, PKG and CBLPK, as well as with a mixture of TRP inhibitors. Thus, copper induces the activation of TRP channels allowing extracellular calcium entry as well as the activation of CaMK, PKA, PKC, PKG and CBLPK leading to increased expression of genes encoding antioxidant enzymes in E. siliculosus. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Therapeutic perspectives in hypertension: novel means for renin-angiotensin-aldosterone system modulation and emerging device-based approaches.

PubMed

Unger, Thomas; Paulis, Ludovit; Sica, Domenic A

2011-11-01

The conventional antihypertensive therapies including renin-angiotensin-aldosterone system antagonists (converting enzyme inhibitors, receptor blockers, renin inhibitors, and mineralocorticoid receptor blockers), diuretics, β-blockers, and calcium channel blockers are variably successful in achieving the challenging target blood pressure values in hypertensive patients. Difficult to treat hypertension is still a commonly observed problem world-wide. A number of drugs are considered to be used as novel therapies for hypertension. Renalase supplementation, vasopeptidase inhibitors, endothelin antagonists, and especially aldosterone antagonists (aldosterone synthase inhibitors and novel selective mineralocorticoid receptor blockers) are considered an option in resistant hypertension. In addition, the aldosterone antagonists as well as (pro)renin receptor blockers or AT(2) receptor agonists might attenuate end-organ damage. This array of medications has now been complemented by a number of new approaches of non-pharmacological strategies including vaccination, genomic interference, controlled breathing, baroreflex activation, and probably most successfully renal denervation techniques. However, the progress on innovative therapies seems to be slow and the problem of resistant hypertension and proper blood pressure control appears to be still persisting. Therefore the regimens of currently available drugs are being fine-tuned, resulting in the establishment of several novel fixed-dose combinations including triple combinations with the aim to facilitate proper blood pressure control. It remains an exciting question which approach will confer the best blood pressure control and risk reduction in this tricky disease.

The role of calcium and calmodulin in freezing-induced freezing resistance of Populus tomentosa cuttings.

PubMed

Lin, Shan-Zhi; Zhang, Zhi-Yi; Lin, Yuan-Zhen; Zhang, Qian; Guo, Huan

2004-02-01

To explore the role of calcium-calmodulin messenger system in the transduction of low temperature signal in woody plants, Populus tomentosa cuttings after being treated with CaCl(2) (10 mmol/L), Ca(2+) chelator EGTA (3 mmol/L), Ca(2+) channel inhibitor LaCl(3) (100 mmol/L) or CaM antagonist CPZ (50 mmol/L) were used for freezing acclimation at -3 degrees C. The changes in the calmodulin (CaM) and malonaldehyde (MDA) contents, the activities of superoxide dismutase (SOD), peroxidase (POD) and Ca(2+)-dependent adenosinetriphosphatase (Ca(2+)-ATPase) of mitochondrial membrane as well as freezing resistance (expressed as LT(50)) of cuttings were investigated to elucidate the physiological mechanisms by which trees adapt to freezing. The results showed that freezing acclimation increased the CaM content, the activities of SOD, POD and Ca(2+)-ATPase of mitochondrial membrane as well as freezing resistance of cuttings, and decreased the MDA content as compared with control cuttings. Treatment with CaCl(2) at the time of freezing acclimation enhanced the effect of freezing acclimation on the above-mentioned indexes, but this enhancement was abolished by Ca(2+)chelator EGTA, Ca(2+) channel inhibitor LaCl(3) or CaM antagonist CPZ, indicating that the calcium-calmodulin messenger system was involved in the course of freezing resistance development. The presence of CaCl(2) at the same time of freezing acclimation also reduced the degree of decline in CaM content, and in SOD, POD and Ca(2+)-ATPase activities caused by freezing stress at -14 degrees C, and enhanced the level of increase in CaM content, and in SOD, POD and Ca(2+)-ATPase activity in the recovery periods at 25 degrees C . The change in CaM content was found to be closely correlated to the levels of SOD, POD and Ca(2+)-ATPase, and to the degree of freezing resistance of cuttings during freezing acclimation either with or without CaCl(2) treatment. It was suggested that the increase of CaM content induced by CaCl(2) treatment promote the formation of Ca(2+)-CaM complexes, which effectively activates the activities of SOD, POD and mitochondrial Ca(2+)-ATPase and then further result in the adaptive changes associated with the development and enhancement of freezing resistance. Thus, It could be concluded that Ca(2+)-calmodulin may be involved in the regulation of the increase in SOD, POD and Ca(2+)-ATPase activities, and the induction of freezing resistance of cuttings.

Contractile responses induced by physalaemin, an analogue of substance P, in the rat esophagus.

PubMed

Shiina, Takahiko; Shima, Takeshi; Hirayama, Haruko; Kuramoto, Hirofumi; Takewaki, Tadashi; Shimizu, Yasutake

2010-02-25

We examined the effects of physalaemin, an agonist of tachykinin receptors, on mechanical responses in the rat esophagus to clarify possible regulatory roles of tachykinins in esophageal motility. Exogenous application of physalaemin caused tonic contractions in rat esophageal segments when tension was recorded in the longitudinal direction but not when tension was recorded in the circular direction. The physalaemin-evoked contractions were blocked by pretreatment with nifedipine, a blocker of L-type calcium channels in both striated and smooth muscle cells. However, tetrodotoxin, a blocker of voltage-dependent sodium channels in striated muscle cells and neurons, did not affect the physalaemin-induced contractions. These results indicate that physalaemin might induce contractile responses in longitudinal smooth muscle of the muscularis mucosa via direct actions on muscle cells but not on neurons. Although pretreatment with a tachykinin NK(1) receptor antagonist, N-acetyl-l-tryptophan 3,5-bis (trifluoromethyl) benzyl ester (L-732,138), did not significantly affect the physalaemin-evoked contractions in rat esophageal segments, a tachykinin NK(2) receptor antagonist, (S)-N-methyl-N[4-(4-acetylamino-4-phenylpiperidino)-2-(3,4-dichlorophenyl) butyl] benzamide (SR48968), and a tachykinin NK(3) receptor antagonist, (S)-(N)-(1-(3-(1-benzoyl-3-(3,4-dichlorophenyl) piperidin-3-yl)propyl)-4-phenylpiperidin-4-yl)-N-methylacetamide (SR142801), significantly inhibited the physalaemin-evoked contractions. These results suggest that tachykinins can activate longitudinal contraction of smooth muscle in the muscularis mucosa, mediated via tachykinin NK(2) and NK(3) receptors on muscle cells, in the rat esophagus.

Pushing the threshold: How NMDAR antagonists induce homeostasis through protein synthesis to remedy depression.

PubMed

Raab-Graham, Kimberly F; Workman, Emily R; Namjoshi, Sanjeev; Niere, Farr

2016-09-15

Healthy neurons have an optimal operating range, coded globally by the frequency of action potentials or locally by calcium. The maintenance of this range is governed by homeostatic plasticity. Here, we discuss how new approaches to treat depression alter synaptic activity. These approaches induce the neuron to recruit homeostatic mechanisms to relieve depression. Homeostasis generally implies that the direction of activity necessary to restore the neuron's critical operating range is opposite in direction to its current activity pattern. Unconventional antidepressant therapies-deep brain stimulation and NMDAR antagonists-alter the neuron's "depressed" state by pushing the neuron's current activity in the same direction but to the extreme edge. These therapies rally the intrinsic drive of neurons in the opposite direction, thereby allowing the cell to return to baseline activity, form new synapses, and restore proper communication. In this review, we discuss seminal studies on protein synthesis dependent homeostatic plasticity and their contribution to our understanding of molecular mechanisms underlying the effectiveness of NMDAR antagonists as rapid antidepressants. Rapid antidepressant efficacy is likely to require a cascade of mRNA translational regulation. Emerging evidence suggests that changes in synaptic strength or intrinsic excitability converge on the same protein synthesis pathways, relieving depressive symptoms. Thus, we address the question: Are there multiple homeostatic mechanisms that induce the neuron and neuronal circuits to self-correct to regulate mood in vivo? Targeting alternative ways to induce homeostatic protein synthesis may provide, faster, safer, and longer lasting antidepressants. This article is part of a Special Issue entitled SI:RNA Metabolism in Disease. Published by Elsevier B.V.

Anticonvulsant and adverse effects of MK-801, LY 235959, and GYKI 52466 in combination with Ca2+ channel inhibitors in mice.

PubMed

Gasior, M; Borowicz, K; Kleinrok, Z; Starownik, R; Czuczwar, S J

1997-04-01

This study was designed to investigate the influence of the calcium (Ca2+) channel inhibitors nicardipine, nifedipine, and flunarizine on the protective action of MK-801, LY 235959 [N-methyl-D-aspartate (NMDA) receptor antagonists], and GYKI 52466 (a non-NMDA receptor antagonist) against electroconvulsions in mice. Unlike nicardipine (15 mg/kg) or flunarizine (10 mg/kg) nifedipine (7.5 and 15 mg/kg) potentiated the protective potency of MK-801 (0.05 mg/kg), as reflected by significant elevation of the convulsive threshold (a CS50 value of the current strength in mA producing tonic hind limb extension in 50% of the animals). The protective activity of LY 235959 and GYKI 52466 was reflected by their ED50 values in mg/kg, at which the drugs were expected to protect 50% of mice against maximal electroshock-induced tonic extension of the hind limbs. Nicardipine (3.75 15 mg/kg), nifedipine (0.94-15 mg/kg), and flunarizine (2.5-10 mg/kg) in a dose-dependent manner markedly potentiated the antiseizure efficacy of LY 235959. Flunarizine (5 and 10 mg/kg) was the only Ca2+ channel inhibitor to enhance the protective action of GYKI 52466 against electroconvulsions. Except with MK-801 + flunarizine (motor performance) or GYKI 52466 + flunarizine (long-term memory), combination of NMDA or non-NMDA receptor antagonists with Ca2+ channel inhibitors produced an impairment of motor performance (evaluated in the chimney test) and long-term memory acquisition (measured in the passive avoidance task) as compared with vehicle treatment.

Block of high-threshold calcium channels by the synthetic polyamines sFTX-3.3 and FTX-3.3.

PubMed

Norris, T M; Moya, E; Blagbrough, I S; Adams, M E

1996-10-01

A polyamine component of Agelenopsis aperta spider venom designated FTX is reported to be a selective antagonist of P-type calcium channels in the mammalian brain. Consequently, this component has frequently been used as a pharmacological tool to determine the presence, distribution, and function of P-type channels in physiological systems. We describe antagonism of calcium channels by the synthesized polyamine FTX-3.3, which has the proposed structure of natural FTX. We also examined a corresponding polyamine amide, sFTX-3.3. These polyamines are critically evaluated for antagonism of three high-threshold calcium channel subtypes in rat neurons through the use of the whole-cell patch-clamp technique. FTX-3.3 (IC50 = approximately 0.13 mM) is approximately twice as potent as sFTX-3.3 (IC50 = approximately 0.24 mM) against P-type channels and approximately 3-fold more potent against N-type channels (FTX-3.3, IC50 = approximately 0.24 mM; sFTX-3.3, IC50 = approximately 0.70 mM). Both polyamines also block L-type calcium channels with similar potencies. sFTX-3.3 (1 mM) and FTX-3.3 (0.5 mM) typically block 50% and 65% of Bay K8644-enhanced L-type current, respectively. Antagonism of each calcium channel subtype is voltage dependent, with less inhibition of Ba2+ currents at more-positive potentials. These data show that both sFTX-3.3 and FTX-3.3 antagonize P-, N-, and L-type calcium channels in mammalian Purkinje and superior cervical ganglia neurons with similar IC50 values.

TRPV2 activation induces apoptotic cell death in human T24 bladder cancer cells: a potential therapeutic target for bladder cancer.

PubMed

Yamada, Takahiro; Ueda, Takashi; Shibata, Yasuhiro; Ikegami, Yosuke; Saito, Masaki; Ishida, Yusuke; Ugawa, Shinya; Kohri, Kenjiro; Shimada, Shoichi

2010-08-01

To investigate the functional expression of the transient receptor potential vanilloid 2 (TRPV2) channel protein in human urothelial carcinoma (UC) cells and to determine whether calcium influx into UC cells through TRPV2 is involved in apoptotic cell death. The expression of TRPV2 mRNA in bladder cancer cell lines (T24, a poorly differentiated UC cell line and RT4, a well-differentiated UC cell line) was analyzed using reverse transcriptase-polymerase chain reaction. The calcium permeability of TRPV2 channels in T24 cells was investigated using a calcium imaging assay that used cannabidiol (CBD), a relatively selective TRPV2 agonist, and ruthenium red (RuR), a nonselective TRPV channel antagonist. The death of T24 or RT4 cells in the presence of CBD was evaluated using a cellular viability assay. Apoptosis of T24 cells caused by CBD was confirmed using an annexin-V assay and small interfering RNA (siRNA) silencing of TRPV2. TRPV2 mRNA was abundantly expressed in T24 cells. The expression level in UC cells was correlated with high-grade disease. The administration of CBD increased intracellular calcium concentrations in T24 cells. In addition, the viability of T24 cells progressively decreased with increasing concentrations of CBD, whereas RT4 cells were mostly unaffected. Cell death occurred via apoptosis caused by continuous influx of calcium through TRPV2. TRPV2 channels in UC cells are calcium-permeable and the regulation of calcium influx through these channels leads directly to the death of UC cells. TRPV2 channels in UC cells may be a potential new therapeutic target, especially in higher-grade UC cells. Copyright 2010 Elsevier Inc. All rights reserved.

[Effects of flunarizine and vitamin C on hemodynamics in rat heart subjected to ischemia-reperfusion].

PubMed

Xian, Y; Lan, T; Wang, Y

1998-09-01

Langendorff perfusion isolated rat heart was subjected to total global ischemia (coronary flow rate is equal to zero) for 10 minutes and reperfusion for 15 minutes. The heart rate (HR), left ventricular developed pressure (LVDP), coronary flow rate (CFR), electrocardiogram (ECG) and the effects of calcium antagonist-flunarizine (FNZ) and/or oxygen free radical scavenger--vitamine C on the above parameters were observed. The results showed that FNZ dilated coronary vessel (P < 0.01) and had a slight negative chronotropic effect, but it had no effect on LVP. Vitamine C did not affect HR, LVP and CFR. The recovery of the product of HR and LVDP-Rate Pressure Product (RPP) in the FNZ + Vit. C group, Vit. C group and FNZ group was significantly higher than that in the control group (P < 0.05) at ten minutes reperfusion. All the results suggest that FNZ and Vit. C may improve the recovery of heart function after reperfusion.

Partial purification and characterization of a Ca(2+)-dependent protein kinase from the green alga, Dunaliella salina

NASA Technical Reports Server (NTRS)

Roux, S. J.

1990-01-01

A calcium-dependent protein kinase was partially purified and characterized from the green alga Dunaliella salina. The enzyme was activated at free Ca2+ concentrations above 10(-7) molar. and half-maximal activation was at about 3 x 10(-7) molar. The optimum pH for its Ca(2+)-dependent activity was 7.5. The addition of various phospholipids and diolein had no effects on enzyme activity and did not alter the sensitivity of the enzyme toward Ca2+. The enzyme was inhibited by calmodulin antagonists, N-(6-aminohexyl)-1-naphthalene sulfonamide and N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide in a dose-dependent manner while the protein kinase C inhibitor, sphingosine, had little effect on enzyme activity up to 800 micromolar. Immunoassay showed some calmodulin was present in the kinase preparations. However, it is unlikely the kinase was calmodulin regulated, since it still showed stimulation by Ca2+ in gel assays after being electrophoretically separated from calmodulin by two different methods. This gel method of detection of the enzyme indicated that a protein band with an apparent molecular weight of 40,000 showed protein kinase activity at each one of the several steps in the purification procedure. Gel assay analysis also showed that after native gel isoelectric focusing the partially purified kinase preparations had two bands with calcium-dependent activity, at isoelectric points 6.7 and 7.1. By molecular weight, by isoelectric point, and by a comparative immunoassay, the Dunaliella kinase appears to differ from at least some of the calcium-dependent, but calmodulin and phospholipid independent kinases described from higher plants.

Endomorphin-2 is Released from Newborn Rat Primary Sensory Neurons in a Frequency- and Calcium- Dependent Manner

PubMed Central

Scanlin, Heather L.; Carroll, Elizabeth A.; Jenkins, Victoria K.; Balkowiec, Agnieszka

2008-01-01

Recent evidence indicates that endomorphins, endogenous mu-opioid receptor (MOR) agonists, modulate synaptic transmission in both somatic and visceral sensory pathways. Here we show that endomorphin-2 (END-2) is expressed in newborn rat dorsal root ganglion (DRG) and nodose-petrosal ganglion complex (NPG) neurons, and rarely co-localizes with brain-derived neurotrophic factor (BDNF). In order to examine activity-dependent release of END-2 from neurons, we established a model using dispersed cultures of DRG and NPG cells activated by patterned electrical field stimulation. To detect release of END-2, we developed a novel rapid capture ELISA, in which END-2 capture antibody was added to neuronal cultures shortly before their electrical stimulation. The conventional assay was effective at reliably detecting END-2 only when the cells were stimulated in the presence of CTAP, a MOR-selective antagonist. This suggests that the strength of the novel assay is related primarily to rapid capture of released END-2 before it binds to endogenous MORs. Using the rapid capture ELISA, we found that stimulation protocols known to induce plastic changes at sensory synapses were highly effective at releasing END-2. Removal of extracellular calcium or blocking voltage-activated calcium channels significantly reduced the release. Together, our data provide the first evidence that END-2 is expressed by newborn DRG neurons of all sizes found in this age group, and can be released from these, as well as from NPG neurons, in an activity-dependent manner. These results point to END-2 as a likely mediator of activity-dependent plasticity in sensory pathways. PMID:18513316

Prostaglandin E2 Prevents Hyperosmolar-Induced Human Mast Cell Activation through Prostanoid Receptors EP2 and EP4

PubMed Central

Torres-Atencio, Ivonne; Ainsua-Enrich, Erola; de Mora, Fernando; Picado, César; Martín, Margarita

2014-01-01

Background Mast cells play a critical role in allergic and inflammatory diseases, including exercise-induced bronchoconstriction (EIB) in asthma. The mechanism underlying EIB is probably related to increased airway fluid osmolarity that activates mast cells to the release inflammatory mediators. These mediators then act on bronchial smooth muscle to cause bronchoconstriction. In parallel, protective substances such as prostaglandin E2 (PGE2) are probably also released and could explain the refractory period observed in patients with EIB. Objective This study aimed to evaluate the protective effect of PGE2 on osmotically activated mast cells, as a model of exercise-induced bronchoconstriction. Methods We used LAD2, HMC-1, CD34-positive, and human lung mast cell lines. Cells underwent a mannitol challenge, and the effects of PGE2 and prostanoid receptor (EP) antagonists for EP1–4 were assayed on the activated mast cells. Beta-hexosaminidase release, protein phosphorylation, and calcium mobilization were assessed. Results Mannitol both induced mast cell degranulation and activated phosphatidyl inositide 3-kinase and mitogen-activated protein kinase (MAPK) pathways, thereby causing de novo eicosanoid and cytokine synthesis. The addition of PGE2 significantly reduced mannitol-induced degranulation through EP2 and EP4 receptors, as measured by beta-hexosaminidase release, and consequently calcium influx. Extracellular-signal-regulated kinase 1/2, c-Jun N-terminal kinase, and p38 phosphorylation were diminished when compared with mannitol activation alone. Conclusions Our data show a protective role for the PGE2 receptors EP2 and EP4 following osmotic changes, through the reduction of human mast cell activity caused by calcium influx impairment and MAP kinase inhibition. PMID:25329458

Glutamate Receptor-Like Channel3.3 Is Involved in Mediating Glutathione-Triggered Cytosolic Calcium Transients, Transcriptional Changes, and Innate Immunity Responses in Arabidopsis1[W][OA

PubMed Central

Li, Feng; Wang, Jing; Ma, Chunli; Zhao, Yongxiu; Wang, Yingchun; Hasi, Agula; Qi, Zhi

2013-01-01

The tripeptide reduced glutathione (GSH; γ-glutamate [Glu]-cysteine [Cys]-glycine) is a major endogenous antioxidant in both animal and plant cells. It also functions as a neurotransmitter mediating communication among neurons in the central nervous system of animals through modulating specific ionotropic Glu receptors (GLRs) in the membrane. Little is known about such signaling roles in plant cells. Here, we report that transient rises in cytosolic calcium triggered by exogenous GSH in Arabidopsis (Arabidopsis thaliana) leaves were sensitive to GLR antagonists and abolished in loss-of-function atglr3.3 mutants. Like the GSH biosynthesis-defective mutant PHYTOALEXIN DEFICIENT2, atglr3.3 showed enhanced susceptibility to the bacterial pathogen Pseudomonas syringae pv tomato DC3000. Pathogen-induced defense marker gene expression was also decreased in atglr3.3 mutants. Twenty-seven percent of genes that were rapidly responsive to GSH treatment of seedlings were defense genes, most of which were dependent on functional AtGLR3.3, while GSH suppressed pathogen propagation through the AtGLR3.3-dependent pathway. Eight previously identified putative AtGLR3.3 ligands, GSH, oxidized glutathione, alanine, asparagine, Cys, Glu, glycine, and serine, all elicited the AtGLR3.3-dependent cytosolic calcium transients, but only GSH and Cys induced the defense response, with the Glu-induced AtGLR3.3-dependent transcription response being much less apparent than that triggered by GSH. Together, these observations suggest that AtGLR3.3 is required for several signaling effects mediated by extracellular GSH, even though these effects may not be causally related. PMID:23656893

Modification of hippocampal markers of synaptic plasticity by memantine in animal models of acute and repeated restraint stress: implications for memory and behavior.

PubMed

Amin, Shaimaa Nasr; El-Aidi, Ahmed Amro; Ali, Mohamed Mostafa; Attia, Yasser Mahmoud; Rashed, Laila Ahmed

2015-06-01

Stress is any condition that impairs the balance of the organism physiologically or psychologically. The response to stress involves several neurohormonal consequences. Glutamate is the primary excitatory neurotransmitter in the central nervous system, and its release is increased by stress that predisposes to excitotoxicity in the brain. Memantine is an uncompetitive N-methyl D-aspartate glutamatergic receptors antagonist and has shown beneficial effect on cognitive function especially in Alzheimer's disease. The aim of the work was to investigate memantine effect on memory and behavior in animal models of acute and repeated restraint stress with the evaluation of serum markers of stress and the expression of hippocampal markers of synaptic plasticity. Forty-two male rats were divided into seven groups (six rats/group): control, acute restraint stress, acute restraint stress with Memantine, repeated restraint stress, repeated restraint stress with Memantine and Memantine groups (two subgroups as positive control). Spatial working memory and behavior were assessed by performance in Y-maze. We evaluated serum cortisol, tumor necrotic factor, interleukin-6 and hippocampal expression of brain-derived neurotrophic factor, synaptophysin and calcium-/calmodulin-dependent protein kinase II. Our results revealed that Memantine improved spatial working memory in repeated stress, decreased serum level of stress markers and modified the hippocampal synaptic plasticity markers in both patterns of stress exposure; in ARS, Memantine upregulated the expression of synaptophysin and brain-derived neurotrophic factor and downregulated the expression of calcium-/calmodulin-dependent protein kinase II, and in repeated restraint stress, it upregulated the expression of synaptophysin and downregulated calcium-/calmodulin-dependent protein kinase II expression.

Contribution of TRPC3 to store-operated calcium entry and inflammatory transductions in primary nociceptors

PubMed Central

2014-01-01

Background Prolonged intracellular calcium elevation contributes to sensitization of nociceptors and chronic pain in inflammatory conditions. The underlying molecular mechanisms remain unknown but store-operated calcium entry (SOCE) components participate in calcium homeostasis, potentially playing a significant role in chronic pain pathologies. Most G protein-coupled receptors activated by inflammatory mediators trigger calcium-dependent signaling pathways and stimulate SOCE in primary afferents. The aim of the present study was to investigate the role of TRPC3, a calcium-permeable non-selective cation channel coupled to phospholipase C and highly expressed in DRG, as a link between activation of pro-inflammatory metabotropic receptors and SOCE in nociceptive pathways. Results Using in situ hybridization, we determined that TRPC3 and TRPC1 constitute the major TRPC subunits expressed in adult rat DRG. TRPC3 was found localized exclusively in small and medium diameter sensory neurons. Heterologous overexpression of TRPC3 channel subunits in cultured primary DRG neurons evoked a significant increase of Gd3+-sensitive SOCE following thapsigargin-induced calcium store depletion. Conversely, using the same calcium add-back protocol, knockdown of endogenous TRPC3 with shRNA-mediated interference or pharmacological inhibition with the selective TRPC3 antagonist Pyr10 induced a substantial decrease of SOCE, indicating a significant role of TRPC3 in SOCE in DRG nociceptors. Activation of P2Y2 purinoceptors or PAR2 protease receptors triggered a strong increase in intracellular calcium in conditions of TRPC3 overexpression. Additionally, knockdown of native TRPC3 or its selective pharmacological blockade suppressed UTP- or PAR2 agonist-evoked calcium responses as well as sensitization of DRG neurons. These data show a robust link between activation of pro-inflammatory receptors and calcium homeostasis through TRPC3-containing channels operating both in receptor- and store-operated mode. Conclusions Our findings highlight a major contribution of TRPC3 to neuronal calcium homeostasis in somatosensory pathways based on the unique ability of these cation channels to engage in both SOCE and receptor-operated calcium influx. This is the first evidence for TRPC3 as a SOCE component in DRG neurons. The flexible role of TRPC3 in calcium signaling as well as its functional coupling to pro-inflammatory metabotropic receptors involved in peripheral sensitization makes it a potential target for therapeutic strategies in chronic pain conditions. PMID:24965271

Carbachol-mediated pigment granule dispersion in retinal pigment epithelium requires Ca2+ and calcineurin.

PubMed

Johnson, Adam S; García, Dana M

2007-12-19

Inside bluegill (Lepomis macrochirus) retinal pigment epithelial cells, pigment granules move in response to extracellular signals. During the process of aggregation, pigment motility is directed toward the cell nucleus; in dispersion, pigment is directed away from the nucleus and into long apical processes. A number of different chemicals have been found to initiate dispersion, and carbachol (an acetylcholine analog) is one example. Previous research indicates that the carbachol-receptor interaction activates a Gq-mediated pathway which is commonly linked to Ca2+ mobilization. The purpose of the present study was to test for involvement of calcium and to probe calcium-dependent mediators to reveal their role in carbachol-mediated dispersion. Carbachol-induced pigment granule dispersion was blocked by the calcium chelator BAPTA. In contrast, the calcium channel antagonist verapamil, and incubation in Ca2+-free medium failed to block carbachol-induced dispersion. The calcineurin inhibitor cypermethrin blocked carbachol-induced dispersion; whereas, two protein kinase C inhibitors (staurosporine and bisindolylmaleimide II) failed to block carbachol-induced dispersion, and the protein kinase C activator phorbol 12-myristate 13-acetate failed to elicit dispersion. A rise in intracellular calcium is necessary for carbachol-induced dispersion; however, the Ca2+ requirement is not dependent on extracellular sources, implying that intracellular stores are sufficient to enable pigment granule dispersion to occur. Calcineurin is a likely Ca2+-dependent mediator involved in the signal cascade. Although the pathway leads to the generation of diacylglycerol and calcium (both required for the activation of certain PKC isoforms), our evidence does not support a significant role for PKC.

Carbachol-mediated pigment granule dispersion in retinal pigment epithelium requires Ca2+ and calcineurin

PubMed Central

Johnson, Adam S; García, Dana M

2007-01-01

Background Inside bluegill (Lepomis macrochirus) retinal pigment epithelial cells, pigment granules move in response to extracellular signals. During the process of aggregation, pigment motility is directed toward the cell nucleus; in dispersion, pigment is directed away from the nucleus and into long apical processes. A number of different chemicals have been found to initiate dispersion, and carbachol (an acetylcholine analog) is one example. Previous research indicates that the carbachol-receptor interaction activates a Gq-mediated pathway which is commonly linked to Ca2+ mobilization. The purpose of the present study was to test for involvement of calcium and to probe calcium-dependent mediators to reveal their role in carbachol-mediated dispersion. Results Carbachol-induced pigment granule dispersion was blocked by the calcium chelator BAPTA. In contrast, the calcium channel antagonist verapamil, and incubation in Ca2+-free medium failed to block carbachol-induced dispersion. The calcineurin inhibitor cypermethrin blocked carbachol-induced dispersion; whereas, two protein kinase C inhibitors (staurosporine and bisindolylmaleimide II) failed to block carbachol-induced dispersion, and the protein kinase C activator phorbol 12-myristate 13-acetate failed to elicit dispersion. Conclusion A rise in intracellular calcium is necessary for carbachol-induced dispersion; however, the Ca2+ requirement is not dependent on extracellular sources, implying that intracellular stores are sufficient to enable pigment granule dispersion to occur. Calcineurin is a likely Ca2+-dependent mediator involved in the signal cascade. Although the pathway leads to the generation of diacylglycerol and calcium (both required for the activation of certain PKC isoforms), our evidence does not support a significant role for PKC. PMID:18093324

Protection against cyanide-induced convulsions with alpha-ketoglutarate.

PubMed

Yamamoto, H

1990-04-30

Protection against convulsions induced by cyanide was observed after treatment with alpha-ketoglutarate, either alone or in combination with sodium thiosulfate, a classical antagonist for cyanide intoxication. However, sodium thiosulfate alone did not protect against cyanide (30 mg/kg)-induced convulsions. gamma-Aminobutyric acid (GABA) levels in brain were decreased by 31% in KCN-treated mice exhibiting convulsions. The combined administration of alpha-ketoglutarate and sodium thiosulfate completely abolished the decrease of GABA levels induced by cyanide. Furthermore, sodium thiosulfate alone also completely abolished the decrease of GABA levels. These results suggest that the depletion of brain GABA levels may not directly contribute to the development of convulsions induced by cyanide. On the other hand, cyanide increased calcium levels by 32% in brain crude mitochondrial fractions in mice with convulsions. The increased calcium levels were completely abolished by the combined administration of alpha-ketoglutarate and sodium thiosulfate, but not affected by sodium thiosulfate alone. These findings support the hypothesis proposed by Johnson et al. (Toxicol. Appl. Pharmacol., 84 (1986) 464) and Robinson et al. (Toxicology, 35 (1985) 59) that calcium may play an important role in mediating cyanide neurotoxicity.

Regulation of axonal and dendritic growth by the extracellular calcium-sensing receptor (CaSR)

PubMed Central

Vizard, Thomas N.; O'Keeffe, Gerard W.; Gutierrez, Humberto; Kos, Claudine H.; Riccardi, Daniela; Davies, Alun M.

2009-01-01

The extracellular calcium-sensing receptor (CaSR) monitors the systemic extracellular free ionized calcium level ([Ca2+]o) in organs involved in systemic [Ca2+]o homeostasis. However, the CaSR is also expressed in the nervous system where its role is unknown. Here we find high levels of the CaSR in perinatal mouse sympathetic neurons when their axons are innervating and branching extensively in their targets. Manipulating CaSR function in these neurons by varying [Ca2+]o, using CaSR agonists and antagonists or expressing a dominant-negative CaSR markedly affects neurite growth in vitro Sympathetic neurons lacking the CaSR have smaller neurite arbors in vitro, and sympathetic innervation density is reduced in CaSR-deficient mice in vivo. Hippocampal pyramidal neurons, which also express the CaSR, have smaller dendrites when transfected with dominant-negative CaSR in postnatal organotypic cultures. Our findings reveal a crucial role for the CaSR in regulating the growth of neural processes in the peripheral and central nervous systems. PMID:18223649

Interaction of SR 33557 with skeletal muscle calcium channel blocker receptors in the baboon: characterization of its binding sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sol-Rolland, J.; Joseph, M.; Rinaldi-Carmona, M.

1991-05-01

A procedure for the isolation of primate skeletal microsomal membranes was initiated. Membranes exhibited specific enzymatic markers such as 5'-nucleotidase, Ca{sup 2}{sup +},Mg({sup 2}{sup +})-adenosine triphosphatase and an ATP-dependent calcium uptake. Baboon skeletal microsomes bound specifically with high-affinity potent Ca{sup 2}{sup +} channel blockers such as dihydropyridine, phenylalkylamine and benzothiazepine derivatives. Scatchard analysis of equilibrium binding assays with ({sup 3}H)(+)-PN 200-110, ({sup 3}H)(-)-desmethoxyverapamil (( {sup 3}H)(-)-D888) and ({sup 3}H)-d-cis-dilitiazem were consistent with a single class of binding sites for the three radioligands. The pharmacological profile of SR 33557, an original compound with calcium antagonist properties, was investigated using radioligand bindingmore » studies. SR 33557 totally inhibited the specific binding of the three main classes of Ca{sup 2}{sup +} channel effectors and interacted allosterically with them. In addition, SR 33557 bound with high affinity to a homogeneous population of binding sites in baboon skeletal muscle.« less

TMEM16A inhibition impedes capacitation and acquisition of hyperactivated motility in guinea pig sperm.

PubMed

Cordero-Martínez, Joaquín; Reyes-Miguel, Tania; Rodríguez-Páez, Lorena; Garduño-Siciliano, Leticia; Maldonado-García, Deneb; Roa-Espitia, Ana L; Hernández-González, Enrique O

2018-07-01

Ca 2+ -activated Cl - channels (CaCCs) are anionic channels that regulate many important physiological functions associated with chloride and calcium flux in some somatic cells. The molecular identity of CaCCs was revealed to be TMEM16A and TMEM16B (also known as Anoctamin or ANO1 and ANO2, respectively) in all eukaryotes. A recent study suggests the presence of TMEM16A in human sperm and a relationship with the rhZP-induced acrosome reaction. However, to the best of our knowledge, little is known about the role of TMEM16A in other spermatic processes such as capacitation or motility. In this study, we evaluated the effects of two TMEM16A antagonists on capacitation, acrosome reaction, and motility in guinea pig sperm; these antagonists were T16Ainh-A01, belonging to a second generation of potent antagonists of TMEM16A, and niflumic acid (NFA), a well-known antagonist of TMEM16A (CaCCs). First of all, we confirmed that the absence of Cl - in the capacitation medium changes motility parameters, capacitation, and the progesterone-induced acrosome reaction. Using a specific antibody, TMEM16A was found as a protein band of ∼120 kDa, which localization was in the apical crest of the acrosome and the middle piece of the flagellum. Inhibition of TMEM16A by T16Ainh-A01 affected sperm physiology by reducing capacitation, blocking the progesterone-induced acrosome reaction under optimal capacitation conditions, inhibiting progressive motility, and the acquisition of hyperactivated motility, diminishing [Ca 2+ ]i, and increasing [Cl - ]i. These changes in sperm kinematic parameters provide new evidence of the important role played by TMEM16A in the production of sperm capable of fertilizing oocytes. © 2018 Wiley Periodicals, Inc.

Acylated and unacylated ghrelin confer neuroprotection to mesencephalic neurons.

PubMed

Wagner, Johanna; Vulinović, Franca; Grünewald, Anne; Unger, Marcus M; Möller, Jens C; Klein, Christine; Michel, Patrick P; Ries, Vincent; Oertel, Wolfgang H; Alvarez-Fischer, Daniel

2017-12-04

The polypeptide ghrelin is an endogenous ligand at the growth hormone secretagogue receptor 1a. To ghrelin multiple functions have been ascribed including promotion of gastrointestinal motility. Postprandial ghrelin levels have been reported to be reduced in patients suffering from Parkinson disease (PD). Experimental studies revealed neuroprotective effects of ghrelin in different PD models. The purpose of the present study was (i) to further elucidate the mechanism underlying the neuroprotective action of ghrelin and (ii) to determine whether these effects occur with both the acylated and the unacylated form. The study was conducted in primary mesencephalic cultures treated with mitochondrial complex I and complex II inhibitors. We show that protective effects of ghrelin against complex I inhibition with MPP + were independent of the acylation status of ghrelin, although acylated ghrelin appeared to be more potent. Protection by both forms was also observed when neurons were exposed to the complex II inhibitor 3-NP. Both forms led to higher oxygen consumption rates upon electron transport chain uncoupling, indicating that the two peptides may exert uncoupling effects themselves. We demonstrate that the rescue provided by ghrelin required calcium influx through L-type voltage-gated calcium channels. Whereas the protective effects of acylated ghrelin required receptor binding, effects of the unacylated form remained unaffected by treatment with a ghrelin receptor antagonist. Importantly, inhibition of ghrelin O-acyltransferase failed to reduce the activity of unacylated ghrelin. Overall, our data suggest that both acylated and unacylated ghrelin afford protection to dopamine neurons but through mechanisms that only partially overlap. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

Glutamate receptors modulate sodium-dependent and calcium-independent vitamin C bidirectional transport in cultured avian retinal cells.

PubMed

Portugal, Camila Cabral; Miya, Vivian Sayuri; Calaza, Karin da Costa; Santos, Rochelle Alberto Martins; Paes-de-Carvalho, Roberto

2009-01-01

Vitamin C is transported in the brain by sodium vitamin C co-transporter 2 (SVCT-2) for ascorbate and glucose transporters for dehydroascorbate. Here we have studied the expression of SVCT-2 and the uptake and release of [(14)C] ascorbate in chick retinal cells. SVCT-2 immunoreactivity was detected in rat and chick retina, specially in amacrine cells and in cells in the ganglion cell layer. Accordingly, SVCT-2 was expressed in cultured retinal neurons, but not in glial cells. [(14)C] ascorbate uptake was saturable and inhibited by sulfinpyrazone or sodium-free medium, but not by treatments that inhibit dehydroascorbate transport. Glutamate-stimulated vitamin C release was not inhibited by the glutamate transport inhibitor l-beta-threo-benzylaspartate, indicating that vitamin C release was not mediated by glutamate uptake. Also, ascorbate had no effect on [(3)H] D-aspartate release, ruling out a glutamate/ascorbate exchange mechanism. 2-Carboxy-3-carboxymethyl-4-isopropenylpyrrolidine (Kainate) or NMDA stimulated the release, effects blocked by their respective antagonists 6,7-initroquinoxaline-2,3-dione (DNQX) or (5R,2S)-(1)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801). However, DNQX, but not MK-801 or 2-amino-5-phosphonopentanoic acid (APV), blocked the stimulation by glutamate. Interestingly, DNQX prevented the stimulation by NMDA, suggesting that the effect of NMDA was mediated by glutamate release and stimulation of non-NMDA receptors. The effect of glutamate was neither dependent on external calcium nor inhibited by 1,2-bis (2-aminophenoxy) ethane-N',N',N',N',-tetraacetic acid tetrakis (acetoxy-methyl ester) (BAPTA-AM), an internal calcium chelator, but was inhibited by sulfinpyrazone or by the absence of sodium. In conclusion, retinal cells take up and release vitamin C, probably through SVCT-2, and the release can be stimulated by NMDA or non-NMDA glutamate receptors.

Mother root of Aconitum carmichaelii Debeaux exerts antinociceptive effect in Complet Freund's Adjuvant-induced mice: roles of dynorpin/kappa-opioid system and transient receptor potential vanilloid type-1 ion channel.

PubMed

Wang, Chao; Sun, Danni; Liu, Chunfang; Zhu, Chunyan; Jing, Xianghong; Chen, Shuping; Liu, Cuiling; Zhi, Kai; Xu, Tengfei; Wang, Hui; Liu, Junling; Xu, Ying; Liu, Zhiqiang; Lin, Na

2015-08-30

Processed Chuanwu (PCW), the mother root of Aconitum carmichaelii Debeauxv, has been widely used as a classic Traditional Chinese Medicine for pain relieve for over two millennia clinically. However, its action on chronic inflammatory pain has not been clarified. Here, we investigated the antinociceptive effect of PCW in complete freund's adjuvant (CFA)-induced mice and its possible mechanisms associated with opioid system and TRPV1 ion channel. Male ICR mice were intraplantarly injected with CFA. PCW (0.34, 0.68 and 1.35 g/kg) was orally given to mice once a day for 7 days. Von frey hairs and planter test were assessed to evaluate the antinociceptive effect of PCW. To investigate the participation of dynorphin/opioid system in PCW antinociception, subtype-specific opioid receptor antagonists or anti-dynorphin A antiserum were used. To eliminate other central mechanisms that contribute to PCW antinociception, hot plate (50 °C) test were performed. Further, involvements of TRPV1 in PCW antinociception were evaluated in CFA-induced TRPV1(-/-) and TRPV1(+/+) C57BL/6 male mice, and in capsaicin-induced nociception ICR naive mice pretreated with nor-BNI. Meanwhile, calcium imaging was performed in HEK293T-TRPV1 cells. Finally, rotarod, open-field tests and body temperature measurement were carried out to assess side effects of PCW. PCW dose-dependently attenuated mechanical and heat hypersensitivities with no tolerance, which could be partially attenuated by coadministration of k-opioid receptor antagonist nor-binaltorphimine (nor-BNI) or anti-dynorphin A (1-13) antiserum. And PCW antinociception was totally erased by pretreatment with nor-BNI in the hot plate test. In addition, PCW antinociception was decreased in TRPV1(-/-) mice compared to TRPV1(+/+) group. And PCW still manifested inhibitory effects in capsaicin-induced nociception with nor-BNI pretreatment. PCW significantly inhibited capsaicin-induced calcium influx in HEK293T-TRPV1 cells. Finally, no detectable side effects were found in naive mice treated with PCW. This study shows PCW's potent antinociceptive effect in inflammatory conditions without obvious side effects. This effect may result from the activation of κ-opioid receptor via dynorpin release and the inhibition of TRPV1. These findings indicate that PCW might be a potential agent for the management of chronic inflammatory pain.

Guanidine-acylguanidine bioisosteric approach in the design of radioligands: synthesis of a tritium-labeled N(G)-propionylargininamide ([3H]-UR-MK114) as a highly potent and selective neuropeptide Y Y1 receptor antagonist.

PubMed

Keller, Max; Pop, Nathalie; Hutzler, Christoph; Beck-Sickinger, Annette G; Bernhardt, Günther; Buschauer, Armin

2008-12-25

Synthesis and characterization of (R)-N(alpha)-(2,2-diphenylacetyl)-N-(4-hydroxybenzyl)-N(omega)-([2,3-(3)H]-propanoyl)argininamide ([(3)H]-UR-MK114), an easily accessible tritium-labeled NPY Y(1) receptor (Y(1)R) antagonist (K(B): 0.8 nM, calcium assay, HEL cells) derived from the (R)-argininamide BIBP 3226, is reported. The radioligand binds with high affinity (K(D), saturation: 1.2 nM, kinetic experiments: 1.1 nM, SK-N-MC cells) and selectivity for Y(1)R over Y(2), Y(4), and Y(5) receptors. The title compound is a useful pharmacological tool for the determination of Y(1)R ligand affinities, quantification of Y(1)R binding sites, and autoradiography.

Current statins show calcium channel blocking activity through voltage gated channels.

PubMed

Ali, Niaz; Begum, Robina; Faisal, Muhammad Saleh; Khan, Aslam; Nabi, Muhammad; Shehzadi, Gulfam; Ullah, Shakir; Ali, Waqar

2016-09-21

Statins are used for treatment of hypercholestremia. Common adverse reports associated with use of statins are generalized bodyache, rhabdomyolysis, muscles weakness and gastrointestinal disorders. The current work is an attempt to explain how smooth muscles of gastrointestinal tissues are affected by the current statins (Simvastatin, atorvastatin, fluvastatin and rosuvastatin). Effects of the current statins were studied on spontaneous activity of isolated rabbits' jejunal preparations. Different molar concentrations (10(-12)-10(-2)M) of the statins were applied on spontaneously contracting rabbits' jejunal preparations. As statins relaxed spontaneous activity, so we tested the statins on KCl (80 mM) induced contractions in similar test concentrations. Positive relaxant statins were tested again through construction of Calcium Concentration Response Curves (CCRCs) in the absence and presence of the statins using verapamil, a standard calcium channel blocker. CCRCs of statins were compared with CCRCs of verapamil. Simvastatin, atorvastatin, fluvastatin and rosuvastatin relaxed the spontaneous and KCl-induced contractions. IC50 for simvastatin on spontaneous rabbit's jejunal preparations is -5.08 ± 0.1 Log 10 M. Similarly, IC50 for KCl-induced contractions is -4.25 ± 0.01 Log 10 M. Mean IC50 (Log 10 M) for atorvastatin on spontaneous rabbit's jejunal preparations and KCl-induced contractions are -5.19 ± 0.07 and -4.37 ± 0.09, respectively. Fluvastatin relaxed spontaneous activity of rabbits' jejunal preparations with an IC50 (Log 10 M) -4.5 ± 0.03. Rosuvastatin relaxed spontaneous as well as KCl (80 mM) induced contractions with respective IC50 (Log 10 M) -3.62 ± 0.04 and -4.57 ± 0.06. In case of CCRCs, tissues pre-treated with 4.6 μg/ml of simvastatin, have IC50 = -1.84 ± 0.03 [log (Ca(++)) M] vs control IC50 = -2.54 ± 0.04 [log (Ca(++)) M]. Similarly, atorvastatin, fluvastatin and rosuvastatin produced significant right shift in IC50 for CCRCs (P ≤ 0.05). In case of verapamil, IC50 for control curves is -2.45 ± 0.06 [log (Ca (++)) M], while IC50 in presence of verapamil (0.1 μM) is -1.69 ± 0.05 [log (Ca (++)) M]. Statins produced right shift in the IC50 of CCRCs. The effects of statins are like that of effects of verapamil, a standard calcium channel blocker. Our findings suggest that current statins have calcium antagonistic effects that act on voltage gated calcium channels that may provide a rationale for cause muscle weakness and gastrointestinal disorders.

A new candidate of calcium channel blocker in silico from Tectona grandis for treatment of gestational hypertension

NASA Astrophysics Data System (ADS)

Azizah, A.; Suselo, Y. H.; Muthmainah, M.; Indarto, D.

2018-05-01

Gestational Hypertension is one of the three main causes of maternal mortality in Indonesia. Nifedipine which blockes the Cav1.2 calcium channel has frequently been used to treat gestational hypertension. However the efficacy of nifedipine has not been established yet and the prevalence of gestational hypertension is still high (27.1 %). Indonesian herbal plants have potential to be developed as natural drugs. Molecular docking, a computational method, is very often used to depict interaction between molecules and target receptor This study was therefore to identify Indonesian herbal plants that could inhibit the calcium channel in silico. This was a bioinformatics study with molecular docking approach. Three-dimensional structure of human calcium channel Cav1.2 was determined by modelling with rabbit calcium channel (ID:5GJW) as template and using the SWISS MODEL software. Nifedipine was used as a standard ligand and obtained from ZINC database with the access code ZINC19594578. Active compounds of Indonesian herbal plants were registered in HerbalDB database and their molecular structure was obtained from PubChem. Binding affinity of human Cav1.2 model-ligand complexes were assesed using AutoDock Vina 1.1.2 software and visualization of molecular conformation used Chimera 1.10 and PyMol 1.3 softwares. The Lipinsky’s rules of five were used to determine active compounds which fullfilled drug criteria. The human Cav1-2 model had 72.35% sequence identity with rabbit Cav1.1. Nifedipine bound to the human Cav1.2 model with -2.1 kcal/mol binding affinity and had binding sites at Gln1060, Phe1129, Ser1132, and Ile1173 residues. A lower binding affinity was observed in 8 phytochemicals but only obtusifolin 2-glucoside (-2.2 kcal/mol) had similar binding sites as nifedipin did. In addition, obtusifolin 2-glucoside met the Lipinsky criteria and the molecule conformation was similar with nifedipine. From the HerbalDB database, obtusifolin 2-glucoside is found in Tectona grandis. Obtusifolin 2-glucoside computationally becomes a potensial candidate of calcium channel blocker. In vitro assays should be performed to evaluate the antagonist effect of obtusifolin 2-glucoside on calcium channel Cav1.2.

Antihypertensive effect of barnidipine 10 mg or amlodipine 5 to 10 mg once daily in treatment-naive patients with essential hypertension: A 24-week, randomized, open-label, pilot study

PubMed Central

Rossetti, Giuseppe; Pizzocri, Samuele; Brasca, Francesco; Pozzi, Marta; Beltrami, Laura M.; Bolla, Giovanni B.; Famiani, Roberta; Caimi, Barbara; Omboni, Stefano; Magrini, Fabio; Carugo, Stefano

2008-01-01

Background: Dihydropyridine calcium antagonists are largely employed for the treatment of hypertension, coronary heart disease, and heart failure. Objective: The aim of our study was to compare the antihypertensive effect of the dihydropyridine calcium antagonists barnidipine and amlodipine. Methods: This was a 24-week, randomized, open-label, pilot study. Consecutive treatment-naive patients with grade I or II essential hypertension (office sitting systolic blood pressure [BP] of 140–179 mm Hg and diastolic BP of 90–109 mm Hg) were enrolled. The primary end points were the effect of treatment with either barnidipine 10 mg or amlodipine 5 mg once daily on office and ambulatory BP, left ventricular mass index (LVMI), and markers of cardiac damage, serum procollagen type I C-terminal propeptide, and plasma amino-terminal pro-B-type natriuretic peptide concentrations. Patients were assessed at enrollment, and 12 and 24 weeks. During each visit, the prevalence of adverse events (AEs) was also monitored using spontaneous reporting, patient interview, and physical examination, the relationship to study drug being determined by the investigators. Compliance with treatment was assessed at each study visit by counting returned tablets. Results: Thirty eligible patients (20 men, 10 women; mean [SD] age, 47 [12] years) were included in the study; all patients completed the 24 weeks of study treatment. Twelve weeks after randomization, 6 patients in the amlodipine group had their dose doubled to 10 mg due to inadequate BP control. Mean BP reductions at study end were not significantly different between the barnidipine and amlodipine groups (office BP, −10.3/−9.4 vs −16.6/−9.1 mm Hg; ambulatory BP, 9.4/6.4 vs 8.1/5.1 mm Hg). Reductions in LVMI and markers of cardiac damage were not significantly different between the 2 groups. Significantly more patients in the amlodipine group reported drug-related AEs compared with those in the barnidipine group (9 [60%] vs 2 [13%]; P < 0.05). Conclusion: In this small sample of treatment-naive hypertensive patients, the antihypertensive effect of barnidipine 10 mg once daily was not significantly different from that of amlodipine 5 to 10 mg once daily. PMID:24692798

Stimulation of airway sensory nerves by cyclosporin A and FK506 in guinea-pig isolated bronchus.

PubMed

Harrison, S; Reddy, S; Page, C P; Spina, D

1998-12-01

We have investigated the contractile property of cyclosporin A and FK506 in guinea-pig isolated bronchus. Cyclosporin A (10 microM) failed to significantly attenuate the excitatory non-adrenergic non-cholinergic (eNANC) and cholinergic contractile response (per cent methacholine Emax) induced by electrical field stimulation (EFS). In contrast, eNANC responses were significantly attenuated by both the neurokinin (NK)-1 and (NK)-2 receptor antagonists, N-acetyl-L-tryptophan 3,5-bis (trifluoromethyl)-benzyl and SR48968, respectively. Cyclosporin A and FK506 caused a concentration-dependent contraction in guinea-pig isolated bronchus, which was significantly attenuated by NK-1 and NK-2 receptor antagonists. The capsaicin receptor antagonist, capsazepine (10 microM) significantly reduced the contractile response to cyclosporin A and capsaicin, but not to FK506. The N-type calcium channel blocker, omega-Conotoxin (omegaCTX: 10 nM), significantly reduced the contractile response to FK506 and the eNANC response following EFS. In contrast, omega-CTX failed to significantly reduce the contractile potency to capsaicin or cyclosporin A. In bronchial preparations desensitized by repeated application of capsaicin (1 microM), the contractile responses to both cyclosporin A (100 microM) and FK506 (100 microM), were significantly reduced. In contrast, the contractile responses to substance P and neurokinin A (10 microM) were not altered. Furthermore, repeated application of cyclosporin A (100 microM) significantly inhibited the contractile response to capsaicin (1 microM). The findings from this study would indicate that cyclosporin A and FK506 mediate contraction of guinea-pig isolated bronchus secondary to the release of neuropeptides from airway sensory nerves. However, the release of sensory neuropeptides appears to be mediated via different mechanisms for cyclosporin A and FK506, the former by stimulation of the vanilloid receptor and the latter via opening of N-type calcium channels.

Calcium Channels, Rho-Kinase, Protein Kinase-C, and Phospholipase-C Pathways Mediate Mercury Chloride-Induced Myometrial Contractions in Rats.

PubMed

Koli, Swati; Prakash, Atul; Choudhury, Soumen; Mandil, Rajesh; Garg, Satish K

2018-05-21

Adverse effects of mercury on female reproduction are reported; however, its effect on myogenic activity of uterus and mechanism thereof is obscure. Present study was undertaken to unravel the mechanistic pathways of mercuric chloride (HgCl 2 )-induced myometrial contraction in rats. Isometric tension in myometrial strips of rats following in vitro exposure to HgCl 2 was recorded using data acquisition system-based physiograph. HgCl 2 produced concentration-dependent (10 nM-100 μM) uterotonic effect which was significantly (p < 0.05) reduced in Ca 2+ -free solution and inhibited in the presence of nifedipine (1 μM), a L-type Ca 2+ channel blocker, thus suggesting the importance of extracellular Ca 2+ and its entry through L-type calcium channels in HgCl 2 -induced myometrial contractions in rats. Cumulative concentration-response curve of HgCl 2 was significantly (p < 0.05) shifted towards right in the presence of Y-27632 (10 μM), a Rho-kinase inhibitor, suggesting the involvement of Ca 2+ -sensitization pathway in mediating HgCl 2 -induced myometrial contraction. HgCl 2 -induced myometrial contraction was also significantly (p < 0.05) inhibited in the presence of methoctramine or para-fluoro-hexahydro-siladifenidol, a selective M 2 and M 3 receptor antagonists, respectively, which evidently suggest that mercury also interacts with M 2 and M 3 muscarinic receptors to produce myometrial contractions. U-73122 and GF-109203X, the respective inhibitors of PLC and PKC-dependent pathways, downstream to the receptor activation, also significantly (p < 0.05) attenuated the uterotonic effect of HgCl 2 on rat uterus. Taken together, present study evidently reveals that HgCl 2 interacts with muscarinic receptors and activates calcium signaling cascades involving calcium channels, Rho-kinase, protein kinase-C, and phospholipase-C pathways to exert uterotonic effect in rats. Graphical Abstract Graphical abstract depicting the mechanism of mercury-induced myometrial contraction in rats. M receptor: Muscarinic receptor; PIP2: phospho-inositol bisphosphate; PLC: phospholipase-C; DAG: diacyl glycerol; IP3: inositol triphosphate; IP3R: inositol triphosphate receptor; PKC; protein kinase-C; MLCP: myosin light chain phosphatise; MYPT: myosin phosphatase; SR: sarco-endoplasmic reticulum.

ICI 182,780 has agonistic effects and synergizes with estradiol-17 beta in fish liver, but not in testis.

PubMed

Pinto, Patrícia I S; Singh, Pratap B; Condeça, João B; Teodósio, Helena R; Power, Deborah M; Canário, Adelino V M

2006-12-27

ICI 182,780 (ICI) belongs to a new class of antiestrogens developed to be pure estrogen antagonists and, in addition to its therapeutic use, it has been used to knock-out estrogen and estrogen receptor (ER) actions in several mammalian species. In the present study, the effects and mechanism of action of ICI were investigated in the teleost fish, sea bream (Sparus auratus). Three independent in vivo experiments were performed in which mature male tilapia (Oreochromis mossambicus) or sea bream received intra-peritoneal implants containing estradiol-17 beta (E2), ICI or a combination of both compounds. The effects of E2 and ICI on plasma calcium levels were measured and hepatic and testicular gene expression of the three ER subtypes, ER alpha, ER beta a and ER beta b, and the estrogen-responsive genes, vitellogenin II and choriogenin L, were analyzed by semi-quantitative RT-PCR in sea bream. E2 treatment caused an increase in calcium levels in tilapia, while ICI alone had no noticeable effect, as expected. However, pretreatment with ICI synergistically potentiated the effect of E2 on plasma calcium in both species. ICI mimicked some E2 actions in gene expression in sea bream liver upregulating ER alpha, vitellogenin II and choriogenin L, although, unlike E2, it did not downregulate ER beta a and ER beta b. In contrast, no effects of E2 or ICI alone were detected in the expression of ERs in testis, while vitellogenin II and choriogenin L were upregulated by E2 but not ICI. Finally, pretreatment with ICI had a synergistic effect on the hepatic E2 down-regulation of ER beta b, but apparently blocked the ER alpha up-regulation by E2. These results demonstrate that ICI has agonistic effects on several typical estrogenic responses in fish, but its actions are tissue-specific. The mechanisms for the ICI agonistic activity are still unknown; although the ICI induced up-regulation of ER alpha mRNA could be one of the factors contributing to the cellular response.

ICI 182,780 has agonistic effects and synergizes with estradiol-17 beta in fish liver, but not in testis

PubMed Central

Pinto, Patrícia IS; Singh, Pratap B; Condeça, João B; Teodósio, Helena R; Power, Deborah M; Canário, Adelino VM

2006-01-01

Background ICI 182,780 (ICI) belongs to a new class of antiestrogens developed to be pure estrogen antagonists and, in addition to its therapeutic use, it has been used to knock-out estrogen and estrogen receptor (ER) actions in several mammalian species. In the present study, the effects and mechanism of action of ICI were investigated in the teleost fish, sea bream (Sparus auratus). Methods Three independent in vivo experiments were performed in which mature male tilapia (Oreochromis mossambicus) or sea bream received intra-peritoneal implants containing estradiol-17 beta (E2), ICI or a combination of both compounds. The effects of E2 and ICI on plasma calcium levels were measured and hepatic and testicular gene expression of the three ER subtypes, ER alpha, ER beta a and ER beta b, and the estrogen-responsive genes, vitellogenin II and choriogenin L, were analyzed by semi-quantitative RT-PCR in sea bream. Results E2 treatment caused an increase in calcium levels in tilapia, while ICI alone had no noticeable effect, as expected. However, pretreatment with ICI synergistically potentiated the effect of E2 on plasma calcium in both species. ICI mimicked some E2 actions in gene expression in sea bream liver upregulating ER alpha, vitellogenin II and choriogenin L, although, unlike E2, it did not downregulate ER beta a and ER beta b. In contrast, no effects of E2 or ICI alone were detected in the expression of ERs in testis, while vitellogenin II and choriogenin L were upregulated by E2 but not ICI. Finally, pretreatment with ICI had a synergistic effect on the hepatic E2 down-regulation of ER beta b, but apparently blocked the ER alpha up-regulation by E2. Conclusion These results demonstrate that ICI has agonistic effects on several typical estrogenic responses in fish, but its actions are tissue-specific. The mechanisms for the ICI agonistic activity are still unknown; although the ICI induced up-regulation of ER alpha mRNA could be one of the factors contributing to the cellular response. PMID:17192186

A novel impedance-based cellular assay for the detection of anti-calcium channel autoantibodies in type 1 diabetes.

PubMed

Jackson, Michael W; Gordon, Tom P

2010-09-30

We have recently postulated that functional autoantibodies (Abs) against L-type voltage-gated calcium channels (VGCCs) contribute to autonomic dysfunction in type 1 diabetes (T1D). Previous studies based on whole-organ assays have proven valuable in establishing the mechanism of anti-VGCC Ab activity, but are complex and unsuitable for screening large patient cohorts. In the current study, we used real-time dynamic monitoring of cell impedance to demonstrate that anti-VGCC Abs from patients with T1D inhibit the adherence of Rin A12 cells. The functional effect of the anti-VGCC Abs was mimicked by the dihydropyridine agonist, Bay K8644, and reversed by the antagonist, nicardipine, providing a pharmacological link to the whole-organ studies. IVIg neutralized the effect on cell adhesion of the anti-VGCC Abs, consistent with the presence of anti-idiotypic Abs in IVIg that may prevent the emergence of pathogenic Abs in healthy individuals. The cell impedance assay can be performed in a 96 well plate format, and represents a simple method for detecting the presence of anti-VGCC activity in patient immunoglobulin (IgG). The new cell assay should prove useful for further studies to determine the prevalence of the Ab and its association with symptoms of autonomic dysfunction in patients with T1D. Crown Copyright © 2010. Published by Elsevier B.V. All rights reserved.

Transient Receptor Potential Melastatin-3 (TRPM3) Mediates Nociceptive-Like Responses in Hydra vulgaris

PubMed Central

Malafoglia, Valentina; Traversetti, Lorenzo; Del Grosso, Floriano; Scalici, Massimiliano; Lauro, Filomena; Russo, Valeria; Persichini, Tiziana; Salvemini, Daniela; Mollace, Vincenzo; Fini, Massimo; Raffaeli, William

2016-01-01

The ability of mammals to feel noxious stimuli lies in a heterogeneous group of primary somatosensory neurons termed nociceptors, which express specific membrane receptors, such as the Transient Receptor Potential (TRP) family. Here, we show that one of the most important nociceptive-like pathways is conserved in the freshwater coelenterate Hydra vulgaris, the most primitive organism possessing a nervous system. In particular, we found that H. vulgaris expresses TRPM3, a nociceptor calcium channel involved in the detection of noxious heat in mammals. Furthermore, we detected that both heat shock and TRPM3 specific agonist (i.e., pregnenolone sulfate) induce the modulation of the heat shock protein 70 (HSP70) and the nitric oxide synthase (NOS), two genes activated by TRP-mediated heat painful stimuli in mammals. As expected, these effects are inhibited by a TRPM3 antagonist (i.e., mefenamic acid). Interestingly, the TRPM3 agonist and heat shock also induce the expression of nuclear transcription erythroid 2-related factor (Nrf2) and superoxide dismutase (SOD), known markers of oxidative stress; noteworthy gene expression was also inhibited by the TRPM3 antagonist. As a whole, our results demonstrate the presence of conserved molecular oxidative/nociceptive-like pathways at the primordial level of the animal kingdom. PMID:26974325

Transient Receptor Potential Melastatin-3 (TRPM3) Mediates Nociceptive-Like Responses in Hydra vulgaris.

PubMed

Malafoglia, Valentina; Traversetti, Lorenzo; Del Grosso, Floriano; Scalici, Massimiliano; Lauro, Filomena; Russo, Valeria; Persichini, Tiziana; Salvemini, Daniela; Mollace, Vincenzo; Fini, Massimo; Raffaeli, William; Muscoli, Carolina; Colasanti, Marco

2016-01-01

The ability of mammals to feel noxious stimuli lies in a heterogeneous group of primary somatosensory neurons termed nociceptors, which express specific membrane receptors, such as the Transient Receptor Potential (TRP) family. Here, we show that one of the most important nociceptive-like pathways is conserved in the freshwater coelenterate Hydra vulgaris, the most primitive organism possessing a nervous system. In particular, we found that H. vulgaris expresses TRPM3, a nociceptor calcium channel involved in the detection of noxious heat in mammals. Furthermore, we detected that both heat shock and TRPM3 specific agonist (i.e., pregnenolone sulfate) induce the modulation of the heat shock protein 70 (HSP70) and the nitric oxide synthase (NOS), two genes activated by TRP-mediated heat painful stimuli in mammals. As expected, these effects are inhibited by a TRPM3 antagonist (i.e., mefenamic acid). Interestingly, the TRPM3 agonist and heat shock also induce the expression of nuclear transcription erythroid 2-related factor (Nrf2) and superoxide dismutase (SOD), known markers of oxidative stress; noteworthy gene expression was also inhibited by the TRPM3 antagonist. As a whole, our results demonstrate the presence of conserved molecular oxidative/nociceptive-like pathways at the primordial level of the animal kingdom.

[Antihypertensive treatment for patients with hypertension and diabetes type II--current clinical research].

PubMed

Rajzer, Marek; Kawecka-Jaszcz, Kalina; Wojciechowska, Wiktoria

2003-01-01

The frequency of arterial hypertension occurrence in polish population amounts to 30-40%, among diabetics is significantly higher-70%. According to the WHO/ISH Guidelines all hypertensive patients with diabetes are included into the "high risk group" independent of hypertension stage. Pharmacological treatment of hypertension is this group of patients has a particular meaning. Among hypertensive patients the degree of blood pressure lowering is more effective for cardiovascular risk reduction than choice of drug. This fact is well documented in clinical trials comparing antihypertensive efficacy of old and new antihypertensive drugs (for example UKPDS, STOP 2, INSIGHT). From the other point of view renal protection and metabolic benefits, as well as reduction of target organ damage are more advantageous for angiotensin converting enzyme inhibitors, angiotensin II receptor antagonists and calcium antagonists than for diuretics and beta-blockers. Despite fast progress in clinical research on new antihypertensive drugs (especially AT1 receptor inhibitors) ACE-I seem to still remain still the "first choice" for hypertensive diabetics. Adequate blood pressure control among diabetic hypertensives is of special importance and usually needs appropriate combined antihypertensive therapy. Our review presents detailed information about treatment advantages and disadvantages of drugs from different antihypertensive classes in light of current clinical trials and international guidelines.

Memantine Inhibits α3β2-nAChRs-Mediated Nitrergic Neurogenic Vasodilation in Porcine Basilar Arteries

PubMed Central

Wu, Celeste Yin-Chieh; Chen, Po-Yi; Chen, Mei-Fang; Kuo, Jon-Son; Lee, Tony Jer-Fu

2012-01-01

Memantine, an NMDA receptor antagonist used for treatment of Alzheimer’s disease (AD), is known to block the nicotinic acetylcholine receptors (nAChRs) in the central nervous system (CNS). In the present study, we examined by wire myography if memantine inhibited α3β2-nAChRs located on cerebral perivascular sympathetic nerve terminals originating in the superior cervical ganglion (SCG), thus, leading to inhibition of nicotine-induced nitrergic neurogenic dilation of isolated porcine basilar arteries. Memantine concentration-dependently blocked nicotine-induced neurogenic dilation of endothelium-denuded basilar arteries without affecting that induced by transmural nerve stimulation, sodium nitroprusside, or isoproterenol. Furthermore, memantine significantly inhibited nicotine-elicited inward currents in Xenopous oocytes expressing α3β2-, α7- or α4β2-nAChR, and nicotine-induced calcium influx in cultured rat SCG neurons. These results suggest that memantine is a non-specific antagonist for nAChR. By directly inhibiting α3β2-nAChRs located on the sympathetic nerve terminals, memantine blocks nicotine-induced neurogenic vasodilation of the porcine basilar arteries. This effect of memantine is expected to reduce the blood supply to the brain stem and possibly other brain regions, thus, decreasing its clinical efficacy in the treatment of Alzheimer’s disease. PMID:22792283

Neuroprotective effects of NPS 846, a novel N-methyl-D-aspartate receptor antagonist, after closed head trauma in rats.

PubMed

Gurevich, B; Artru, A A; Lam, A M; Mueller, A L; Merkind, V; Talmor, D; Katchko, L; Shapira, Y

1998-06-01

The authors sought to determine whether 3,3-bis (3-fluorophenyl) propylamine (NPS 846), a novel noncompetitive N-methyl-D-aspartate receptor antagonist, alters outcome after closed head trauma in rats. The experimental variables were: presence or absence of closed head trauma, treatment with NPS 846 or no treatment, and time at which the rats were killed (24 or 48 hours). The NPS 846 (1 mg/kg) was administered intraperitoneally at 1 and 3 hours after closed head trauma or sham operation. Outcome measures were the neurological severity score (NSS), ischemic tissue volume, hemorrhagic necrosis volume, and specific gravity, water content, and concentrations of calcium, sodium, potassium, and magnesium in brain tissue. The following closed head trauma-induced changes in the injured hemisphere (expressed as the mean +/- the standard deviation) were reversed by NPS 846: decreased specific gravity of 1.035 +/- 0.006 at 24 hours was increased to 1.042 +/- 0.004; the decreased potassium level of 0.583 +/- 0.231 mg/L at 48 hours and at 24 hours was increased to 2.442 +/- 0.860 mg/L; the increased water content of 84.7 +/- 2.6% at 24 hours was decreased to 79.8 +/- 2%; the increased calcium level of 0.592 +/- 0.210 mg/L at 24 hours was decreased to 0.048 +/- 0.029 mg/L; and the increased sodium level of 2.035 +/- 0.649 mg/L was decreased to 0.631 +/- 0.102 mg/L. Administration of NPS 846 also lowered the NSS (improved neurological status) at 48 hours (7 +/- 3) and caused no significant changes in ischemic tissue or hemorrhagic necrosis volumes in the injured hemisphere at 24 or 48 hours. In this model of closed head trauma, NPS 846 improved neurological outcome, delayed the onset of brain edema, and improved brain tissue ion homeostasis.

PACAP Protects Against Ethanol and Nicotine Toxicity in SH-SY5Y Cells: Implications for Drinking-Smoking Co-morbidity.

PubMed

Manavalan, Sridharan; Getachew, Bruk; Manaye, Kebreten F; Khundmiri, Syed J; Csoka, Antonei B; McKinley, Raechel; Tamas, Andrea; Reglodi, Dora; Tizabi, Yousef

2017-07-01

The detrimental effects of heavy drinking and smoking are multiplied when the two are combined. Treatment modalities for each and especially for the combination are very limited. Although in low concentration, alcohol and nicotine, each may have beneficial effects including neuroprotection, their combination, instead of providing additive protection, may actually lead to toxicity in cell cultures. Pituitary adenylate cyclase-activating polypeptide (PACAP) is an endogenous 38 amino-acid peptide with demonstrated protection against neuronal injury, trauma as well as various endogenous and exogenous toxic agents. The aim of this study was to investigate whether PACAP may also protect against toxicity induced by high alcohol, high nicotine, or the combination of low alcohol and nicotine concentrations, and if so, whether this effect was mediated via PAC1 receptor. We used the neuroblastoma-derived SH-SY5Y cells and applied various colorimetric assays for determination of cell viability or toxicity. Results indicate that PACAP blocks toxicity induced by high alcohol and high nicotine as well as their combination at low concentrations. The effects of PACAP in turn were blocked by the PACAP antagonist (PACAP 6-38), indicating involvement of the PACAP receptor PAC1 and possibly vasoactive intestinal peptide (VIP) receptors in PACAP's protection. Moreover, no combined toxicity of low alcohol and low nicotine could be detected in calcium-free medium. These findings suggest possible beneficial effects of PACAP in preventing alcohol and nicotine toxicity and that calcium contributes to the damage induced by combination of low alcohol and nicotine in SH-SY5Y cells.

Role of opioid system in verapamil-induced antinociception in a rat model of orofacial pain

PubMed Central

Tamaddonfard, Esmaeal; Erfanparast, Amir; Taati, Mina; Dabbaghi, Milad

2014-01-01

Calcium, through its various channels involves in local, spinal and supra-spinal transmission of pain. In the present study, we investigated the separate and combined treatment effects of verapamil (a calcium channel blocker), morphine (an opioid agonist) and naloxone (an opioid antagonist) on pain in the orofacial region of rats. Orofacial pain was induced by subcutaneous (SC) injection of formalin (50 µL, 1.5%) into the left upper lip side, and the time durations spent face rubbing with epsilateral forepaw were recorded in three min blocks for a period of 45 min. Formalin induced a biphasic pattern (first phase: 0-3 min; second phase: 15-33 min) of pain. Intraperitoneal (IP) injections of verapamil (2 and 8 mg kg-1) and morphine (2 and 4 mg kg-1) suppressed orofacial pain. Co-administration of sub-analgesic doses of verapamil (0.5 mg kg-1) and morphine (1 mg kg-1) produced second phase analgesia. Both phases of formalin-induced pain were suppressed when an analgesic dose (2 mg kg-1) of verapamil co-administered with a sub-analgesic dose (1 mg kg-1) of morphine. The SC injection of naloxone (2 mg kg-1) alone with no effect on pain intensity, prevented the antinociceptive effects induced by morphine (2 mg kg-1), but not verapamil (2 mg kg-1). The obtained results showed antinociceptive effects for verapamli and morphine on orofacial pain. Co-administrations of verapamil and morphine produced antinociceptive effects. It seems that opioid analgesic system may not have a role in the verapamil-induced antinociception. PMID:25568692

Lipid effects on neutrophil calcium signaling induced by opsonized particles: platelet activating factor is only part of the story.

PubMed

Wanten, Geert; Kusters, Anneke; van Emst-de Vries, Sjenet E; Tool, Anton; Roos, Dirk; Naber, Ton; Willems, Peter

2004-08-01

Total parenteral nutrition is frequently used in clinical practice to improve the nutritional status of patients. However, the risk for infectious complications remains a drawback in which immune-modulating effects of the lipid component may play a role. To characterize these lipid effects we investigated neutrophil activation by opsonized yeast particles under influence of lipid emulsions derived from fish oil (VLCT), olive oil (LCT-MUFA), soybean oil (LCT), and a physical mixture of coconut and soybean oil (LCT-MCT). Serum-treated zymosan (STZ) evoked a biphasic increase in cytosolic Ca2+ concentration ([Ca2+]c) with an initial slow rise that turned into a second fast rise until a plateau was reached. LCT-MCT (5 mM) pretreatment markedly increased the rate of [Ca2+]c rise during the initial phase, abolished the second phase and lowered the plateau. These effects of LCT-MCT were mimicked by the protein kinase C (PKC) activating phorbol ester PMA. LCT, LCT-MUFA and VLCT, on the other hand, decreased the rate of [Ca2+]c rise during both phases and lowered the plateau. The platelet-activating factor (PAF) receptor antagonist WEB 2086 inhibited the second phase, demonstrating that PAF acts as an intercellular messenger in STZ-induced Ca2+ mobilization, but did not interfere with the stimulatory effect of LCT-MCT or PMA on the initial rate of [Ca2+]c rise. Structurally different lipids act only in part through PAF to distinctively modulate neutrophil calcium signaling in response to activation by opsonized particles. Copyright 2003 Elsevier Ltd.

Use of intralesional verapamil to dissolve Peyronie's disease plaque: a long-term single-blind study.

PubMed

Rehman, J; Benet, A; Melman, A

1998-04-01

Multiple conservative therapies for the treatment of Peyronie's disease have been offered with variable and poor response rates. Calcium channel blockers have been shown in vitro and in vivo to inhibit secretion and synthesis of extracellular matrix, including collagen, glycosaminoglycans, and fibronectin, as well as causing increased collagenase and anti transforming growth factor-beta activity. Calcium antagonists, including verapamil, are effective in stimulating the remodeling and degradation of extracellular matrix in tissue by altering the metabolic pathways of fibroblasts. Recently, a pilot study (1994) showed preliminary promising results in treating plaque caused by Peyronie's disease. This randomized single-blind placebo-based study (1994 to 1996) was undertaken to confirm the hypothesis. In this randomized single-blind study, 14 patients completed the study and were divided into two groups: the verapamil treatment group (n = 7) or the control saline group (n = 7). Verapamil or saline was injected directly into the Peyronie's plaque once a week for 6 months. Patients were evaluated before and after treatment with duplex ultrasound to confirm the extent of the lesion and to measure volume of the plaque, and by interview and mailed questionnaire 3 months after treatment. Patients being treated with oral calcium antagonists were excluded from the study. A decreased plaque volume was measured in 57% of the verapamil-treated men versus 28% in the control group (P <0.04). Penile curvature demonstrated an improvement trend of 37.71 +/- 9.3 degrees to 29.57 +/- 7.3 degrees in the verapamil-treated patients, but the difference was not significant (P <0.07). Plaque softening was noted in all patients treated with verapamil. There was significant objective improvement in plaque-associated penile narrowing in all patients in the verapamil group. Subjective plaque-associated erectile dysfunction (quality of erection) showed improvement in 42.87% of the verapamil group versus none in the control group (P <0.02). There was no local or systemic toxicity except for an occasional ecchymosis/bruise at the injection site. After a positive clinical response, plaque size, penile angulation, and symptoms continued to improve. Decrease in plaque size was noted in each of the responders in the first 3 months. This randomized single-blind study suggests that intralesional injection of calcium channel blocker may be a reasonable approach in some selected patients for the treatment of Peyronie's disease with noncalcified plaque and penile angulation of less than 30 degrees. Patients whose plaque failed to respond to intralesional verapamil therapy within 3 months or whose angulation was greater than 30 degrees at presentation were more likely to benefit from surgery.

Autophagy inhibition attenuates hyperoxaluria-induced renal tubular oxidative injury and calcium oxalate crystal depositions in the rat kidney.

PubMed

Duan, Xiaolu; Kong, Zhenzhen; Mai, Xin; Lan, Yu; Liu, Yang; Yang, Zhou; Zhao, Zhijian; Deng, Tuo; Zeng, Tao; Cai, Chao; Li, Shujue; Zhong, Wen; Wu, Wenqi; Zeng, Guohua

2018-06-01

Hyperoxaluria-induced oxidative injury of renal tubular epithelial cell is a casual and essential factor in kidney calcium oxalate (CaOx) stone formation. Autophagy has been shown to be critical for the regulation of oxidative stress-induced renal tubular injury; however, little is known about its role in kidney CaOx stone formation. In the present study, we found that the autophagy antagonist chloroquine could significantly attenuate oxalate-induced autophagy activation, oxidative injury and mitochondrial damage of renal tubular cells in vitro and in vivo, as well as hyperoxaluria-induced CaOx crystals depositions in rat kidney, whereas the autophagy agonist rapamycin exerted contrasting effects. In addition, oxalate-induced p38 phosphorylation was significantly attenuated by chloroquine pretreatment but was markedly enhanced by rapamycin pretreatment, whereas the protective effect of chloroquine on rat renal tubular cell oxidative injury was partly reversed by a p38 protein kinase activator anisomycin. Furthermore, the knockdown of Beclin1 represented similar effects to chloroquine on oxalate-induced cell oxidative injury and p38 phosphorylation in vitro. Taken together, our results revealed that autophagy inhibition could attenuate oxalate-induced oxidative injury of renal tubular cell and CaOx crystal depositions in the rat kidney via, at least in part, inhibiting the activation of p38 signaling pathway, thus representing a novel role of autophagy in the regulation of oxalate-induced renal oxidative injury and CaOx crystal depositions for the first time. Copyright © 2018. Published by Elsevier B.V.

Involvement of peripheral mechanism in the verapamil-induced potentiation of morphine analgesia in mice.

PubMed

Shimizu, Norifumi; Kishioka, Shiroh; Maeda, Takehiko; Fukazawa, Yohji; Dake, Yoshihiro; Yamamoto, Chizuko; Ozaki, Masanobu; Yamamoto, Hiroyuki

2004-08-01

Morphine's analgesic actions are thought to be mediated through both the central and peripheral nervous systems. L-type calcium channel blockers have been reported to potentiate the analgesic effects of morphine, but the locus of this interaction is not known. In this experiment, we examined the site of verapamil-induced potentiation of morphine analgesia in mice using the quaternary opioid receptor antagonist naloxone-methiodide (NLX-M). Subcutaneous injections of morphine increased locomotor activity and serum corticosterone level, which are mediated by the central nervous system. These central effects were not antagonized by 0.1 mg/kg of NLX-M, whereas this dose of NLX-M partially antagonized the analgesic effect of morphine. Treatment with verapamil potentiated morphine analgesia in a dose-dependent manner. The verapamil-induced potentiation of morphine analgesia was abolished by pretreatment with NLX-M (0.1 and 1 mg/kg). These findings suggest that peripheral mechanisms partially contribute to morphine analgesia and mediate the potentiation of morphine analgesia by verapamil.

BDNF Up-Regulates α7 Nicotinic Acetylcholine Receptor Levels on Subpopulations of Hippocampal Interneurons

PubMed Central

Massey, Kerri A.; Zago, Wagner M.; Berg, Darwin K.

2006-01-01

In the hippocampus, brain-derived neurotrophic factor (BDNF) regulates a number of synaptic components. Among these are nicotinic acetylcholine receptors containing α7 subunits (α7-nAChRs), which are interesting because of their relative abundance in the hippocampus and their high relative calcium permeability. We show here that BDNF elevates surface and intracellular pools of α7-nAChRs on cultured hippocampal neurons and that glutamatergic activity is both necessary and sufficient for the effect. Blocking transmission through NMDA receptors with APV blocked the BDNF effect; increasing spontaneous excitatory activity with the GABAA receptor antagonist bicuculline replicated the BDNF effect. BDNF antibodies blocked the BDNF-mediated increase but not the bicuculline one, consistent with enhanced glutamatergic activity acting downstream from BDNF. Increased α7-nAChR clusters were most prominent on interneuron subtypes known to innervate directly excitatory neurons. The results suggest that BDNF, acting through glutamatergic transmission, can modulate hippocampal output in part by controlling α7-nAChR levels. PMID:17029981

Clevidipine for hypertension treatment in pheochromocytoma surgery.

PubMed

Luis-García, C; Arbonés-Aran, E; Teixell-Aleu, C; Lorente-Poch, L; Trillo-Urrutia, L

2018-04-01

Pheochromocytoma is a catecholamine-producing tumour and laparoscopic adrenalectomy is its treatment of choice. During pneumoperitoneum insufflation and tumour handling there is a high risk of massive catecholamine release and hypertensive crisis. After tumour excision, severe arterial hypotension is a common effect, due to relative vasodilation and the residual effect of antihypertensive drugs. We report the case of a patient with pheochromocytoma who was treated with laparoscopic adrenalectomy. During surgical manipulation there was a sudden hypertensive peak that could be controlled quickly with clevidipine infusion. After tumour resection, clevidipine perfusion was stopped and there were no arterial hypotension episodes. Clevidipine is a new intravenous calcium antagonist with rapid onset of action and short half-life that has no residual effect and does not produce arterial hypotension after tumour resection. For these reasons, it can be a first-choice drug for this kind of surgery. Copyright © 2017 Sociedad Española de Anestesiología, Reanimación y Terapéutica del Dolor. Publicado por Elsevier España, S.L.U. All rights reserved.

Spasmogenic and spasmolytic activities of Agastache mexicana ssp. mexicana and A. mexicana ssp. xolocotziana methanolic extracts on the guinea pig ileum.

PubMed

Ventura-Martínez, Rosa; Rodríguez, Rodolfo; González-Trujano, María Eva; Ángeles-López, Guadalupe E; Déciga-Campos, Myrna; Gómez, Claudia

2017-01-20

Agastache mexicana has been used in traditional medicine for relief of abdominal pain and treatment of other diseases. Two subspecies have been identified: A. mexicana ssp. mexicana (AMM) and A. mexicana ssp. xolocotziana (AMX) and both are used traditionally without distinction or in combination. To determine the effect of methanol extracts of A. mexicana ssp. mexicana and A. mexicana ssp. xolocotziana on gut motility and their possible mechanism of action. The effect of AMM and AMX methanol extracts were tested on the spontaneous activity in the isolated guinea pig ileum and on tissues pre-contracted with KCl, electrical field stimulation (EFS) or ACh. In addition, the possible mechanism of action of each subspecies on gut motility was analyzed in the presence of hexametonium, indomethacin, L-NAME, verapamil, atropine or pyrylamine. A comparative chromatographic profile of these extracts was also done to indicate the most abundant flavonoids presents in methanol extracts of both subspecies. AMM, but not AMX, induced a contractile effect in the guinea pig ileum. This spasmogenic effect was partially inhibited by atropine, antagonist of muscarinic receptors; and pyrilamine, antagonist of H 1 receptors. In contrast, AMX, but not AMM, diminished the contractions induced by KCl, EFS or ACh. The spasmolytic activity of AMX was partially inhibited by hexamethonium, ganglionic blocker; and indomethacin, inhibitor of the synthesis of prostaglandins; but not by L-NAME, inhibitor of nitric oxide synthase. In addition, AMX diminished the maximal contraction induced by CaCl 2 in a calcium-free medium. Chromatographic analyses of these methanol extracts showed the presence of acacetin and tilanin in both. These results suggest that in folk medicine only AMX should be used as spasmolytic, and not in combination with AMM as traditionally occurs, due to the spasmogenic effects of the latter. In addition, activation of nicotinic receptors, prostaglandins and calcium channels, but not nitric oxide mechanisms, could be responsible for the spasmolytic activity of AMX. On the other hand, release of ACh and histamine could be involved in the spasmogenic effect induced by AMM. Acacetin and tilanin are present in methanol extracts of both subspecies and both flavonoids were more abundant in AMX than AMM. Our findings contribute to the validation of the traditional use of Agastache mexicana in relieving gastrointestinal disorders, but indicate that the subspecie that should be used for this effect is A. mexicana ssp. xolocotziana. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Mechanism of action of relaxant effect of Agastache mexicana ssp.mexicana essential oil in guinea-pig trachea smooth muscle.

PubMed

Navarrete, Andrés; Ávila-Rosas, Natalia; Majín-León, Mateo; Balderas-López, José Luis; Alfaro-Romero, Alejandro; Tavares-Carvalho, José Carlos

2017-12-01

Agastache mexicana ssp. mexicana (Kunth) Lint & Epling (Lamiaceae), popularly known as 'toronjil morado', is used in Mexican traditional medicine for the treatment of several diseases such as hypertension, anxiety and respiratory disorders. This study investigates the relaxant action mechanism of A. mexicana ssp. mexicana essential oil (AMEO) in guinea-pig isolated trachea model. AMEO was analyzed by GC/MS. The relaxant effect of AMEO (5-50 μg/mL) was tested in guinea-pig trachea pre-contracted with carbachol (3 × 10  -   6  M) or histamine (3 × 10  -   5  M) in the presence or absence of glibenclamide (10  -   5  M), propranolol (3 × 10  -   6  M) or 2',5'-dideoxyadenosine (10  -   5  M). The antagonist effect of AMEO (10-300 μg/mL) against contractions elicited by carbachol (10  -   15 -10  -   3  M), histamine (10  -   15 -10  -   3  M) or calcium (10-300 μg/mL) was evaluated. Essential oil composition was estragole, d-limonene and linalyl anthranilate. AMEO relaxed the carbachol (EC 50  =   18.25 ± 1.03 μg/mL) and histamine (EC 50  =   13.3 ± 1.02 μg/mL)-induced contractions. The relaxant effect of AMEO was not modified by the presence of propranolol, glibenclamide or 2',5'-dideoxyadenosine, suggesting that effect of AMEO is not related to β 2 -adrenergic receptors, ATP-sensitive potassium channels or adenylate cyclase activation. AMEO was more potent to antagonize histamine (pA 2 ' = -1.507 ± 0.122) than carbachol (pA 2 ' = -2.180 ± 0.357). Also, AMEO antagonized the calcium chloride-induced contractions. The results suggest that relaxant effect of AMEO might be due to blockade of calcium influx in guinea-pig trachea smooth muscle. It is possible that estragole and d-limonene could contribute majority in the relaxant effect of AMEO.

Abscisic Acid Acts as a Blocker of the Bitter Taste G Protein-Coupled Receptor T2R4.

PubMed

Pydi, Sai P; Jaggupilli, Appalaraju; Nelson, Ken M; Abrams, Suzanne R; Bhullar, Rajinder P; Loewen, Michele C; Chelikani, Prashen

2015-04-28

Bitter taste receptors (T2Rs) belong to the G protein-coupled receptor superfamily. In humans, 25 T2Rs mediate bitter taste sensation. In addition to the oral cavity, T2Rs are expressed in many extraoral tissues, including the central nervous system, respiratory system, and reproductive system. To understand the mechanistic roles of the T2Rs in oral and extraoral tissues, novel blockers or antagonists are urgently needed. Recently, we elucidated the binding pocket of T2R4 for its agonist quinine, and an antagonist and inhibitory neurotransmitter, γ-aminobutyric acid. This structure-function information about T2R4 led us to screen the plant hormone abscisic acid (ABA), its precursor (xanthoxin), and catabolite phaseic acid for their ability to bind and activate or inhibit T2R4. Molecular docking studies followed by functional assays involving calcium imaging confirmed that ABA is an antagonist with an IC50 value of 34.4 ± 1.1 μM. However, ABA precursor xanthoxin acts as an agonist on T2R4. Interestingly, molecular model-guided site-directed mutagenesis suggests that the T2R4 residues involved in quinine binding are also predominantly involved in binding to the novel antagonist, ABA. The antagonist ability of ABA was tested using another T2R4 agonist, yohimbine. Our results suggest that ABA does not inhibit yohimbine-induced T2R4 activity. The discovery of natural bitter blockers has immense nutraceutical and physiological significance and will help in dissecting the T2R molecular pathways in various tissues.

Can heart rate predict blood pressure response to anti-hypertensive drug therapy?

PubMed

Owens, P E; Lyons, S; O'Brien, E

1998-04-01

The use of heart rate in clinical practice is limited by its variability under measurement situations. The mean heart rate on ambulatory monitoring provides a more robust statistic for clinical use. We examined the relationship between mean heart rate on initial referral ambulatory blood pressure monitoring (ABPM) to the BP-lowering efficacy of the four main groups of anti-hypertensive medications, in a referral hypertensive population. Patients were retrospectively identified by review of the BP database, and data collected from the initial referral BP monitor off medication, and the subsequent ABPM after treatment with either beta-blockers, diuretics, calcium antagonists or angiotensin-converting enzyme (ACE) inhibitors. The change in mean arterial BP from the initial to the subsequent ABPM (ie, as a result of treatment) was correlated with the mean heart rate on the initial ABPM. A moderate association was found for initial daytime heart rate and BP response to beta-blockers (r = 0.24, P = 0.02), and ACE inhibitors (r = 0.14, P = 0.05). No such association was found for calcium antagonists or diuretics. When the groups were divided into those with a mean daytime heart rate <75 and > or =75 beats per min, BP reduction from beta-blocker and ACE inhibitor therapy was significantly greater in those patients with a higher daytime heart rate. We conclude that average daytime heart rate on pre-treatment ABPM can be useful as a predictor of BP response to beta-blockade or ACE inhibition.

Seventeen-Year Nationwide Trends in Antihypertensive Drug Use in Denmark.

PubMed

Sundbøll, Jens; Adelborg, Kasper; Mansfield, Kathryn E; Tomlinson, Laurie A; Schmidt, Morten

2017-12-15

Recent trends in use of antihypertensive drugs are unknown. From Danish nationwide prescription data, we obtained information on primary care use of angiotensin-converting enzyme inhibitors, angiotensin II receptor blockers, beta blockers, diuretics, aldosterone receptor antagonists, and calcium channel blockers. During 1999 to 2015, the use of antihypertensive drugs per 1,000 inhabitants/day increased from 184 to 379 defined daily doses (DDD), corresponding to a rise in the prevalence proportion of users from ≈20% to ≈35%. From 1999 to 2015, a notable increase was observed for angiotensin-converting enzyme inhibitors (from 29 to 105 DDD per 1,000 inhabitants/day ≈260%) and angiotensin II receptor blockers (from 13 to 73 DDD per 1,000 inhabitants/day ≈520%). For diuretics the use remained stable, with a slight decrease (from 89 to 81 DDD per 1,000 inhabitants/day ≈-10%). The use of aldosterone receptor antagonists increased until 2007 and remained unchanged at around 3.5 DDD per 1,000 inhabitants/day thereafter (average change ≈65%). The use of beta blockers doubled during the study period (from 17 to 34 DDD per 1,000 inhabitants/day ≈100%), entirely driven by increasing use of metoprolol. Similar trends were observed for calcium channel blockers (from 34 to 82 DDD per 1,000 inhabitants/day ≈140%), where amlodipine drove the overall increase. In conclusion, antihypertensive drug use has increased remarkably during the past 2 decades. Copyright © 2017 Elsevier Inc. All rights reserved.

Therapeutic perspectives in hypertension: novel means for renin–angiotensin–aldosterone system modulation and emerging device-based approaches

PubMed Central

Unger, Thomas; Paulis, Ludovit; Sica, Domenic A.

2011-01-01

The conventional antihypertensive therapies including renin–angiotensin–aldosterone system antagonists (converting enzyme inhibitors, receptor blockers, renin inhibitors, and mineralocorticoid receptor blockers), diuretics, β-blockers, and calcium channel blockers are variably successful in achieving the challenging target blood pressure values in hypertensive patients. Difficult to treat hypertension is still a commonly observed problem world-wide. A number of drugs are considered to be used as novel therapies for hypertension. Renalase supplementation, vasopeptidase inhibitors, endothelin antagonists, and especially aldosterone antagonists (aldosterone synthase inhibitors and novel selective mineralocorticoid receptor blockers) are considered an option in resistant hypertension. In addition, the aldosterone antagonists as well as (pro)renin receptor blockers or AT2 receptor agonists might attenuate end-organ damage. This array of medications has now been complemented by a number of new approaches of non-pharmacological strategies including vaccination, genomic interference, controlled breathing, baroreflex activation, and probably most successfully renal denervation techniques. However, the progress on innovative therapies seems to be slow and the problem of resistant hypertension and proper blood pressure control appears to be still persisting. Therefore the regimens of currently available drugs are being fine-tuned, resulting in the establishment of several novel fixed-dose combinations including triple combinations with the aim to facilitate proper blood pressure control. It remains an exciting question which approach will confer the best blood pressure control and risk reduction in this tricky disease. PMID:21951628

T-type calcium channels in synaptic plasticity

PubMed Central

Lambert, Régis C.

2017-01-01

ABSTRACT The role of T-type calcium currents is rarely considered in the extensive literature covering the mechanisms of long-term synaptic plasticity. This situation reflects the lack of suitable T-type channel antagonists that till recently has hampered investigations of the functional roles of these channels. However, with the development of new pharmacological and genetic tools, a clear involvement of T-type channels in synaptic plasticity is starting to emerge. Here, we review a number of studies showing that T-type channels participate to numerous homo- and hetero-synaptic plasticity mechanisms that involve different molecular partners and both pre- and post-synaptic modifications. The existence of T-channel dependent and independent plasticity at the same synapse strongly suggests a subcellular localization of these channels and their partners that allows specific interactions. Moreover, we illustrate the functional importance of T-channel dependent synaptic plasticity in neocortex and thalamus. PMID:27653665

Treatment for calcium channel blocker poisoning: A systematic review

PubMed Central

Dubé, P.-A.; Gosselin, S.; Guimont, C.; Godwin, J.; Archambault, P. M.; Chauny, J.-M.; Frenette, A. J.; Darveau, M.; Le sage, N.; Poitras, J.; Provencher, J.; Juurlink, D. N.; Blais, R.

2014-01-01

Context Calcium channel blocker poisoning is a common and sometimes life-threatening ingestion. Objective To evaluate the reported effects of treatments for calcium channel blocker poisoning. The primary outcomes of interest were mortality and hemodynamic parameters. The secondary outcomes included length of stay in hospital, length of stay in intensive care unit, duration of vasopressor use, functional outcomes, and serum calcium channel blocker concentrations. Methods Medline/Ovid, PubMed, EMBASE, Cochrane Library, TOXLINE, International pharmaceutical abstracts, Google Scholar, and the gray literature up to December 31, 2013 were searched without time restriction to identify all types of studies that examined effects of various treatments for calcium channel blocker poisoning for the outcomes of interest. The search strategy included the following Keywords: [calcium channel blockers OR calcium channel antagonist OR calcium channel blocking agent OR (amlodipine or bencyclane or bepridil or cinnarizine or felodipine or fendiline or flunarizine or gallopamil or isradipine or lidoflazine or mibefradil or nicardipine or nifedipine or nimodipine or nisoldipine or nitrendipine or prenylamine or verapamil or diltiazem)] AND [overdose OR medication errors OR poisoning OR intoxication OR toxicity OR adverse effect]. Two reviewers independently selected studies and a group of reviewers abstracted all relevant data using a pilot-tested form. A second group analyzed the risk of bias and overall quality using the STROBE (STrengthening the Reporting of OBservational studies in Epidemiology) checklist and the Thomas tool for observational studies, the Institute of Health Economics tool for Quality of Case Series, the ARRIVE (Animal Research: Reporting In Vivo Experiments) guidelines, and the modified NRCNA (National Research Council for the National Academies) list for animal studies. Qualitative synthesis was used to summarize the evidence. Of 15,577 citations identified in the initial search, 216 were selected for analysis, including 117 case reports. The kappa on the quality analysis tools was greater than 0.80 for all study types. Results The only observational study in humans examined high-dose insulin and extracorporeal life support. The risk of bias across studies was high for all interventions and moderate to high for extracorporeal life support. High-dose insulin. High-dose insulin (bolus of 1 unit/kg followed by an infusion of 0.5–2.0 units/kg/h) was associated with improved hemodynamic parameters and lower mortality, at the risks of hypoglycemia and hypokalemia (low quality of evidence). Extracorporeal life support. Extracorporeal life support was associated with improved survival in patients with severe shock or cardiac arrest at the cost of limb ischemia, thrombosis, and bleeding (low quality of evidence). Calcium, dopamine, and norepinephrine. These agents improved hemodynamic parameters and survival without documented severe side effects (very low quality of evidence). 4-Aminopyridine. Use of 4-aminopyridine was associated with improved hemodynamic parameters and survival in animal studies, at the risk of seizures. Lipid emulsion therapy. Lipid emulsion was associated with improved hemodynamic parameters and survival in animal models of intravenous verapamil poisoning, but not in models of oral verapamil poisoning. Other studies. Studies on decontamination, atropine, glucagon, pacemakers, levosimendan, and plasma exchange reported variable results, and the methodologies used limit their interpretation. No trial was documented in humans poisoned with calcium channel blockers for Bay K8644, CGP 28932, digoxin, cyclodextrin, liposomes, bicarbonate, carnitine, fructose 1,6-diphosphate, PK 11195, or triiodothyronine. Case reports were only found for charcoal hemoperfusion, dialysis, intra-aortic balloon pump, Impella device and methylene blue. Conclusions The treatment for calcium channel blocker poisoning is supported by low-quality evidence drawn from a heterogeneous and heavily biased literature. High-dose insulin and extracorporeal life support were the interventions supported by the strongest evidence, although the evidence is of low quality. PMID:25283255

Effect of methanolic extract of Musa sapientum leaves on gastrointestinal transit time in normal and alloxan induced diabetic rats: possible mechanism of action.

PubMed

Adewoye, E O; Ige, A O; Latona, C T

2011-11-23

Disorders of gastrointestinal motility have been associated with diabetes mellitus. Hyperglycaemia particularly has been reported to inhibit gastrointestinal transit time while glibenclamide, a sulphonylurea and insulin, both increased transit time. Musa sapientum has also been reported as an antidiabetic agent but there is dearth of information on the effect of this plant on gastrointestinal motility. This study was therefore carried out to investigate the effect of methanolic extract of Musa sapientum leaves (MEMSL) on gastrointestinal transit time (GITT) in male albino rats with and without hyperglycaemia and to elucidate possible mechanism by which this extract functions. Fifty five albino rats were divided into 11 groups of five animals each. All animals were fasted for 24hrs before the begining of the experiment. Group 1 served as control; while the remaining groups (2 - 11) were treated with 250mg/kg; 500mg/kg MEMSL; diabetic control; diabetic treated with 250mg/kg; 500mg/kg MEMSL; diabetic treated with glibenclamide (5mg/kg); normal rats treated with nifedipine (50mg/kg); normal rats treated with calcium chloride (CaCl2) only (10mg/kg); groups 10 and 11 were both pretreated with CaCl2 and subsequently treated with 250mg/kg and 500mg/kg MEMSL respectively. All plant extracts used for treatments were dissolved in normal saline and administered orally using orogastric tube. Charcoal meal was used as marker in the estimation of GITT. The study showed significant decrease in GITT in the normal rats treated with 250mg/kg and 500mg/kg of extract. However, in the diabetic rats treated with 500mg/kg MEMSL, there was significant increase in GITT and this is comparable with the gut response to glibenclamide (5mg/kg). Musa sapientum extract produced significant decrease in transit time in the calcium chloride pre-treated normal rats and this is comparable to the effect observed in Nifedipine treated group. The significant reduction in GITT produced by MEMSL in the normal rats reflects a strong possibility of MEMSL acting as calcium channel antagonist through the voltage gated calcium channel which may be due to the presence of alkaloids, saponins, cardenolides. There is the possibility of the extract acting as an inhibitor of potassium channel at higher concentration as observed in glibenclamide treated groups.

Neurotrophic Effect of Citrus 5-Hydroxy-3,6,7,8,3′,4′-Hexamethoxyflavone: Promotion of Neurite Outgrowth via cAMP/PKA/CREB Pathway in PC12 Cells

PubMed Central

Lai, Hui-Chi; Wu, Ming-Jiuan; Chen, Pei-Yi; Sheu, Ting-Ting; Chiu, Szu-Ping; Lin, Meng-Han; Ho, Chi-Tang; Yen, Jui-Hung

2011-01-01

5-Hydroxy-3,6,7,8,3′,4′-hexamethoxyflavone (5-OH-HxMF), a hydroxylated polymethoxyflavone, is found exclusively in the Citrus genus, particularly in the peels of sweet orange. In this research, we report the first investigation of the neurotrophic effects and mechanism of 5-OH-HxMF in PC12 pheochromocytoma cells. We found that 5-OH-HxMF can effectively induce PC12 neurite outgrowth accompanied with the expression of neuronal differentiation marker protein growth-associated protein-43(GAP-43). 5-OH-HxMF caused the enhancement of cyclic AMP response element binding protein (CREB) phosphorylation, c-fos gene expression and CRE-mediated transcription, which was inhibited by 2-naphthol AS-E phosphate (KG-501), a specific antagonist for the CREB-CBP complex formation. Moreover, 5-OH-HxMF-induced both CRE transcription activity and neurite outgrowth were inhibited by adenylate cyclase and protein kinase A (PKA) inhibitor, but not MEK1/2, protein kinase C (PKC), phosphatidylinositol 3-kinase (PI3K) or calcium/calmodulin-dependent protein kinase (CaMK) inhibitor. Consistently, 5-OH-HxMF treatment increased the intracellular cAMP level and downstream component, PKA activity. We also found that addition of K252a, a TrKA antagonist, significantly inhibited NGF- but not 5-OH-HxMF-induced neurite outgrowth. These results reveal for the first time that 5-OH-HxMF is an effective neurotrophic agent and its effect is mainly through a cAMP/PKA-dependent, but TrKA-independent, signaling pathway coupling with CRE-mediated gene transcription. A PKC-dependent and CREB-independent pathway was also involved in its neurotrophic action. PMID:22140566

Dronedarone: an amiodarone analogue.

PubMed

Doggrell, Sheila A; Hancox, Jules C

2004-04-01

Of the antiarrhythmic drugs in current use, amiodarone is one of the most effective and is associated with a comparatively low risk of drug-induced pro-arrhythmia, probably due to its multiple pharmacological actions on cardiac ion channels and receptors. However, amiodarone is associated with significant extra-cardiac side effects and this has driven development of amiodarone analogues. These analogues include short acting analogues (e.g., AT-2001) with similar acute effects to amiodarone, the thyroid receptor antagonist KB-130015 and dronedarone. Dronedarone, (SR-33589; Sanofi-Synthelabo), is a non-iodinated amiodarone derivative that inhibits Na +, K + and Ca 2+ currents. It is a potent inhibitor of the acetylcholine-activated K + current from atrial and sinoatrial nodal tissue, and inhibits the rapid delayed rectifier more potently than slow and inward rectifier K + currents and inhibits L-type calcium current. Dronedarone is an antagonist at alpha- and beta-adrenoceptors and unlike amiodarone, has little effect at thyroid receptors. Dronedarone is more potent than amiodarone in inhibiting arrhythmias and death in animal models of ischaemia- and reperfusion-induced arrhythmias. In the Dronedarone Atrial Fibrillation Study After Electrical Cardioversion (DAFNE) clinical trial, dronedarone 800 mg/day appeared to be effective and safe for the prevention of atrial fibrillation relapses after cardioversion. The Antiarrhythmic Trial with Dronedarone in Moderate-to-Severe Congestive Heart Failure Evaluating Morbidity Decrease (ANDROMEDA) trial was stopped due to a potential increased risk of death in the dronedarone group. Trials of dronedarone in the maintenance of sinus rhythm in patients with atrial fibrillation and a safety and tolerability study in patients with an implantable cardioverter defibrillator are ongoing. Further experimental and clinical studies are required before we have a definitive answer to whether dronedarone has advantages over amiodarone and other amiodarone analogues.

Delay of iris flower senescence by cytokinins and jasmonates.

PubMed

van Doorn, Wouter G; Çelikel, Fisun G; Pak, Caroline; Harkema, Harmannus

2013-05-01

It is not known whether tepal senescence in Iris flowers is regulated by hormones. We applied hormones and hormone inhibitors to cut flowers and isolated tepals of Iris × hollandica cv. Blue Magic. Treatments with ethylene or ethylene antagonists indicated lack of ethylene involvement. Auxins or auxin inhibitors also did not change the time to senescence. Abscisic acid (ABA) hastened senescence, but an inhibitor of ABA synthesis (norflurazon) had no effect. Gibberellic acid (GA3 ) slightly delayed senescence in some experiments, but in other experiments it was without effect, and gibberellin inhibitors [ancymidol or 4-hydroxy-5-isopropyl-2-methylphenyltrimethyl ammonium chloride-1-piperidine carboxylate (AMO-1618)] were ineffective as well. Salicylic acid (SA) also had no effect. Ethylene, auxins, GA3 and SA affected flower opening, therefore did reach the flower cells. Jasmonates delayed senescence by about 2.0 days. Similarly, cytokinins delayed senescence by about 1.5-2.0 days. Antagonists of the phosphatidylinositol signal transduction pathway (lithium), calcium channels (niguldipine and verapamil), calmodulin action [fluphenazine, trifluoroperazine, phenoxybenzamide and N-(6-aminohexyl)-5-chloro-1-naphtalenesulfonamide hydrochloride (W-7)] or protein kinase activity [1-(5-isoquinolinesulfonyl)-2-methylpiperazine hydrochloride (H-7), N-[2-(methylamino)ethyl]-5-isoquinolinesulfonamide hydrochloride (H-8) and N-(2-aminoethyl)-5-isoquinolinesulfonamide dihydrochloride (H-9)] had no effect on senescence, indicating no role of a few common signal transduction pathways relating to hormone effects on senescence. The results indicate that tepal senescence in Iris cv. Blue Magic is not regulated by endogenous ethylene, auxin, gibberellins or SA. A role of ABA can at present not be excluded. The data suggest the hypothesis that cytokinins and jasmonates are among the natural regulators. Copyright © Physiologia Plantarum 2012.

Modafinil attenuates reinstatement of cocaine seeking: role for cystine-glutamate exchange and metabotropic glutamate receptors.

PubMed

Mahler, Stephen V; Hensley-Simon, Megan; Tahsili-Fahadan, Pouya; LaLumiere, Ryan T; Thomas, Charles; Fallon, Rebecca V; Kalivas, Peter W; Aston-Jones, Gary

2014-01-01

Modafinil may be useful for treating stimulant abuse, but the mechanisms by which it acts to do so are unknown. Indeed, a primary effect of modafinil is to inhibit dopamine transport, which typically promotes rather than inhibits motivated behavior. Therefore, we examined the role of nucleus accumbens extracellular glutamate and the group II metabotropic glutamate receptor (mGluR2/3) in modafinil effects. One group of rats was trained to self-administer cocaine for 10 days and extinguished, then given priming injections of cocaine to elicit reinstatement. Modafinil (300 mg/kg, intraperitoneal) inhibited reinstated cocaine seeking (but did not alter extinction responding by itself), and this effect was prevented by pre-treatment with bilateral microinjections of the mGluR2/3 antagonist LY-341495 (LY) into nucleus accumbens core. No reversal of modafinil effects was seen after unilateral accumbens core LY, or bilateral LY in the rostral pole of accumbens. Next, we sought to explore effects of modafinil on extracellular glutamate levels in accumbens after chronic cocaine. Separate rats were administered non-contingent cocaine, and after 3 weeks of withdrawal underwent accumbens microdialysis. Modafinil increased extracellular accumbens glutamate in chronic cocaine, but not chronic saline-pre-treated animals. This increase was prevented by reverse dialysis of cystine-glutamate exchange or voltage-dependent calcium channel antagonists. Voltage-dependent sodium channel blockade partly attenuated the increase in glutamate, but mGluR1 blockade did not. We conclude that modafinil increases extracellular glutamate in nucleus accumbens from glial and neuronal sources in cocaine-exposed rats, which may be important for its mGluR2/3-mediated antirelapse properties. © 2012 The Authors, Addiction Biology © 2012 Society for the Study of Addiction.

Pharmacological modulation of cortical excitability shifts induced by transcranial direct current stimulation in humans.

PubMed

Nitsche, M A; Fricke, K; Henschke, U; Schlitterlau, A; Liebetanz, D; Lang, N; Henning, S; Tergau, F; Paulus, W

2003-11-15

Transcranial direct current stimulation (tDCS) of the human motor cortex results in polarity-specific shifts of cortical excitability during and after stimulation. Anodal tDCS enhances and cathodal stimulation reduces excitability. Animal experiments have demonstrated that the effect of anodal tDCS is caused by neuronal depolarisation, while cathodal tDCS hyperpolarises cortical neurones. However, not much is known about the ion channels and receptors involved in these effects. Thus, the impact of the sodium channel blocker carbamazepine, the calcium channel blocker flunarizine and the NMDA receptor antagonist dextromethorphane on tDCS-elicited motor cortical excitability changes of healthy human subjects were tested. tDCS-protocols inducing excitability alterations (1) only during tDCS and (2) eliciting long-lasting after-effects were applied after drug administration. Carbamazepine selectively eliminated the excitability enhancement induced by anodal stimulation during and after tDCS. Flunarizine resulted in similar changes. Antagonising NMDA receptors did not alter current-generated excitability changes during a short stimulation, which elicits no after-effects, but prevented the induction of long-lasting after-effects independent of their direction. These results suggest that, like in other animals, cortical excitability shifts induced during tDCS in humans also depend on membrane polarisation, thus modulating the conductance of sodium and calcium channels. Moreover, they suggest that the after-effects may be NMDA receptor dependent. Since NMDA receptors are involved in neuroplastic changes, the results suggest a possible application of tDCS in the modulation or induction of these processes in a clinical setting. The selective elimination of tDCS-driven excitability enhancements by carbamazepine proposes a role for this drug in focussing the effects of cathodal tDCS, which may have important future clinical applications.

Pharmacological modulation of cortical excitability shifts induced by transcranial direct current stimulation in humans

PubMed Central

Nitsche, M A; Fricke, K; Henschke, U; Schlitterlau, A; Liebetanz, D; Lang, N; Henning, S; Tergau, F; Paulus, W

2003-01-01

Transcranial direct current stimulation (tDCS) of the human motor cortex results in polarity-specific shifts of cortical excitability during and after stimulation. Anodal tDCS enhances and cathodal stimulation reduces excitability. Animal experiments have demonstrated that the effect of anodal tDCS is caused by neuronal depolarisation, while cathodal tDCS hyperpolarises cortical neurones. However, not much is known about the ion channels and receptors involved in these effects. Thus, the impact of the sodium channel blocker carbamazepine, the calcium channel blocker flunarizine and the NMDA receptor antagonist dextromethorphane on tDCS-elicited motor cortical excitability changes of healthy human subjects were tested. tDCS-protocols inducing excitability alterations (1) only during tDCS and (2) eliciting long-lasting after-effects were applied after drug administration. Carbamazepine selectively eliminated the excitability enhancement induced by anodal stimulation during and after tDCS. Flunarizine resulted in similar changes. Antagonising NMDA receptors did not alter current-generated excitability changes during a short stimulation, which elicits no after-effects, but prevented the induction of long-lasting after-effects independent of their direction. These results suggest that, like in other animals, cortical excitability shifts induced during tDCS in humans also depend on membrane polarisation, thus modulating the conductance of sodium and calcium channels. Moreover, they suggest that the after-effects may be NMDA receptor dependent. Since NMDA receptors are involved in neuroplastic changes, the results suggest a possible application of tDCS in the modulation or induction of these processes in a clinical setting. The selective elimination of tDCS-driven excitability enhancements by carbamazepine proposes a role for this drug in focussing the effects of cathodal tDCS, which may have important future clinical applications. PMID:12949224

Management of preterm labor: atosiban or nifedipine?

PubMed

de Heus, Roel; Mulder, Eduard J H; Visser, Gerard H A

2010-08-09

Preterm birth is strongly associated with neonatal death and long-term neurological morbidity. The purpose of tocolytic drug administration is to postpone threatening preterm delivery for 48 hours to allow maximal effect of antenatal corticosteroids and maternal transportation to a center with specialized neonatal care facilities. There is uncertainty about the value of atosiban (oxytocin receptor antagonist) and nifedipine (calcium channel blocker) as first-line tocolytic drugs in the management of preterm labor. For nifedipine, concerns have been raised about unproven safety, lack of placebo-controlled trials, and its off-label use. The tocolytic efficacy of atosiban has also been questioned because of a lack of reduction in neonatal morbidity. This review discusses the available evidence, the pros and cons of either drug and aims to provide information to support a balanced choice of first-line tocolytic drug: atosiban or nifedipine?

The results of modipin monotherapy in coronary heart disease patients with arterial hypertension (secondary coronary prevention).

PubMed

Kapanadze, S; Dolidze, N; Bakhutashvili, Z; Latsabidze, N; Chapidze, L

2005-01-01

The goal of the present study was to evaluate the safety and efficacy of the third generation calcium antagonist -- modipin (amlodipin, Asfarma, Turkey) in 29 patients with coronary atherosclerosis and arterial hypertension. In addition, some pleiotropic actions were examined. On the background of 5-10 mg/day modipin monotherapy during the 3-month study period the target systolic and diastolic blood pressure were achieved in 64 and 51% of cases. Modipin revealed some antiatherogenic efficacy as well. Pleiotropic effects of the drug were particularly expressed in restoring endothelial function reducing degree of hyperlipoperoxidemia and inhibition of platelet aggregation. There were positive changes in functional class of angina. Clinical safety was good. Consequently the present trial supports the use of modipine in all coronary artery disease patients with moderate or severe arterial hypertension.

[Traffic-related PM2.5 regulates IL-2 releasing in Jurkat T cells by calcium signaling pathway].

PubMed

Tong, Guoqiang; Zhang, Zhihong; Han, Jianbiao; Qiu, Yong; Xu, Jianjun

2013-09-01

To explore the effects of traffic-related PM2.5 on interleukin-2 (IL-2) in Jurkat T cells and the regulatory action of calcium signaling pathway. The cells were exposed to 100 microg/ml of PM2.5 for 3, 6 and 24 h. Normal saline group, blank filter group, calcium chelating agent EGTA group and the calcineurin antagonist cyclosporine A (CSA) group were as parallel control. The level of IL-2 was detected by ELISA kits, the mRNA expression of CaN, NFAT were determined by QRT-PCR. The nuclear distribution of NFAT was observed by immunofluorescence microscopy. The level of IL-2 in Jurkat T cells exposed to 100 microg/ml PM2.5 was significantly lower than parallel groups, but higher than PM2.5 + CSA group and PM2.5 + EGTA group (P < 0.05). With the increase of time, the releasing level of IL-2 appeared reducing trend in 100 microg/ml of PM2.5 group. The mRNA expression level of NFAT and CaN were higher than parallel groups, PM2.5 + CSA group and PM2.5 + EGTA group (P < 0.05). PM2.5 can induce NFAT protein with dephosphorylation and be activated, and NFAT protein can shift into nuclear. The level of IL-2 was negatively associated with the expression level of NFAT and CaN gene (P < 0.05). Traffic-related PM2.5 may inhibit the releasing of IL-2, Ca(2+)-CaN-NFAT signal pathway may involve in the regulation of IL-2.

Pathogen variation and urea influence selection and success of Streptomyces mixtures in biological control.

PubMed

Otto-Hanson, L K; Grabau, Z; Rosen, C; Salomon, C E; Kinkel, L L

2013-01-01

Success in biological control of plant diseases remains inconsistent in the field. A collection of well-characterized Streptomyces antagonists (n = 19 isolates) was tested for their capacities to inhibit pathogenic Streptomyces scabies (n = 15 isolates). There was significant variation among antagonists in ability to inhibit pathogen isolates and among pathogens in their susceptibility to inhibition. Only one antagonist could inhibit all pathogens, and antagonist-pathogen interactions were highly specific, highlighting the limitations of single-strain inoculum in biological control. However, the collection of pathogens could be inhibited by several combinations of antagonists, suggesting the potential for successful antagonist mixtures. Urea generally increased effectiveness of antagonists at inhibiting pathogens in vitro (increased mean inhibition zones) but its specific effects varied among antagonist-pathogen combinations. In greenhouse trials, urea enhanced the effectiveness of antagonist mixtures relative to individual antagonists in controlling potato scab. Although antagonist mixtures were frequently antagonistic in the absence of urea, all n= 2 and n = 3 antagonist-isolate combinations were synergistic in the presence of urea. This work provides insights into the efficacy of single- versus multiple-strain inocula in biological control and on the potential for nutrients to influence mixture success.

The Target Residence Time of Antihistamines Determines Their Antagonism of the G Protein-Coupled Histamine H1 Receptor

PubMed Central

Bosma, Reggie; Witt, Gesa; Vaas, Lea A. I.; Josimovic, Ivana; Gribbon, Philip; Vischer, Henry F.; Gul, Sheraz; Leurs, Rob

2017-01-01

The pharmacodynamics of drug-candidates is often optimized by metrics that describe target binding (Kd or Ki value) or target modulation (IC50). However, these metrics are determined at equilibrium conditions, and consequently information regarding the onset and offset of target engagement and modulation is lost. Drug-target residence time is a measure for the lifetime of the drug-target complex, which has recently been receiving considerable interest, as target residence time is shown to have prognostic value for the in vivo efficacy of several drugs. In this study, we have investigated the relation between the increased residence time of antihistamines at the histamine H1 receptor (H1R) and the duration of effective target-inhibition by these antagonists. Hela cells, endogenously expressing low levels of the H1R, were incubated with a series of antihistamines and dissociation was initiated by washing away the unbound antihistamines. Using a calcium-sensitive fluorescent dye and a label free, dynamic mass redistribution based assay, functional recovery of the H1R responsiveness was measured by stimulating the cells with histamine over time, and the recovery was quantified as the receptor recovery time. Using these assays, we determined that the receptor recovery time for a set of antihistamines differed more than 40-fold and was highly correlated to their H1R residence times, as determined with competitive radioligand binding experiments to the H1R in a cell homogenate. Thus, the receptor recovery time is proposed as a cell-based and physiologically relevant metric for the lead optimization of G protein-coupled receptor antagonists, like the H1R antagonists. Both, label-free or real-time, classical signaling assays allow an efficient and physiologically relevant determination of kinetic properties of drug molecules. PMID:29033838

Odontoblasts as sensory receptors: transient receptor potential channels, pannexin-1, and ionotropic ATP receptors mediate intercellular odontoblast-neuron signal transduction.

PubMed

Shibukawa, Yoshiyuki; Sato, Masaki; Kimura, Maki; Sobhan, Ubaidus; Shimada, Miyuki; Nishiyama, Akihiro; Kawaguchi, Aya; Soya, Manabu; Kuroda, Hidetaka; Katakura, Akira; Ichinohe, Tatsuya; Tazaki, Masakazu

2015-04-01

Various stimuli induce pain when applied to the surface of exposed dentin. However, the mechanisms underlying dentinal pain remain unclear. We investigated intercellular signal transduction between odontoblasts and trigeminal ganglion (TG) neurons following direct mechanical stimulation of odontoblasts. Mechanical stimulation of single odontoblasts increased the intracellular free calcium concentration ([Ca(2+)]i) by activating the mechanosensitive-transient receptor potential (TRP) channels TRPV1, TRPV2, TRPV4, and TRPA1, but not TRPM8 channels. In cocultures of odontoblasts and TG neurons, increases in [Ca(2+)]i were observed not only in mechanically stimulated odontoblasts, but also in neighboring odontoblasts and TG neurons. These increases in [Ca(2+)]i were abolished in the absence of extracellular Ca(2+) and in the presence of mechanosensitive TRP channel antagonists. A pannexin-1 (ATP-permeable channel) inhibitor and ATP-degrading enzyme abolished the increases in [Ca(2+)]i in neighboring odontoblasts and TG neurons, but not in the stimulated odontoblasts. G-protein-coupled P2Y nucleotide receptor antagonists also inhibited the increases in [Ca(2+)]i. An ionotropic ATP (P2X3) receptor antagonist inhibited the increase in [Ca(2+)]i in neighboring TG neurons, but not in stimulated or neighboring odontoblasts. During mechanical stimulation of single odontoblasts, a connexin-43 blocker did not have any effects on the [Ca(2+)]i responses observed in any of the cells. These results indicate that ATP, released from mechanically stimulated odontoblasts via pannexin-1 in response to TRP channel activation, transmits a signal to P2X3 receptors on TG neurons. We suggest that odontoblasts are sensory receptor cells and that ATP released from odontoblasts functions as a neurotransmitter in the sensory transduction sequence for dentinal pain.

Memantine alters striatal plasticity inducing a shift of synaptic responses toward long-term depression.

PubMed

Mancini, Maria; Ghiglieri, Veronica; Bagetta, Vincenza; Pendolino, Valentina; Vannelli, Anna; Cacace, Fabrizio; Mineo, Desireé; Calabresi, Paolo; Picconi, Barbara

2016-02-01

Memantine is an open channel blocker that antagonizes NMDA receptors reducing the inappropriate calcium (Ca(2+)) influx occurring in presence of moderately increased glutamate levels. At the same time, memantine has the ability to preserve the transient physiological activation of NMDA receptor, essential for learning and memory formation at synaptic level. In the present study we investigated the effects exerted by memantine on striatal synaptic plasticity in rat striatal spiny projection neurons (SPNs). In vitro application of memantine in striatal slices elicited a disruption of long-term potentiation (LTP) induction and maintenance, and revealed, in the majority of the recorded neurons, a long-term depression (LTD), whose amplitude was concentration-dependent (0.3-10 μM). Interestingly, preincubation with the dopamine (DA) D2 receptor antagonist sulpiride (10 μM) prevented memantine-induced LTD and restored LTP. Moreover, the DA D2 agonist quinpirole (10 μM), similarly to memantine, induced LTD in a subgroup of SPNs. In addition, memantine-induced LTD was also prevented by the CB1 endocannabinoid receptor antagonist AM 251 (1 μM). These results suggest that the actions exerted by memantine on striatal synaptic plasticity, and in particular the induction of LTD observed in SPNs, could be attributed to its ability to activate DA D2 receptors. By contrast, blockade of NMDA receptor is not involved in memantine-induced LTD since APV (30 μM) and MK801 (10 μM), two NMDA receptor antagonists, failed to induce this form of synaptic plasticity. Our data indicate that memantine could be used as treatment of neurological disorders in which DA D2 receptor represents a possible therapeutic target. Copyright © 2015 Elsevier Ltd. All rights reserved.

Sigma-1 receptor agonist increases axon outgrowth of hippocampal neurons via voltage-gated calcium ions channels.

PubMed

Li, Dong; Zhang, Shu-Zhuo; Yao, Yu-Hong; Xiang, Yun; Ma, Xiao-Yun; Wei, Xiao-Li; Yan, Hai-Tao; Liu, Xiao-Yan

2017-12-01

Sigma-1 receptors (Sig-1Rs) are unique endoplasmic reticulum proteins that have been implicated in both neurodegenerative and ischemic diseases, such as Alzheimer's disease and stroke. Accumulating evidence has suggested that Sig-1R plays a role in neuroprotection and axon outgrowth. The underlying mechanisms of Sig-1R-mediated neuroprotection have been well elucidated. However, the mechanisms underlying the effects of Sig-1R on axon outgrowth are not fully understood. To clarify this issue, we utilized immunofluorescence to compare the axon lengths of cultured naïve hippocampal neurons before and after the application of the Sig-1R agonist, SA4503. Then, electrophysiology and immunofluorescence were used to examine voltage-gated calcium ion channel (VGCCs) currents in the cell membranes and growth cones. We found that Sig-1R activation dramatically enhanced the axonal length of the naïve hippocampal neurons. Application of the Sig-1R antagonist NE100 and gene knockdown techniques both demonstrated the effects of Sig-1R. The growth-promoting effect of SA4503 was accompanied by the inhibition of voltage-gated Ca 2+ influx and was recapitulated by incubating the neurons with the L-type, N-type, and P/Q-type VGCC blockers, nimodipine, MVIIA and ω-agatoxin IVA, respectively. This effect was unrelated to glial cells. The application of SA4503 transformed the growth cone morphologies from complicated to simple, which favored axon outgrowth. Sig-1R activation can enhance axon outgrowth and may have a substantial influence on neurogenesis and neurodegenerative diseases. © 2017 John Wiley & Sons Ltd.

Protective effects of AT1-receptor blocker and CA antagonist combination on renal function in salt loaded spontaneously hypertensive rats.

PubMed

Gjorgjievska, K; Zafirov, D; Jurhar-Pavlova, M; Cekovska, S; Atanasovska, E; Pavlovska, K; Zendelovska, D

2015-01-01

Salt sensitive hypertension is known to be a contributing factor for the progression of kidney disease. This study was undertaken to investigate the role of excessive dietary salt on renal function and to evaluate the effect of valsartan and amlodipin given as a combination therapy on blood pressure and parameters specific to the renal function in salt loaded SHR rats. 48 male SHR rats at age of 20 weeks and body weight ranging between 270-350 g were used. SHR rats were divided into 3 groups: control group of rats -SHRC (n = 16) given tab water ad libitum and two salt treated groups in which tab water was replaced with a solution of NaCl (1%) from age of 8 weeks given ad libitum: SHRVAL+AMLO group (n = 16) where investigated drugs were administered at a dose of 10 mg/kg/ b.w. (valsartan) and 5 mg/kg/ b.w. (amlodipin) by gavage and SHR NaCl group (n = 16) that received saline in the same volume and the same time intervals as the SHRVAL+AMLO group. For a period of 12 weeks we have investigated the effect of the VAL+AMLO drug combination on systolic blood pressure (SBP), body weight and renal function tests. Salt loading with 1% solution in the SHR NaCl group has lead to significant increase of blood pressure, proteinuria and decrease in creatinine clearance. Combined treatment with AT1 receptor blocker and calcium antagonist has managed to control blood pressure and ameliorated renal damage.

Renal effects of felodipine: a review of experimental evidence and clinical data.

PubMed

DiBona, G F

1990-01-01

The dihydropyridine calcium channel antagonist felodipine has a wide spectrum of effects on the kidney. From a variety of studies in normotensive and hypertensive animals and human subjects, felodipine produces a decrease in renal vascular resistance that, although predominantly dependent on the decrease in mean arterial pressure (MAP), may be associated with an increase in renal blood flow (RBF). The glomerular filtration rate (GFR) is unchanged. In response to acute felodipine administration, the significant diuresis and natriuresis observed is caused by a direct inhibitory effect on net renal tubular sodium and water reabsorption. While the acute natriuretic response to felodipine administration is modulated by compensatory adaptations over the remainder of the 24-h period and during chronic treatment, the negative sodium balance established is sustained over the duration of the treatment. Renal sodium and water retention are not observed and there is little effect on renal potassium handling. As a vasodilator antihypertensive agent, felodipine produces renal vasodilatation (normal or increased but not decreased RBF) without adverse effects on the GFR or renal sodium and water retention.

Delineating unique cellular responses to PDT (Invited paper)

NASA Astrophysics Data System (ADS)

Kessel, David

2005-04-01

Photodamage to mitochondria, endoplasmic reticulum (ER) or lysosomes can lead to activation of the apoptotic program, as can exposure of cells to the non-peptidic Bcl-2/Bcl-xL antagonist HA14-1. Many signaling pathways are evoked by photodynamic therapy (PDT), presumably from oxidative stress effects. To discover which of the latter effects might be unique to PDT, we compared some photodynamic effects with HA14-1 treatment, using murine leukemia L1210 cells in culture. Two photosensitizers were employed: the porphycene CPO and the chlorin NPe6. The former targets the endoplasmic reticulum (ER) and causes Bcl-2 photodamage, while NPe6 targets lysosomes, resulting in protease-induced cleavage and activation of Bid to form the pro-apoptotic product t-Bid. PDT at either target will lead to loss of the mitochondrial membrane potential ΔΨm, translocation of cytochrome c to the cytosol and an apoptotic response. Photodynamic effects of CPO or NPe6 led to activation of several 'stress proteins' and intracellular oxidation of the probe dihydrodichlorofluorescein (H2DCF). All of these effects were mimicked by HA14-1, indicating that these early responses to PDT result from initiation of apoptosis, however achieved. After CPO-catalyzed PDT or HA14-1 treatment, we observed a prompt release of Ca2+ into the cytosol, but this was insufficient to significantly alter mitochondrial calcium levels. The apoptotic response to HA14-1 or Bcl-2 photodamage was markedly promoted by the protein kinase C (PKC) inhibitor staurosporin (STS). These effects were not observed after photodamage catalyzed by NPe6, indicating that calcium release and PKC interactions are associated with loss of Bcl-2 function, but not Bid activation.

Occupation of low-affinity cholecystokinin (CCK) receptors by CCK activates signal transduction and stimulates amylase secretion in pancreatic acinar cells.

PubMed

Vinayek, R; Patto, R J; Menozzi, D; Gregory, J; Mrozinski, J E; Jensen, R T; Gardner, J D

1993-03-10

Based on the effects of monensin on binding of 125I-CCK-8 and its lack of effect on CCK-8-stimulated amylase secretion we previously proposed that pancreatic acinar cells possess three classes of CCK receptors: high-affinity receptors, low-affinity receptors and very low-affinity receptors [1]. In the present study we treated pancreatic acini with carbachol to induce a complete loss of high-affinity CCK receptors and then examined the action of CCK-8 on inositol trisphosphate IP3(1,4,5), cytosolic calcium and amylase secretion in an effort to confirm and extend our previous hypothesis. We found that first incubating pancreatic acini with 10 mM carbachol decreased binding of 125I-CCK-8 measured during a second incubation by causing a complete loss of high-affinity CCK receptors with no change in the low-affinity CCK receptors. Carbachol treatment of acini, however, did not alter the action of CCK-8 on IP3(1,4,5), cytosolic calcium or amylase secretion or the action of CCK-JMV-180 on amylase secretion or on the supramaximal inhibition of amylase secretion caused by CCK-8. The present findings support our previous hypothesis that pancreatic acinar cells possess three classes of CCK receptors and suggest that high-affinity CCK receptors do not mediate the action of CCK-8 on enzyme secretion, that low-affinity CCK receptors may mediate the action of CCK on cytosolic calcium that does not involve IP3(1,4,5) and produce the upstroke of the dose-response curve for CCK-8-stimulated amylase secretion and that very low-affinity CCK receptors mediate the actions of CCK on IP3(1,4,5) and cytosolic calcium and produce the downstroke of the dose-response curve for CCK-8-stimulated amylase secretion. Moreover, CCK-JMV-180 is a full agonist for stimulating amylase secretion by acting at low-affinity CCK receptors and is an antagonist at very low-affinity CCK receptors.

Histamine up-regulates fibroblast growth factor receptor 1 and increases FOXP2 neurons in cultured neural precursors by histamine type 1 receptor activation: conceivable role of histamine in neurogenesis during cortical development in vivo.

PubMed

Molina-Hernández, Anayansi; Rodríguez-Martínez, Griselda; Escobedo-Ávila, Itzel; Velasco, Iván

2013-03-07

During rat development, histamine (HA) is one of the first neuroactive molecules to appear in the brain, reaching its maximal value at embryonic day 14, a period when neurogenesis of deep layers is occurring in the cerebral cortex, suggesting a role of this amine in neuronal specification. We previously reported, using high-density cerebrocortical neural precursor cultures, that micromolar HA enhanced the effect of fibroblast growth factor (FGF)-2 on proliferation, and that HA increased neuronal differentiation, due to HA type 1 receptor (H(1)R) activation. Clonal experiments performed here showed that HA decreased colony size and caused a significant increase in the percentage of clones containing mature neurons through H(1)R stimulation. In proliferating precursors, we studied whether HA activates G protein-coupled receptors linked to intracellular calcium increases. Neural cells presented an increase in cytoplasmic calcium even in the absence of extracellular calcium, a response mediated by H(1)R. Since FGF receptors (FGFRs) are known to be key players in cell proliferation and differentiation, we determined whether HA modifies the expression of FGFRs1-4 by using RT-PCR. An important transcriptional increase in FGFR1 was elicited after H(1)R activation. We also tested whether HA promotes differentiation specifically to neurons with molecular markers of different cortical layers by immunocytochemistry. HA caused significant increases in cells expressing the deep layer neuronal marker FOXP2; this induction of FOXP2-positive neurons elicited by HA was blocked by the H(1)R antagonist chlorpheniramine in vitro. Finally, we found a notable decrease in FOXP2+ cortical neurons in vivo, when chlorpheniramine was infused in the cerebral ventricles through intrauterine injection. These results show that HA, by activating H(1)R, has a neurogenic effect in clonal conditions and suggest that intracellular calcium elevation and transcriptional up-regulation of FGFR1 participate in HA-induced neuronal differentiation to FOXP2 cells in vitro; furthermore, H(1)R blockade in vivo resulted in decreased cortical FOXP2+ neurons.

Narcotic antagonists. Treatment tool for addiction.

PubMed

Valentine, N M; Meyer, R E

1976-09-01

Narcotic antagonists have recently gained attention through research aimed at evaluating both biochemical effects and treatment potential for opiate addiction. Narcotic antagonists are a classification of drugs which block the euphoric (and all other) effects of opiates. Naltrexone is the most promising narcotic antagonist based on ability to produce blockade, length of duration, and relative absence of side effects. The narcotic antagonists offer an adjunctive or alternative method of treatment for opiate addicts based on Wikler's biobehavioral theory of conditioned abstinence. Narcotic antagonists are presently being investigated at seven research centers throughout the United States and may be available for clinical use in the future.

Involvement of the Calcium-sensing Receptor in Human Taste Perception

PubMed Central

Ohsu, Takeaki; Amino, Yusuke; Nagasaki, Hiroaki; Yamanaka, Tomohiko; Takeshita, Sen; Hatanaka, Toshihiro; Maruyama, Yutaka; Miyamura, Naohiro; Eto, Yuzuru

2010-01-01

By human sensory analyses, we found that various extracellular calcium-sensing receptor (CaSR) agonists enhance sweet, salty, and umami tastes, although they have no taste themselves. These characteristics are known as “kokumi taste” and often appear in traditional Japanese cuisine. Although GSH is a typical kokumi taste substance (taste enhancer), its mode of action is poorly understood. Here, we demonstrate how the kokumi taste is enhanced by the CaSR, a close relative of the class C G-protein-coupled receptors T1R1, T1R2, and T1R3 (sweet and umami receptors). We identified a large number of CaSR agonist γ-glutamyl peptides, including GSH (γ-Glu-Cys-Gly) and γ-Glu-Val-Gly, and showed that these peptides elicit the kokumi taste. Further analyses revealed that some known CaSR agonists such as Ca2+, protamine, polylysine, l-histidine, and cinacalcet (a calcium-mimetic drug) also elicit the kokumi taste and that the CaSR-specific antagonist, NPS-2143, significantly suppresses the kokumi taste. This is the first report indicating a distinct function of the CaSR in human taste perception. PMID:19892707

Fenspiride and membrane transduction signals in rat alveolar macrophages.

PubMed

Féray, J C; Mohammadi, K; Taouil, K; Brunet, J; Garay, R P; Hannaert, P

1997-07-15

Fenspiride inhibits the calcium signal evoked by the inflammatory peptide formyl-Met-Leu-Phe (fMLP) in peritoneal macrophages, but at concentrations (approximately 1 mM) far above the therapeutic range (approximately 1 microM). Here, in rat alveolar macrophages, high fenspiride concentrations (1 mM) were required to inhibit the calcium signals evoked by the calcium agonist Bay K8644 or by ionomycin. Moreover, fenspiride (1 mM) was a poor inhibitor of the cell membrane depolarization induced by gramicidine D. By contrast, fenspiride blocked Na+-H+ antiport activation by (i) fMLP with an IC50 = 3.1 +/- 1.9 nM and (ii) PMA (phorbol 12-myristate 13-acetate) with an IC50 = 9.2 +/- 3.1 nM. Finally, protein kinase C (PKC) activity of macrophage homogenate was not significantly modified by 10 or 100 microM fenspiride (at 100 microM: 2.57 +/- 1.60 vs. 2.80 +/- 1.71 pmol/10(6) cells/min). In conclusion, fenspiride inhibits fMLP- and PMA-induced pH signals in rat alveolar macrophages, probably by acting distally on the PKC transduction signal. This pH antagonistic action may be relevant for the antiinflammatory mechanism of fenspiride and requires further investigation.

Effect of a 5-lipoxygenase inhibitor and leukotriene antagonist (PF 5901) on antigen-induced airway responses in neonatally immunized rabbits.

PubMed Central

Herd, C. M.; Donigi-Gale, D.; Shoupe, T. S.; Burroughs, D. A.; Yeadon, M.; Page, C. P.

1994-01-01

1. The effect of a single intratracheal dose (10 mg) of PF 5901 (2-[3(1-hydroxyhexyl) phenoxymethyl] quinoline hydrochloride, a specific inhibitor of the 5-lipoxygenase pathway of arachidonic acid metabolism and a leukotriene D4 antagonist) on airway changes induced in response to Alternaria tenuis aerosol challenge was assessed in adult rabbits neonatally immunized. Leukotriene generation was determined in vivo by measuring leukotriene B4 (LTB4) levels in bronchoalveolar lavage (BAL) fluid and ex vivo by measuring calcium ionophore-stimulated production of LTB4 in whole blood. 2. While PF 5901 (10 mg) had no significant effect on the acute bronchoconstriction induced by antigen, this dose was sufficient to inhibit significantly the increase in airway responsiveness to inhaled histamine 24 h following antigen challenge (P < 0.05). 3. Total leucocyte infiltration into the airways induced by antigen, as assessed by bronchoalveolar lavage, was significantly inhibited by pretreatment with PF 5901 (10 mg). However, the pulmonary infiltration of neutrophils and eosinophils induced by antigen was unaltered by prior treatment with PF 5901 (10 mg). 4. PF 5901 (10 mg) had no effect on ex vivo LTB4 synthesis in whole blood. However, the antigen-induced increase in LTB4 levels in BAL 24 h following challenge was significantly inhibited (P < 0.05). 5. We suggest from the results of the present study that the antigen-induced airway hyperresponsiveness to inhaled histamine in immunized rabbits is mediated, at least in part, by products of the 5-lipoxygenase metabolic pathway, and is not dependent on the extent of eosinophil or neutrophil influx into the airway lumen. PMID:8032653

In Vitro Biologic Activities of the Antimicrobials Triclocarban, Its Analogs, and Triclosan in Bioassay Screens: Receptor-Based Bioassay Screens

PubMed Central

Ahn, Ki Chang; Zhao, Bin; Chen, Jiangang; Cherednichenko, Gennady; Sanmarti, Enio; Denison, Michael S.; Lasley, Bill; Pessah, Isaac N.; Kültz, Dietmar; Chang, Daniel P.Y.; Gee, Shirley J.; Hammock, Bruce D.

2008-01-01

Background Concerns have been raised about the biological and toxicologic effects of the antimicrobials triclocarban (TCC) and triclosan (TCS) in personal care products. Few studies have evaluated their biological activities in mammalian cells to assess their potential for adverse effects. Objectives In this study, we assessed the activity of TCC, its analogs, and TCS in in vitro nuclear-receptor–responsive and calcium signaling bioassays. Materials and methods We determined the biological activities of the compounds in in vitro, cell-based, and nuclear-receptor–responsive bioassays for receptors for aryl hydrocarbon (AhR), estrogen (ER), androgen (AR), and ryanodine (RyR1). Results Some carbanilide compounds, including TCC (1–10 μM), enhanced estradiol (E2)-dependent or testosterone-dependent activation of ER- and AR-responsive gene expression up to 2.5-fold but exhibited little or no agonistic activity alone. Some carbanilides and TCS exhibited weak agonistic and/or antagonistic activity in the AhR-responsive bioassay. TCS exhibited antagonistic activity in both ER- and AR-responsive bioassays. TCS (0.1–10 μM) significantly enhanced the binding of [3H]ryanodine to RyR1 and caused elevation of resting cytosolic [Ca2+] in primary skeletal myotubes, but carbanilides had no effect. Conclusions Carbanilides, including TCC, enhanced hormone-dependent induction of ER- and AR-dependent gene expression but had little agonist activity, suggesting a new mechanism of action of endocrine-disrupting compounds. TCS, structurally similar to noncoplanar ortho-substituted poly-chlorinated biphenyls, exhibited weak AhR activity but interacted with RyR1 and stimulated Ca2+ mobilization. These observations have potential implications for human and animal health. Further investigations are needed into the biological and toxicologic effects of TCC, its analogs, and TCS. PMID:18795164

Neuroprotective effect of noscapine on cerebral oxygen-glucose deprivation injury.

PubMed

Vahabzadeh, Gelareh; Rahbar-Roshandel, Nahid; Ebrahimi, Soltan-Ahmad; Mahmoudian, Massoud

2015-04-01

The present study aims to investigate the effect of noscapine (0.5-2.5 μM), an alkaloid from the opium poppy, on primary murine fetal cortical neurons exposed to oxygen-glucose deprivation (OGD), an in vitro model of ischemia. Cells were transferred to glucose-free DMEM and were exposed to hypoxia in a small anaerobic chamber. Cell viability and nitric oxide production were evaluated by MTT assay and the Griess method, respectively. The neurotoxicities produced by all three hypoxia durations tested were significantly inhibited by 0.5 μM noscapine. Increasing noscapine concentration up to 2.5 μM produced a concentration-dependent inhibition of neurotoxicity. Pretreatment of cells with MK-801 (10 μM), a non-competitive NMDA antagonist, and nimodipine (10nM), an L-type Ca(2+) channel blockers, increased cell viability after 30 min OGD, while the application of NBQX (30 μM), a selective AMPA-kainate receptor antagonist partially attenuated cell injury. Subsequently, cells treated with noscapine in the presence of thapsigargin (1 μM), an inhibitor of endoplasmic reticulum Ca(2+) ATPases. After 60 min OGD, noscapine could inhibit the cell damage induced by thapsigargin. However, noscapine could not reduce cell damage induced by 240 min OGD in the presence of thapsigargin. Noscapine attenuated nitric oxide (NO) production in cortical neurons after 30 min OGD. We concluded that noscapine had a neuroprotective effect, which could be due to its interference with multiple targets in the excitotoxicity process. These effects could be mediated partially by a decrease in NO production and the modulation of intracellular calcium levels. Copyright © 2014 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

Muscarinic modulation of TREK currents in mouse sympathetic superior cervical ganglion neurons.

PubMed

Rivas-Ramírez, P; Cadaveira-Mosquera, A; Lamas, J A; Reboreda, A

2015-07-01

Muscarinic receptors play a key role in the control of neurotransmission in the autonomic ganglia, which has mainly been ascribed to the regulation of potassium M-currents and voltage-dependent calcium currents. Muscarinic agonists provoke depolarization of the membrane potential and a reduction in spike frequency adaptation in postganglionic neurons, effects that may be explained by M-current inhibition. Here, we report the presence of a riluzole-activated current (IRIL ) that flows through the TREK-2 channels, and that is also inhibited by muscarinic agonists in neurons of the mouse superior cervical ganglion (mSCG). The muscarinic agonist oxotremorine-M (Oxo-M) inhibited the IRIL by 50%, an effect that was abolished by pretreatment with atropine or pirenzepine, but was unaffected in the presence of himbacine. Moreover, these antagonists had similar effects on single-channel TREK-2 currents. IRIL inhibition was unaffected by pretreatment with pertussis toxin. The protein kinase C blocker bisindolylmaleimide did not have an effect, and neither did the inositol triphosphate antagonist 2-aminoethoxydiphenylborane. Nevertheless, the IRIL was markedly attenuated by the phospholipase C (PLC) inhibitor ET-18-OCH3. Finally, the phosphatidylinositol-3-kinase/phosphatidylinositol-4-kinase inhibitor wortmannin strongly attenuated the IRIL , whereas blocking phosphatidylinositol 4,5-bisphosphate (PIP2 ) depletion consistently prevented IRIL inhibition by Oxo-M. These results demonstrate that TREK-2 currents in mSCG neurons are inhibited by muscarinic agonists that activate M1 muscarinic receptors, reducing PIP2 levels via a PLC-dependent pathway. The similarities between the signaling pathways regulating the IRIL and the M-current in the same neurons reflect an important role of this new pathway in the control of autonomic ganglia excitability. © 2015 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

Reduction of intimal hyperplasia and enhanced reactivity of experimental vein bypass grafts with verapamil treatment.

PubMed Central

el-Sanadiki, M N; Cross, K S; Murray, J J; Schuman, R W; Mikat, E; McCann, R L; Hagen, P O

1990-01-01

Recent studies have shown that calcium antagonists exert an antiatherogenic effect in animals fed cholesterol. Vein graft intimal hyperplasia is believed to be an early event in atherosclerotic lesion formation, which is a significant cause of graft failure. Altered vasoreactivity has also been postulated in the etiology of vein graft failure. Therefore this study examined the effect of verapamil treatment on the development of intimal hyperplasia and the vasoreactivity of experimental vein bypass grafts. The right external jugular vein was grafted into the right carotid artery of 30 male New Zealand white rabbits fed normal rabbit chow. The left external jugular vein was used as the control vein. Fifteen animals received verapamil (1.25 mg/day for 28 days) via the femoral vein by means of an osmotic pump. In 15 control animals the pump contained saline. Plasma verapamil concentration was 50.9 +/- 13.2 ng/mL (x +/- SEM), a dose that showed no effect on either blood pressure, total serum cholesterol, or in vitro platelet aggregation to ADP. Fourteen of fifteen grafts were patent in each group, for a patency rate of 93%. Histologic examination using computer morphometry showed significant reduction of intimal hyperplasia at the proximal, middle, and distal graft segments (p less than 0.05). In addition in vitro isometric tension studies of the vein grafts and control veins showed that verapamil causes enhanced reactivity of both vein grafts and control veins in response to norepinephrine and histamine (p less than 0.05). Reactivity of vein grafts to serotonin was unaltered. While none of the normal veins in the control group responded to serotonin, normal veins treated with verapamil contracted readily in response to serotonin. Endothelial-dependent relaxation to acetylcholine was absent in both control and verapamil-treated vein grafts, while normal veins from both groups responded to the same extent to acetylcholine. Because we could not demonstrate any difference in platelet or endothelium function between untreated and verapamil-treated animals, we examined the direct effect of verapamil on smooth muscle. Verapamil significantly inhibited [3H]-thymidine incorporation into DNA in vascular smooth muscle cells in culture in a dose-dependent manner. Verapamil treatment significantly reduces intimal hyperplasia in experimental vein grafts and inhibits smooth muscle cell proliferation in culture. Furthermore the enhanced reactivity to norepinephrine and histamine in the verapamil-treated vessels has no detrimental effect on the patency rate at 4 weeks. Thus by inhibiting intimal hyperplasia, calcium antagonists may improve the long-term patency of vein bypass grafts. Images Figs. 1A-C. PMID:2363608

Stimulant effects of adenosine antagonists on operant behavior: differential actions of selective A2A and A1 antagonists

PubMed Central

Randall, Patrick A.; Nunes, Eric J.; Janniere, Simone L.; Stopper, Colin M.; Farrar, Andrew M.; Sager, Thomas N.; Baqi, Younis; Hockemeyer, Jörg; Müller, Christa E.

2012-01-01

Rationale Adenosine A2A antagonists can reverse many of the behavioral effects of dopamine antagonists, including actions on instrumental behavior. However, little is known about the effects of selective adenosine antagonists on operant behavior when these drugs are administered alone. Objective The present studies were undertaken to investigate the potential for rate-dependent stimulant effects of both selective and nonselective adenosine antagonists. Methods Six drugs were tested: two nonselective adenosine antagonists (caffeine and theophylline), two adenosine A1 antagonists (DPCPX and CPT), and two adenosine A2A antagonists (istradefylline (KW6002) and MSX-3). Two schedules of reinforcement were employed; a fixed interval 240-s (FI-240 sec) schedule was used to generate low baseline rates of responding and a fixed ratio 20 (FR20) schedule generated high rates. Results Caffeine and theophylline produced rate-dependent effects on lever pressing, increasing responding on the FI-240 sec schedule but decreasing responding on the FR20 schedule. The A2A antagonists MSX-3 and istradefylline increased FI-240 sec lever pressing but did not suppress FR20 lever pressing in the dose range tested. In fact, there was a tendency for istradefylline to increase FR20 responding at a moderate dose. A1 antagonists failed to increase lever pressing rate, but DPCPX decreased FR20 responding at higher doses. Conclusions These results suggest that adenosine A2A antagonists enhance operant response rates, but A1 antagonists do not. The involvement of adenosine A2A receptors in regulating aspects of instrumental response output and behavioral activation may have implications for the treatment of effort-related psychiatric dysfunctions, such as psychomotor slowing and anergia in depression. PMID:21347642

Changes in the response to excitatory antagonists, agonists, and spasmolytic agents in circular colonic smooth muscle strips from patients with diverticulosis.

PubMed

Alvarez-Berdugo, D; Espín, F; Arenas, C; López, I; Clavé, P; Gallego, D

2015-11-01

Colonic samples from asymptomatic diverticulosis (DS) patients presented enhanced electrical field stimulation (EFS)-contractions, in an earlier study of ours, suggesting increased endogenous responses. The aim of this study was to explore changes in excitatory neuromuscular transmission and to assess the pharmacodynamics of spasmolytic agents in DS. Circular muscle strips from sigmoid colon of DS patients (n = 30; 69.5 ± 14.8 years) and controls (n = 32; 64.7 ± 16.2 years) were studied using organ baths to evaluate the direct effect of excitatory agonists (carbachol, neurokinin A [NKA] and substance P [SP]), and the effect of antagonists (atropine and NK2 antagonist GR94800) and spasmolytic drugs (otilonium bromide [OB] and N-butyl-hyoscine) on the contractions induced by EFS-stimulation of excitatory motorneurons. qRT-PCR was also performed to compare mRNA expression of M2 , M3 , NK2 receptors and L-type calcium channels. Contractions to carbachol (Emax : 663.7 ± 305.6% control vs 2698.0 ± 439.5% DS; p < 0.0005) and NKA (Emax : 387.8 ± 35.6% vs 1102.0 ± 190.1%; p < 0.0005) were higher in DS group, without differences for SP. Higher potency for DS patients was observed in the concentration-response curves for atropine (pIC50  = 8.56 ± 0.15 control vs pIC50  = 9.95 ± 0.18 DS group; p < 0.005) and slightly higher for GR94800 (pIC50  = 7.21 ± 0.18 control vs pIC50  = 7.97 ± 0.32 group; p < 0.0001). Lower efficacy (Emax ) and potency (pIC50 ) was observed for spasmolytic drugs in DS, whereas no differences were found regarding the relative expression of the receptors evaluated between groups. The greater response to cholinergic and tachykinergic agonists and greater potency for muscarinic and NK2 antagonists observed in DS might play a role in the spasticity found in diverticular disease. © 2015 John Wiley & Sons Ltd.

Dopamine modulation of transient receptor potential vanilloid type 1 (TRPV1) receptor in dorsal root ganglia neurons.

PubMed

Chakraborty, Saikat; Rebecchi, Mario; Kaczocha, Martin; Puopolo, Michelino

2016-03-15

The transient receptor potential vanilloid type 1 (TRPV1) receptor plays a key role in the modulation of nociceptor excitability. To address whether dopamine can modulate the activity of TRPV1 channels in nociceptive neurons, the effects of dopamine and dopamine receptor agonists were tested on the capsaicin-activated current recorded from acutely dissociated small diameter (<27 μm) dorsal root ganglia (DRG) neurons. Dopamine or SKF 81297 (an agonist at D1/D5 receptors), caused inhibition of both inward and outward currents by ∼60% and ∼48%, respectively. The effect of SKF 81297 was reversed by SCH 23390 (an antagonist at D1/D5 receptors), confirming that it was mediated by activation of D1/D5 dopamine receptors. In contrast, quinpirole (an agonist at D2 receptors) had no significant effect on the capsaicin-activated current. Inhibition of the capsaicin-activated current by SKF 81297 was mediated by G protein coupled receptors (GPCRs), and highly dependent on external calcium. The inhibitory effect of SKF 81297 on the capsaicin-activated current was not affected when the protein kinase A (PKA) activity was blocked with H89, or when the protein kinase C (PKC) activity was blocked with bisindolylmaleimide II (BIM). In contrast, when the calcium-calmodulin-dependent protein kinase II (CaMKII) was blocked with KN-93, the inhibitory effect of SKF 81297 on the capsaicin-activated current was greatly reduced, suggesting that activation of D1/D5 dopamine receptors may be preferentially linked to CaMKII activity. We suggest that modulation of TRPV1 channels by dopamine in nociceptive neurons may represent a way for dopamine to modulate incoming noxious stimuli. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Dopamine Receptor Activation Reorganizes Neuronal Ensembles during Hippocampal Sharp Waves In Vitro

PubMed Central

Miyawaki, Takeyuki; Norimoto, Hiroaki; Ishikawa, Tomoe; Watanabe, Yusuke; Matsuki, Norio; Ikegaya, Yuji

2014-01-01

Hippocampal sharp wave (SW)/ripple complexes are thought to contribute to memory consolidation. Previous studies suggest that behavioral rewards facilitate SW occurrence in vivo. However, little is known about the precise mechanism underlying this enhancement. Here, we examined the effect of dopaminergic neuromodulation on spontaneously occurring SWs in acute hippocampal slices. Local field potentials were recorded from the CA1 region. A brief (1 min) treatment with dopamine led to a persistent increase in the event frequency and the magnitude of SWs. This effect lasted at least for our recording period of 45 min and did not occur in the presence of a dopamine D1/D5 receptor antagonist. Functional multineuron calcium imaging revealed that dopamine-induced SW augmentation was associated with an enriched repertoire of the firing patterns in SW events, whereas the overall tendency of individual neurons to participate in SWs and the mean number of cells participating in a single SW were maintained. Therefore, dopaminergic activation is likely to reorganize cell assemblies during SWs. PMID:25089705

Effect of a Hypocretin/Orexin Antagonist on Neurocognitive Performance

DTIC Science & Technology

2010-09-30

a novel hypocretiniorexin antagonist, almorexant (ALM), to a standard hypnotic , zolpidem (ZOL), and placebo (PBO) on neurocognitive performance at...Placebo-Controlled, Randomized, Parallel- Group Study Comparing the Effect of a Novel HypocretiniOrexin Antagonist (Almorexant) Versus a Standard Hypnotic ...Group Study Comparing the Effect of a Novel HypocretiniOrexin Antagonist (Almorexant) Versus a Standard Hypnotic (Zolpidem) and Placebo on

EHB1 and AGD12, two calcium-dependent proteins affect gravitropism antagonistically in Arabidopsis thaliana.

PubMed

Dümmer, Michaela; Michalski, Christian; Essen, Lars-Oliver; Rath, Magnus; Galland, Paul; Forreiter, Christoph

2016-11-01

The ADP-RIBOSYLATION FACTOR GTPase-ACTIVATING PROTEIN (AGD) 12, a member of the ARF-GAP protein family, affects gravitropism in Arabidopsis thaliana. A loss-of-function mutant lacking AGD12 displayed diminished gravitropism in roots and hypocotyls indicating that both organs are affected by this regulator. AGD12 is structurally related to ENHANCED BENDING (EHB) 1, previously described as a negative effector of gravitropism. In contrast to agd12 mutants, ehb1 loss-of function seedlings displayed enhanced gravitropic bending. While EHB1 and AGD12 both possess a C-terminal C2/CaLB-domain, EHB1 lacks the N-terminal ARF-GAP domain present in AGD12. Subcellular localization analysis using Brefeldin A indicated that both proteins are elements of the trans Golgi network. Physiological analyses provided evidence that gravitropic signaling might operate via an antagonistic interaction of ARF-GAP (AGD12) and EHB1 in their Ca 2+ -activated states. Copyright © 2016 Elsevier GmbH. All rights reserved.

Treatment of essential hypertension with calcium channel blockers: what is the place of lercanidipine?

PubMed

Burnier, Michel; Pruijm, Menno; Wuerzner, Gregoire

2009-08-01

In all actual clinical guidelines, dihydropyridine calcium channel blockers (CCBs) belong to the recommended first line antihypertensive drugs to treat essential hypertension. Several recent large clinical trials have confirmed their efficacy not only in lowering blood pressure but also in reducing cardiovascular morbidity and mortality in hypertensive patients with a normal or high cardiovascular risk profile. In clinical trials such as ALLHAT, VALUE or ASCOT, an amlodipine-based therapy was at least as effective, when not slightly superior, in lowering blood pressure and sometimes more effective in preventing target organ damages than blood pressure lowering strategies based on the use of diuretics, beta-blockers and blockers of the renin-angiotensin system. One of the main clinical side effects of the first and second generation CCBs including amlodipine is the development of peripheral edema. The incidence of leg edema can be markedly reduced by combining the CCB with a blocker of the renin-angiotensin system. This strategy has now led to the development of several fixed-dose combinations of amlodipine and angiotensin II receptor antagonists. Another alternative to lower the incidence of edema is to use CCBs of the third generation such as lercanidipine. Indeed, although no major clinical trials have been conducted with this compound, clinical studies have shown that lercanidipine and amlodipine have a comparable antihypertensive efficacy but with significantly less peripheral edema in patients receiving lercanidipine. In some countries, lercanidipine is now available in a single-pill association with an ACE inhibitor thereby further improving its efficacy and tolerability profile.

Angiotensin II stimulates calcineurin activity in proximal tubule epithelia through AT-1 receptor-mediated tyrosine phosphorylation of the PLC-gamma1 isoform.

PubMed

Lea, Janice P; Jin, Shao G; Roberts, Brian R; Shuler, Michael S; Marrero, Mario B; Tumlin, James A

2002-07-01

Angiotensin II (AngII) contributes to the maintenance of extracellular fluid volume by regulating sodium transport in the nephron. In nonepithelial cells, activation of phospholipase C (PLC) by AT-1 receptors stimulates the generation of 1,4,5-trisphosphate (IP(3)) and the release of intracellular calcium. Calcineurin, a serine-threonine phosphatase, is activated by calcium and calmodulin, and both PLC and calcineurin have been linked to sodium transport in the proximal tubule. An examination of whether AngII activates calcineurin in a model of proximal tubule epithelia (LLC-PK1 cells) was performed; AngII increased calcineurin activity within 30 s. An examination of whether AngII activates PLC in proximal tubule epithelia was also performed after first showing that all three families of PLC isoforms are present in LLC-PK1 cells. Application of AngII increased IP(3) generation by 60% within 15 s, which coincided with AngII-induced tyrosine phosphorylation of the PLC-gamma1 isoform also observed at 15 s. AngII-induced tyrosine phosphorylation was blocked by the AT-1 receptor antagonist, Losartan. Subsequently, an inhibitor of tyrosine phosphorylation blocked the AngII-induced activation of calcineurin, as did coincubation with an inhibitor of PLC activity and with an antagonist of the AT-1 receptor. It is therefore concluded that AngII stimulates calcineurin phosphatase activity in proximal tubule epithelial cells through a mechanism involving AT-1 receptor-mediated tyrosine phosphorylation of the PLC isoform.

Calcium ion involvement in growth inhibition of mechanically stressed soybean (Glycine max) seedlings

NASA Technical Reports Server (NTRS)

Jones, R. S.; Mitchell, C. A.

1989-01-01

A 40-50% reduction in soybean [Glycine max (L.) Merr. cv. Century 84] hypocotyl elongation occurred 24 h after application of mechanical stress. Exogenous Ca2+ at 10 mM inhibited growth by 28% if applied with the Ca2+ ionophore A23187 to the zone of maximum hypocotyl elongation. La3+ was even more inhibitory than Ca2+, especially above 5 mM. Treatment with ethyleneglycol-bis-(beta-aminoethylether)-N, N, N', N'-tetraacetic acid (EGTA) alone had no effect on growth of non-stressed seedlings at the concentrations used but negated stress-induced growth reduction by 36% at 4 mM when compared to non-treated, stressed controls. Treatment with EDTA was ineffective in negating stress-induced growth inhibition. Calmodulin antagonists calmidazolium, chlorpromazine, and 48/80 also negated stress-induced growth reduction by 23, 50, and 35%, respectively.

Epidemiology of Gout

PubMed Central

Choi, Hyon

2014-01-01

Synopsis Gout is the most prevalent inflammatory arthritis in men. The findings of several epidemiological studies from a diverse range of countries suggest that the prevalence of gout has risen over the last few decades. Whilst incidence data are scarce, data from the US suggests that the incidence of gout is also rising. Evidence from prospective epidemiological studies has confirmed dietary factors (animal purines, alcohol and fructose), obesity, the metabolic syndrome, hypertension, diuretic use, and chronic kidney disease as clinically relevant risk factors for hyperuricemia and gout. Low-fat dairy products, coffee, and vitamin C appear to have a protective effect. Further prospective studies are required to examine other proposed risk factors for hyperuricaemia and gout such as the use of β-blockers and angiotension-II receptor antagonists (other than losartan), obstructive sleep apnoea, and osteoarthritis, and putative protective factors such as calcium-channel blockers and losartan. PMID:24703341

In Silico Identification of Novel APRIL Peptide Antagonists and Binding Insights by Molecular Modeling and Immunosorbent Assays.

PubMed

Silva, Joao H M da; Calmon-Hamaty, Flavia; Savino, Wilson; Hahne, Michael; Caffarena, Ernesto R

2015-01-01

The "A proliferation inducing ligand" protein (APRIL) is a cytokine over-expressed in many transformed and tumoral cells acting onto two distinct receptors of the Tumoral Necrosis Factor B cell maturation antigen (BCMA) and the transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI). We herein describe, through a detailed computational approach, the molecular interactions between TACI and its ligands APRIL and another structurally similar protein called B-cell activating factor (BAFF) by means of molecular dynamics. Dynamical analysis suggests R84 and D85 residues from TACI as possible mutation candidates, yielding increased affinity between TACI and APRIL. The association of computational simulations, site directed mutagenesis and peptide design could be a powerful tool, driving to better in vitro experiments. Our results contribute to the elucidation of APRIL signaling and help clarify the effects of blocking interaction between APRIL and its receptors through the use of particular peptides.

Leiurus quinquestriatus venom inhibits BRL 34915-induced /sup 86/Rb/sup +/ efflux from the rat portal vein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Quast, U.; Cook, N.S.

1988-01-01

The effect of the crude venom of the Israeli scorpion Leiurus quinquestriatus hebraeus on the /sup 86/Rb/sup +/ efflux stimulated by the K/sup +/ channel opener BRL 34915 in the rat portal vein was examined. Applied alone, the venom greatly increased the spontaneous mechanical activity of and the concomitant /sup 86/Rb/sup +/ efflux from the vessel. When the excitability of the vein was suppressed by the dihydropyridine calcium antagonist, PN 200-110, the /sup 86/Rb/sup +/ efflux stimulated by BRL 34915 could be shown to be inhibited by the venom. From the concentration dependence of this inhibition an IC/sub 50/ valuemore » of 0.17 +/- 0.01 mg/ml was estimated. This venom is thus the most potent blocker of BRL 34915-evoked /sup 86/Rb/sup +/ efflux reported so far. 17 references, 2 figures.« less

Tacripyrines, the first tacrine-dihydropyridine hybrids, as multitarget-directed ligands for the treatment of Alzheimer's disease.

PubMed

Marco-Contelles, José; León, Rafael; de los Ríos, Cristóbal; Samadi, Abdelouahid; Bartolini, Manuela; Andrisano, Vincenza; Huertas, Oscar; Barril, Xavier; Luque, F Javier; Rodríguez-Franco, María I; López, Beatriz; López, Manuela G; García, Antonio G; Carreiras, María do Carmo; Villarroya, Mercedes

2009-05-14

Tacripyrines (1-14) have been designed by combining an AChE inhibitor (tacrine) with a calcium antagonist such as nimodipine and are targeted to develop a multitarget therapeutic strategy to confront AD. Tacripyrines are selective and potent AChE inhibitors in the nanomolar range. The mixed type inhibition of hAChE activity of compound 11 (IC(50) 105 +/- 15 nM) is associated to a 30.7 +/- 8.6% inhibition of the proaggregating action of AChE on the Abeta and a moderate inhibition of Abeta self-aggregation (34.9 +/- 5.4%). Molecular modeling indicates that binding of compound 11 to the AChE PAS mainly involves the (R)-11 enantiomer, which also agrees with the noncompetitive inhibition mechanism exhibited by p-methoxytacripyrine 11. Tacripyrines are neuroprotective agents, show moderate Ca(2+) channel blocking effect, and cross the blood-brain barrier, emerging as lead candidates for treating AD.

A simple, broad-scope nickel(0) precatalyst system for direct amination of allyl alcohols.

PubMed

Sweeney, Joseph B; Ball, Anthony; Lawrence, Philippa; Sinclair, Mackenzie; Smith, Luke

2018-06-25

The preparation of allylic amines is traditionally accomplished by reaction of amines with reactive electrophiles, such as allylic halides, sulfonates or oxyphosphonium species; such methods involve hazardous reagents, generate stoichiometric waste streams, and often suffer from side-reactions (such as over-alkylation). We report here the first nickel-catalysed direct amination of allyl alcohols which enables allylation of primary, secondary and electron-deficient amines, using an inexpensive Ni(II)-Zn couple, without need for glove-box techniques. Under mild conditions, primary and secondary aliphatic amines react smoothly with a range of allyl alcohols, giving secondary and tertiary amines efficiently. This 'totally catalytic' method can be also applied to electron-deficient nitrogen nucleophiles with effective results; the practicality of the process has been demonstrated in an efficient, gram-scale preparation of the calcium antagonist drug substance flunarizine (Sibelium®). © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Use of nitrates in ischemic heart disease.

PubMed

Giuseppe, Cocco; Paul, Jerie; Hans-Ulrich, Iselin

2015-01-01

Short-acting nitrates are beneficial in acute myocardial ischemia. However, many unresolved questions remain about the use of long-acting nitrates in stable ischemic heart disease. The use of long-acting nitrates is weakened by the development of endothelial dysfunction and tolerance. Also, we currently ignore whether lower doses of transdermal nitroglycerin would be better than those presently used. Multivariate analysis data from large nonrandomized studies suggested that long-acting nitrates increase the incidence of acute coronary syndromes, while data from another multivariate study indicate that they have positive effects. Because of methodological differences and open questions, the two studies cannot be compared. A study in Japanese patients with vasospastic angina has shown that, when compared with calcium antagonists, long-acting nitrates do not improve long-term prognosis and that the risk for cardiac adverse events increases with the combined therapy. We have many unanswered questions.

Mortality Benefit of Recombinant Human Interleukin-1 Receptor Antagonist for Sepsis Varies by Initial Interleukin-1 Receptor Antagonist Plasma Concentration.

PubMed

Meyer, Nuala J; Reilly, John P; Anderson, Brian J; Palakshappa, Jessica A; Jones, Tiffanie K; Dunn, Thomas G; Shashaty, Michael G S; Feng, Rui; Christie, Jason D; Opal, Steven M

2018-01-01

Plasma interleukin-1 beta may influence sepsis mortality, yet recombinant human interleukin-1 receptor antagonist did not reduce mortality in randomized trials. We tested for heterogeneity in the treatment effect of recombinant human interleukin-1 receptor antagonist by baseline plasma interleukin-1 beta or interleukin-1 receptor antagonist concentration. Retrospective subgroup analysis of randomized controlled trial. Multicenter North American and European clinical trial. Five hundred twenty-nine subjects with sepsis and hypotension or hypoperfusion, representing 59% of the original trial population. Random assignment of placebo or recombinant human interleukin-1 receptor antagonist × 72 hours. We measured prerandomization plasma interleukin-1 beta and interleukin-1 receptor antagonist and tested for statistical interaction between recombinant human interleukin-1 receptor antagonist treatment and baseline plasma interleukin-1 receptor antagonist or interleukin-1 beta concentration on 28-day mortality. There was significant heterogeneity in the effect of recombinant human interleukin-1 receptor antagonist treatment by plasma interleukin-1 receptor antagonist concentration whether plasma interleukin-1 receptor antagonist was divided into deciles (interaction p = 0.046) or dichotomized (interaction p = 0.028). Interaction remained present across different predicted mortality levels. Among subjects with baseline plasma interleukin-1 receptor antagonist above 2,071 pg/mL (n = 283), recombinant human interleukin-1 receptor antagonist therapy reduced adjusted mortality from 45.4% to 34.3% (adjusted risk difference, -0.12; 95% CI, -0.23 to -0.01), p = 0.044. Mortality in subjects with plasma interleukin-1 receptor antagonist below 2,071 pg/mL was not reduced by recombinant human interleukin-1 receptor antagonist (adjusted risk difference, +0.07; 95% CI, -0.04 to +0.17), p = 0.230. Interaction between plasma interleukin-1 beta concentration and recombinant human interleukin-1 receptor antagonist treatment was not statistically significant. We report a heterogeneous effect of recombinant human interleukin-1 receptor antagonist on 28-day sepsis mortality that is potentially predictable by plasma interleukin-1 receptor antagonist in one trial. A precision clinical trial of recombinant human interleukin-1 receptor antagonist targeted to septic patients with high plasma interleukin-1 receptor antagonist may be worthy of consideration.

Amine dependence of proliferative activity in two transplantable lines of mouse colonic carcinoma.

PubMed

Tutton, P J; Barkla, D H

1987-01-01

Serotonin, histamine and their antagonists have previously been shown to influence both the cell proliferation rate and the volumetric growth rate of colonic tumours. Of these earlier studies, those on cell proliferation could not distinguish between direct effects on tumour cells and indirect effects on the host, whereas those on the volumetric growth rate of tumours, whilst suggesting an outcome related to the individual properties of the tumour rather than the host, could not distinguish between influences on cell gain, cell loss or stromal changes. In an attempt to distinguish between these possibilities the current experiments on the mitotic rate in two lines of transplantable mouse colonic carcinoma were undertaken. One line of tumour proved to be sensitive to inhibition by a histamine H2 receptor antagonist and a dopamine D2 antagonist but resistant to serotonin antagonists; the inhibition by histamine antagonists was surmountable by co-administration of histamine. The other line proved to be highly sensitive to the inhibitory effects of serotonin antagonist and less so to antagonists of the other two amines and in this case the effect of serotonin antagonists was surmountable by serotonin. These results suggest that the variations between different colonic tumours in the response to amine antagonists is due to differences in the extent of inhibition of cell proliferation rather than differences in cell loss or stromal effects. Thus it appears likely that amine antagonists are able to directly interfere with the proliferation of some colonic tumour cells.

Meningitic Escherichia coli K1 penetration and neutrophil transmigration across the blood-brain barrier are modulated by alpha7 nicotinic receptor.

PubMed

Chi, Feng; Wang, Lin; Zheng, Xueye; Wu, Chun-Hua; Jong, Ambrose; Sheard, Michael A; Shi, Wei; Huang, Sheng-He

2011-01-01

Alpha7 nicotinic acetylcholine receptor (nAChR), an essential regulator of inflammation, is abundantly expressed in hippocampal neurons, which are vulnerable to bacterial meningitis. However, it is unknown whether α7 nAChR contributes to the regulation of these events. In this report, an aggravating role of α7 nAChR in host defense against meningitic E. coli infection was demonstrated by using α7-deficient (α7(-/-)) mouse brain microvascular endothelial cells (BMEC) and animal model systems. As shown in our in vitro and in vivo studies, E. coli K1 invasion and polymorphonuclear neutrophil (PMN) transmigration across the blood-brain barrier (BBB) were significantly reduced in α7(-/-) BMEC and α7(-/-) mice. Stimulation by nicotine was abolished in the α7(-/-) cells and animals. The same blocking effect was achieved by methyllycaconitine (α7 antagonist). The tight junction molecules occludin and ZO-1 were significantly reduced in the brain cortex of wildtype mice infected with E. coli and treated with nicotine, compared to α7(-/-) cells and animals. Decreased neuronal injury in the hippocampal dentate gyrus was observed in α7(-/-) mice with meningitis. Proinflammatory cytokines (IL-1β, IL-6, TNFα, MCP-1, MIP-1alpha, and RANTES) and adhesion molecules (CD44 and ICAM-1) were significantly reduced in the cerebrospinal fluids of the α7(-/-) mice with E. coli meningitis. Furthermore, α7 nAChR is the major calcium channel for nicotine- and E. coli K1-increased intracellular calcium concentrations of mouse BMEC. Taken together, our data suggest that α7 nAChR plays a detrimental role in the host defense against meningitic infection by modulation of pathogen invasion, PMN recruitment, calcium signaling and neuronal inflammation.

Meningitic Escherichia coli K1 Penetration and Neutrophil Transmigration Across the Blood–Brain Barrier are Modulated by Alpha7 Nicotinic Receptor

PubMed Central

Zheng, Xueye; Wu, Chun-Hua; Jong, Ambrose; Sheard, Michael A.; Shi, Wei; Huang, Sheng-He

2011-01-01

Alpha7 nicotinic acetylcholine receptor (nAChR), an essential regulator of inflammation, is abundantly expressed in hippocampal neurons, which are vulnerable to bacterial meningitis. However, it is unknown whether α7 nAChR contributes to the regulation of these events. In this report, an aggravating role of α7 nAChR in host defense against meningitic E. coli infection was demonstrated by using α7-deficient (α7-/-) mouse brain microvascular endothelial cells (BMEC) and animal model systems. As shown in our in vitro and in vivo studies, E. coli K1 invasion and polymorphonuclear neutrophil (PMN) transmigration across the blood-brain barrier (BBB) were significantly reduced in α7-/- BMEC and α7-/- mice. Stimulation by nicotine was abolished in the α7-/- cells and animals. The same blocking effect was achieved by methyllycaconitine (α7 antagonist). The tight junction molecules occludin and ZO-1 were significantly reduced in the brain cortex of wildtype mice infected with E. coli and treated with nicotine, compared to α7-/- cells and animals. Decreased neuronal injury in the hippocampal dentate gyrus was observed in α7-/- mice with meningitis. Proinflammatory cytokines (IL-1β, IL-6, TNFα, MCP-1, MIP-1alpha, and RANTES) and adhesion molecules (CD44 and ICAM-1) were significantly reduced in the cerebrospinal fluids of the α7-/- mice with E. coli meningitis. Furthermore, α7 nAChR is the major calcium channel for nicotine- and E. coli K1-increased intracellular calcium concentrations of mouse BMEC. Taken together, our data suggest that α7 nAChR plays a detrimental role in the host defense against meningitic infection by modulation of pathogen invasion, PMN recruitment, calcium signaling and neuronal inflammation. PMID:21966399

Neonatal vaginal irritation results in long-term visceral and somatic hypersensitivity and increased hypothalamic–pituitary–adrenal axis output in female mice

PubMed Central

Pierce, Angela N.; Zhang, Zhen; Fuentes, Isabella M.; Wang, Ruipeng; Ryals, Janelle M.; Christianson, Julie A.

2015-01-01

Abstract Experiencing early life stress or injury increases a woman's likelihood of developing vulvodynia and concomitant dysregulation of the hypothalamic–pituitary–adrenal (HPA) axis. To investigate the outcome of neonatal vaginal irritation (NVI), female mouse pups were administered intravaginal zymosan on postnatal days 8 and 10 and were assessed as adults for vaginal hypersensitivity by measuring the visceromotor response to vaginal balloon distension (VBD). Western blotting and calcium imaging were performed to measure transient receptor potential ankyrin 1 (TRPA1) and vanilloid 1 (TRPV1) in the vagina and innervating primary sensory neurons. Serum corticosterone (CORT), mast cell degranulation, and corticotropin-releasing factor receptor 1 (CRF1) expression were measured as indicators of peripheral HPA axis activation. Colorectal and hind paw sensitivity were measured to determine cross-sensitization resulting from NVI. Adult NVI mice had significantly larger visceromotor response during VBD than naive mice. TRPA1 protein expression was significantly elevated in the vagina, and calcium transients evoked by mustard oil (TRPA1 ligand) or capsaicin (TRPV1 ligand) were significantly decreased in dorsal root ganglion from NVI mice, despite displaying increased depolarization-evoked calcium transients. Serum CORT, vaginal mast cell degranulation, and CRF1 protein expression were all significantly increased in NVI mice, as were colorectal and hind paw mechanical and thermal sensitivity. Neonatal treatment with a CRF1 antagonist, NBI 35965, immediately before zymosan administration largely attenuated many of the effects of NVI. These results suggest that NVI produces chronic hypersensitivity of the vagina, as well as of adjacent visceral and distant somatic structures, driven in part by increased HPA axis activation. PMID:26098441

Exogenous Calcium Alleviates Photoinhibition of PSII by Improving the Xanthophyll Cycle in Peanut (Arachis Hypogaea) Leaves during Heat Stress under High Irradiance

PubMed Central

Yang, Sha; Wang, Fang; Guo, Feng; Meng, Jing-Jing; Li, Xin-Guo; Dong, Shu-Ting; Wan, Shu-Bo

2013-01-01

Peanut is one of the calciphilous plants. Calcium (Ca) serves as a ubiquitous central hub in a large number of signaling pathways. The effect of exogenous calcium nitrate [Ca(NO3)2] (6 mM) on the dissipation of excess excitation energy in the photosystem II (PSII) antenna, especially on the level of D1 protein and the xanthophyll cycle in peanut plants under heat (40°C) and high irradiance (HI) (1 200 µmol m−2 s−1) stress were investigated. Compared with the control plants [cultivated in 0 mM Ca(NO3)2 medium], the maximal photochemical efficiency of PSII (Fv/Fm) in Ca2+-treated plants showed a slighter decrease after 5 h of stress, accompanied by higher non-photochemical quenching (NPQ), higher expression of antioxidative genes and less reactive oxygen species (ROS) accumulation. Meanwhile, higher content of D1 protein and higher ratio of (A+Z)/(V+A+Z) were also detected in Ca2+-treated plants under such stress. These results showed that Ca2+ could help protect the peanut photosynthetic system from severe photoinhibition under heat and HI stress by accelerating the repair of D1 protein and improving the de-epoxidation ratio of the xanthophyll cycle. Furthermore, EGTA (a chelant of Ca ion), LaCl3 (a blocker of Ca2+ channel in cytoplasmic membrane), and CPZ [a calmodulin (CaM) antagonist] were used to analyze the effects of Ca2+/CaM on the variation of (A+Z)/(V+A+Z) (%) and the expression of violaxanthin de-epoxidase (VDE). The results indicated that CaM, an important component of the Ca2+ signal transduction pathway, mediated the expression of the VDE gene in the presence of Ca to improve the xanthophyll cycle. PMID:23940721

Mechanism of resveratrol-induced relaxation of the guinea pig fundus.

PubMed

Tsai, Ching-Chung; Tey, Shu-Leei; Lee, Ming-Che; Liu, Ching-Wen; Su, Yu-Tsun; Huang, Shih-Che

2018-04-01

Resveratrol is a polyphenolic compound that can be isolated from plants and also is a constituent of red wine. Resveratrol induces relaxation of vascular smooth muscle and may prevent cardiovascular diseases. Impaired gastric accommodation plays an important role in functional dyspepsia and fundic relaxation and is a therapeutic target of functional dyspepsia. Although drugs for fundic relaxation have been developed, these types of drugs are still rare. The purpose of this study was to investigate the relaxant effects of resveratrol in the guinea pig fundus. We studied the relaxant effects of resveratrol in the guinea pig fundus. In addition, we investigated the mechanism of resveratrol-induced relaxation on the guinea pig fundus by using tetraethylammonium (a non-selective potassium channel blocker), apamine (a selective inhibitor of the small conductance calcium-activated potassium channel), iberiotoxin (an inhibitor of large conductance calcium-activated potassium channels), glibenclamide (an ATP-sensitive potassium channel blocker), KT 5720 (a cAMP-dependent protein kinase A inhibitor), KT 5823 (a cGMP-dependent protein kinase G inhibitor), NG-nitro-L-arginine (a competitive inhibitor of nitric oxide synthase), tetrodotoxin (a selective neuronal Na + channel blocker), ω-conotoxin GVIA (a selective neuronal Ca 2+ channel blocker) and G-15 (a G-protein coupled estrogen receptor antagonist). The results of this study showed that resveratrol has potent and dose-dependent relaxant effects on the guinea pig fundic muscle. In addition, the results showed that resveratrol-induced relaxation of the guinea pig fundus occurs through nitric oxide and ATP-sensitive potassium channels. This study provides the first evidence concerning the relaxant effects of resveratrol in the guinea pig fundic muscle strips. Furthermore, resveratrol may be a potential drug to relieve gastrointestinal dyspepsia. Copyright © 2018 Elsevier GmbH. All rights reserved.

Regulation of alternative splicing of Slo K+ channels in adrenal and pituitary during the stress-hyporesponsive period of rat development.

PubMed

Lai, Guey-Jen; McCobb, David P

2006-08-01

Stress triggers release of ACTH from the pituitary, glucocorticoids from the adrenal cortex, and epinephrine from the adrenal medulla. Although functions differ, these hormone systems interact in many ways. Previous evidence indicates that pituitary and steroid hormones regulate alternative splicing of the Slo gene at the stress axis-regulated exon (STREX), with functional implications for the calcium-activated K+ channels prominent in adrenal medullary and pituitary cells. Here we examine the role of corticosterone in Slo splicing regulation in pituitary and adrenal tissues during the stress-hyporesponsive period of early rat postnatal life. The sharp drop in plasma corticosterone (CORT) that defines this period offers a unique opportunity to test CORT's role in Slo splicing. We report that in both adrenal and pituitary tissues, the percentage of Slo transcripts having STREX declines and recovers in parallel with CORT. Moreover, addition of 500 nm CORT to cultures of anterior pituitary cells from 13-, 21-, and 30-d postnatal animals increased the percentage of Slo transcripts with STREX, whereas 20 microm CORT reduced STREX representation. Applied to adrenal chromaffin cells, 20 microm CORT decreased STREX inclusion, whereas neither 500 nm nor 2 microm had any effect. The mineralocorticoid receptor antagonist RU28318 abolished the effect of 500 nm CORT on splicing in pituitary cells, whereas the glucocorticoid receptor antagonist RU38486 blocked the effect of 20 microm CORT on adrenal chromaffin cells. These results support the hypothesis that the abrupt, transient drop in CORT during the stress-hyporesponsive period drives the transient decline in STREX splice variant representation in pituitary, but not adrenal.

Curtailing effect of awakening on visual responses of cortical neurons by cholinergic activation of inhibitory circuits.

PubMed

Kimura, Rui; Safari, Mir-Shahram; Mirnajafi-Zadeh, Javad; Kimura, Rie; Ebina, Teppei; Yanagawa, Yuchio; Sohya, Kazuhiro; Tsumoto, Tadaharu

2014-07-23

Visual responsiveness of cortical neurons changes depending on the brain state. Neural circuit mechanism underlying this change is unclear. By applying the method of in vivo two-photon functional calcium imaging to transgenic rats in which GABAergic neurons express fluorescent protein, we analyzed changes in visual response properties of cortical neurons when animals became awakened from anesthesia. In the awake state, the magnitude and reliability of visual responses of GABAergic neurons increased whereas the decay of responses of excitatory neurons became faster. To test whether the basal forebrain (BF) cholinergic projection is involved in these changes, we analyzed effects of electrical and optogenetic activation of BF on visual responses of mouse cortical neurons with in vivo imaging and whole-cell recordings. Electrical BF stimulation in anesthetized animals induced the same direction of changes in visual responses of both groups of neurons as awakening. Optogenetic activation increased the frequency of visually evoked action potentials in GABAergic neurons but induced the delayed hyperpolarization that ceased the late generation of action potentials in excitatory neurons. Pharmacological analysis in slice preparations revealed that photoactivation-induced depolarization of layer 1 GABAergic neurons was blocked by a nicotinic receptor antagonist, whereas non-fast-spiking layer 2/3 GABAergic neurons was blocked only by the application of both nicotinic and muscarinic receptor antagonists. These results suggest that the effect of awakening is mediated mainly through nicotinic activation of layer 1 GABAergic neurons and mixed nicotinic/muscarinic activation of layer 2/3 non-fast-spiking GABAergic neurons, which together curtails the visual responses of excitatory neurons. Copyright © 2014 the authors 0270-6474/14/3410122-12$15.00/0.

Case study: the link between hypertension and diabetes.

PubMed

Bakris, George L; Gonzalez, Edgar R

2007-06-01

Many diabetics develop hypertension, and it is a major risk factor for cardiovascular and microvascular complications. To review a case study of a patient with poorly controlled hypertension and diabetes. Further assessment of this case study shows that the patient has poorly controlled hypertension, despite multiple medications. The patient also has metabolic syndrome complicated.by diabetes, microalbuminuria and peripheral arterial disease. The patient's hypertensive treatment options must be evaluated in light of the fact that polypharmacy has made it more difficult for her to achieve glycemic control. A panoply of drugs and drug classes are available from which to choose: diuretics, beta-blockers, calcium channel blockers, angiotensin-converting enzyme inhibitors, angiotensin II receptor blockers, and aldosterone antagonists. New vasodilatory betablockers reduce adverse drug reactions and produce beneficial effects on arterial vasculature. Various beta~blockers' effects on insulin sensitivity are compared. Older beta-blockers have been shown to have detrimental effects on glucose or lipid parameters. Newer agents such as nebivolol do not impact lipid, glucose, insulin, or high-density lipoproteins. Instead, nebivolol stimulates endothelial nitric oxide release in renal arteries and improves renal function.

Pharmacological modulation of platelet function in hypertension.

PubMed

Blann, Andrew D; Nadar, Sunil; Lip, Gregory Y H

2003-07-01

Platelets exert a considerable influence on human morbidity and mortality. The rationale for their study in hypertension follows the observation that the major consequences of hypertension are stroke and myocardial infarction. However, the etiology of these consequences in hypertension is, paradoxically, not hemorrhagic (as might be expected from the effects of high blood pressure), but occlusive, with thrombus being the culprit lesion. Mechanisms of platelet activation include high shear force, activation of the renin-angiotensin system, endothelial changes, and the presence of comorbidity, such as atrial fibrillation. The treatment of high blood pressure brings about a reversal of the changes seen in the cell. This could be in part due to the direct effect of the drug on the megakaryocyte and/or the platelets themselves, or it might simply be due to the reduction in blood pressure. Some drugs, such as calcium channel antagonists and angiotensin II receptor blockers, however, might have direct effects on platelet biochemistry other than reducing blood pressure. Finally, antiplatelet drugs are becoming an important part of the management of high-risk hypertensives, which aim to minimize vascular complications.

The 5-HT2A receptor antagonist M100907 is more effective in counteracting NMDA antagonist- than dopamine agonist-induced hyperactivity in mice.

PubMed

Carlsson, M L; Martin, P; Nilsson, M; Sorensen, S M; Carlsson, A; Waters, S; Waters, N

1999-01-01

The purpose of the present study was to compare the effectiveness of the selective 5-HT2A antagonist M100907 in different psychosis models. The classical neuroleptic haloperidol was used as reference compound. Two hyperdopaminergia and two hypoglutamatergia mouse models were used. Hyperdopaminergia was produced by the DA releaser d-amphetamine or the DA uptake inhibitor GBR 12909. Hypoglutamatergia was produced by the un-competitive NMDA receptor antagonist MK-801 or the competitive NMDA receptor antagonist D-CPPene. M100907 was found to counteract the locomotor stimulant effects of the NMDA receptor antagonists MK-801 and D-CPPene, but spontaneous locomotion, d-amphetamine- and GBR-12909-induced hyperactivity were not significantly affected. Haloperidol, on the other hand, antagonized both NMDA antagonist- and DA agonist-induced hyperactivity, as well as spontaneous locomotion in the highest dose used. Based on the present and previous results we draw the conclusion that 5-HT2A receptor antagonists are particularly effective against behavioural anomalies resulting from hypoglutamatergia of various origins. The clinical implications of our results and conclusions would be that a 5-HT2A receptor antagonist, due to i a the low side effect liability, could be the preferable treatment strategy in various disorders associated with hypoglutamatergia; such conditions might include schizophrenia, childhood autism and dementia disorders.

Hibiscus Sabdariffa L. Flowers and Olea Europea L. Leaves Extract-Based Formulation for Hypertension Care: In Vitro Efficacy and Toxicological Profile.

PubMed

Micucci, Matteo; Angeletti, Andrea; Cont, Massimiliano; Corazza, Ivan; Aldini, Rita; Donadio, Elisa; Chiarini, Alberto; Budriesi, Roberta

2016-05-01

Olea europaea L. leaves extract (Oe) and Hybiscus sabdariffa L. flowers extract (Hs) have calcium antagonistic properties. Aim of this work was to study the cardiovascular effects of Pres Phytum(®), a nutraceutical formulation containing a mixture of the two extracts and the excipients, and investigate its possible off-target effects, using in vitro biological assays on guinea pig isolated organs. Cardiovascular effects were assessed using guinea pig atria and aorta. The effects of Pres Phytum on spontaneous gastrointestinal, urinary, and respiratory tracts smooth muscle contractility were evaluated. Pres Phytum exerted a vasorelaxant effect (IC50 = 2.38 mg/mL) and a negative chronotropic effect (IC50 = 1.04 mg/mL) at concentrations lower than those producing smooth muscle spontaneous contractility alterations in the other organs. Compared to Pres Phytum, the mixture did not exert negative inotropic activity, while it maintained a negative chronotropic efficacy (IC50 = 1.04 mg/mL). These experimental data suggest a possible nutraceutical use of this food supplement for the management of preclinical hypertension.

Mycolactone-mediated neurite degeneration and functional effects in cultured human and rat DRG neurons

PubMed Central

Sinisi, M; Fox, M; MacQuillan, A; Quick, T; Korchev, Y; Bountra, C; McCarthy, T; Anand, P

2016-01-01

Background Mycolactone is a polyketide toxin secreted by the mycobacterium Mycobacterium ulcerans, responsible for the extensive hypoalgesic skin lesions characteristic of patients with Buruli ulcer. A recent pre-clinical study proposed that mycolactone may produce analgesia via activation of the angiotensin II type 2 receptor (AT2R). In contrast, AT2R antagonist EMA401 has shown analgesic efficacy in animal models and clinical trials for neuropathic pain. We therefore investigated the morphological and functional effects of mycolactone in cultured human and rat dorsal root ganglia (DRG) neurons and the role of AT2R using EMA401. Primary sensory neurons were prepared from avulsed cervical human DRG and rat DRG; 24 h after plating, neurons were incubated for 24 to 96 h with synthetic mycolactone A/B, followed by immunostaining with antibodies to PGP9.5, Gap43, β tubulin, or Mitotracker dye staining. Acute functional effects were examined by measuring capsaicin responses with calcium imaging in DRG neuronal cultures treated with mycolactone. Results Morphological effects: Mycolactone-treated cultures showed dramatically reduced numbers of surviving neurons and non-neuronal cells, reduced Gap43 and β tubulin expression, degenerating neurites and reduced cell body diameter, compared with controls. Dose-related reduction of neurite length was observed in mycolactone-treated cultures. Mitochondria were distributed throughout the length of neurites and soma of control neurons, but clustered in the neurites and soma of mycolactone-treated neurons. Functional effects: Mycolactone-treated human and rat DRG neurons showed dose-related inhibition of capsaicin responses, which were reversed by calcineurin inhibitor cyclosporine and phosphodiesterase inhibitor 3-isobutyl-1-Methylxanthine, indicating involvement of cAMP/ATP reduction. The morphological and functional effects of mycolactone were not altered by Angiotensin II or AT2R antagonist EMA401. Conclusion Mycolactone induces toxic effects in DRG neurons, leading to impaired nociceptor function, neurite degeneration, and cell death, resembling the cutaneous hypoalgesia and nerve damage in individuals with M. Ulcerans infection. PMID:27325560

Activation of brain nitric oxide synthase in depolarized human temporal cortex slices: differential role of voltage-sensitive calcium channels

PubMed Central

Fontana, Giovanni; Fedele, Ernesto; Cossu, Massimo; Munari, Claudio; Raiteri, Maurizio

1997-01-01

Nitric oxide (NO) synthase activity was studied in slices of human temporal cortex samples obtained in neurosurgery by measuring the conversion of L-[3H]-arginine to L-[3H]-citrulline. Elevation of extracellular K+ to 20, 35 or 60 mM concentration-dependently augmented L-[3H]-citrulline production. The response to 35 mM KCl was abolished by NG-nitro-L-arginine (100 μM) demonstrating NO synthase specific conversion of L-arginine to L-citrulline. Increasing extracellular MgCl2 concentration up to 10 mM also prevented the K+ (35 mM)-induced NO synthase activation, suggesting the absolute requirement of external calcium ions for enzyme activity. However, the effect of high K+ (35 mM) on citrulline synthesis was insensitive to the antagonists of ionotropic and metabotropic glutamate receptors dizocilpine (MK-801), 6-nitro-7-sulphamoylbenzo(f)quinoxaline-2-3-dione (NBQX) or L-2-amino-3-phosphonopropionic acid (L-AP3) as well as to the nicotinic receptor antagonist, mecamylamine. The 35 mM K+ response was insensitive to ω-conotoxin GVIA (1 μM) and nifedipine (100 μM), but could be prevented in part by ω-agatoxin IVA (0.1 and 1 μM). The inhibition caused by 0.1 μM ω-agatoxin IVA (∼30%) was enhanced by adding ω-conotoxin GVIA (1 μM) or nifedipine (100 μM). Further inhibition (up to above 70%) could be observed when the three Ca2+ channel blockers were added together. Similarly, synthetic FTX 3.3 arginine polyamine (sFTX) prevented (50% at 100 μM) the K+-evoked NO synthase activation. This effect of sFTX was further enhanced (up to 70%) by adding 1 μM ω-conotoxin GVIA plus 100 μM nifedipine. No further inhibition could be observed upon addition of MK-801 or/and NBQX. It was concluded that elevation of extracellular [K+] causes NO synthase activation by external Ca+ entering cells mainly through channels of the P/Q-type. Other Ca2+ channels (L- and N-type) appear to contribute when P/Q-channels are blocked. PMID:9384511

Activation of brain nitric oxide synthase in depolarized human temporal cortex slices: differential role of voltage-sensitive calcium channels.

PubMed

Fontana, G; Fedele, E; Cossu, M; Munari, C; Raiteri, M

1997-11-01

1. Nitric oxide (NO) synthase activity was studied in slices of human temporal cortex samples obtained in neurosurgery by measuring the conversion of L-[3H]-arginine to L-[3H]-citrulline. 2. Elevation of extracellular K+ to 20, 35 or 60 mM concentration-dependently augmented L-[3H]-citrulline production. The response to 35 mM KCl was abolished by N(G)-nitro-L-arginine (100 microM) demonstrating NO synthase specific conversion of L-arginine to L-citrulline. Increasing extracellular MgCl2 concentration up to 10 mM also prevented the K+ (35 mM)-induced NO synthase activation, suggesting the absolute requirement of external calcium ions for enzyme activity. 3. However, the effect of high K+ (35 mM) on citrulline synthesis was insensitive to the antagonists of ionotropic and metabotropic glutamate receptors dizocilpine (MK-801), 6-nitro-7-sulphamoylbenzo(f)-quinoxaline-2-3-dione (NBQX) or L-2-amino-3-phosphonopropionic acid (L-AP3) as well as to the nicotinic receptor antagonist, mecamylamine. 4. The 35 mM K+ response was insensitive to omega-conotoxin GVIA (1 microM) and nifedipine (100 microM), but could be prevented in part by omega-agatoxin IVA (0.1 and 1 microM). The inhibition caused by 0.1 microM omega-agatoxin IVA (approximately 30%) was enhanced by adding omega-conotoxin GVIA (1 microM) or nifedipine (100 microM). Further inhibition (up to above 70%) could be observed when the three Ca2+ channel blockers were added together. Similarly, synthetic FTX 3.3 arginine polyamine (sFTX) prevented (50% at 100 microM) the K+-evoked NO synthase activation. This effect of sFTX was further enhanced (up to 70%) by adding 1 microM omega-conotoxin GVIA plus 100 microM nifedipine. No further inhibition could be observed upon addition of MK-801 or/and NBQX. 5. It was concluded that elevation of extracellular [K+] causes NO synthase activation by external Ca2+ entering cells mainly through channels of the P/Q-type. Other Ca2+ channels (L- and N-type) appear to contribute when P/Q-channels are blocked.

Clinical protocol for the management of malignant hyperthermia.

PubMed

Kollmann-Camaiora, A; Alsina, E; Domínguez, A; Del Blanco, B; Yepes, M J; Guerrero, J L; García, A

2017-01-01

Malignant hyperthermia is a hypermetabolic syndrome that appears in susceptible patients after exposure to certain anaesthetic drugs (succinylcholine, inhalation anaesthetics). Its incidence in Spain is 1 in 40,000 adults, with a 10% mortality rate. It is induced by an abnormal regulation of the ryanodine receptors, producing a massive release of calcium from the sarcoplasmic reticulum in the striate muscle. Clinical manifestations include: CO 2 increase, tachycardia, haemodynamic instability, metabolic and respiratory acidosis, profuse sweating, hyperpyrexia, CPK increase, myoglobinuria, kidney failure, disseminated intravascular coagulation (DIC), and ending in cardiac arrest. Dantrolene sodium is a ryanodine receptor antagonist, and inhibits the release of intracellular calcium. Definitive diagnosis is achieved by the exposure of muscle fibres to caffeine and halothane. Protocols can help guarantee a reliable and secure management when this severe event occurs. Copyright © 2016 Sociedad Española de Anestesiología, Reanimación y Terapéutica del Dolor. Publicado por Elsevier España, S.L.U. All rights reserved.

TRPM2, calcium and neurodegenerative diseases

PubMed Central

Xie, Yu-Feng; MacDonald, John F; Jackson, Michael F

2010-01-01

NMDA receptor overactivation triggers intracellular Ca2+ dysregulation, which has long been thought to be critical for initiating excitotoxic cell death cascades associated with stroke and neurodegenerative disease. The inability of NMDA receptor antagonists to afford neuroprotection in clinical stroke trials has led to a re-evaluation of excitotoxic models of cell death and has focused research efforts towards identifying additional Ca2+ influx pathways. Recent studies indicate that TRPM2, a member of the TRPM subfamily of Ca2+-permeant, non-selective cation channel, plays an important role in mediating cellular responses to a wide range of stimuli that, under certain situations, can induce cell death. These include reactive oxygen and nitrogen species, tumour necrosis factor as well as soluble oli-gomers of amyloid beta. However, the molecular basis of TRPM2 channel involvement in these processes is not fully understood. In this review, we summarize recent studies about the regulation of TRPM2, its interaction with calcium and the possible implications for neurodegenerative diseases. PMID:21383889

Synthesis, structural elucidation and pharmacological properties of some 5-acetyl-3,4-dihydro-6-methyl-4-(substituted phenyl)-2(1H) -pyrimidinones.

PubMed

Yarim, M; Sarac, S; Ertan, M; Batu, O S; Erol, K

1999-06-30

In this study, the synthesis of some new 5-acetyl-3,4-dihydro-6-methyl-4-(substituted phenyl)-2(1H)-pyrimidinones has been reported. The compounds were prepared by the Biginelli reaction of acetylacetone with aromatic aldehydes and urea. The structures of the compounds were characterized by UV, IR, 1H NMR, 13C NRM, mass spectra and elementary analysis. The calcium antagonistic activity of these compounds was tested in vitro on rat ileum precontracted with 4 x 10(-3) M barium chloride.

Effects of Cannabinoid Agonists and Antagonists on Sleep and Breathing in Sprague-Dawley Rats.

PubMed

Calik, Michael W; Carley, David W

2017-09-01

There are no pharmacological treatments for obstructive sleep apnea syndrome, but dronabinol showed promise in a small pilot study. In anesthetized rats, dronabinol attenuates reflex apnea via activation of cannabinoid (CB) receptors located on vagal afferents; an effect blocked by cannabinoid type 1 (CB1) and/or type 2 (CB2) receptor antagonists. Here, using a natural model of central sleep apnea, we examine the effects of dronabinol, alone and in combination with selective antagonists in conscious rats chronically instrumented to stage sleep and measure cessation of breathing. Adult male Sprague-Dawley rats were anesthetized and implanted with bilateral stainless steel screws into the skull for electroencephalogram recording and bilateral wire electrodes into the nuchal muscles for electromyogram recording. Each animal was recorded by polysomnography on multiple occasions separated by at least 3 days. The study was a fully nested, repeated measures crossover design, such that each rat was recorded following each of 8 intraperitoneal injections: vehicle; vehicle and CB1 antagonist (AM 251); vehicle and CB2 antagonist (AM 630); vehicle and CB1/CB2 antagonist; dronabinol; dronabinol and CB1 antagonist; dronabinol and CB2 antagonist; and dronabinol and CB1/CB2 antagonist. Dronabinol decreased the percent time spent in rapid eye movement (REM) sleep. CB receptor antagonists did not reverse this effect. Dronabinol also decreased apneas during sleep, and this apnea suppression was reversed by CB1 or CB1/CB2 receptor antagonism. Dronabinol's effects on apneas were dependent on CB1 receptor activation, while dronabinol's effects on REM sleep were CB receptor-independent. © Sleep Research Society 2017. Published by Oxford University Press on behalf of the Sleep Research Society. All rights reserved. For permissions, please e-mail journals.permissions@oup.com.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Orton, Frances; Ermler, Sibylle; Kugathas, Subramaniam

Many xenobiotics have been identified as in vitro androgen receptor (AR) antagonists, but information about their ability to produce combined effects at low concentrations is missing. Such data can reveal whether joint effects at the receptor are induced at low levels and may support the prioritisation of in vivo evaluations and provide orientations for the grouping of anti-androgens in cumulative risk assessment. Combinations of 30 AR antagonists from a wide range of sources and exposure routes (pesticides, antioxidants, parabens, UV-filters, synthetic musks, bisphenol-A, benzo(a)pyrene, perfluorooctane sulfonate and pentabromodiphenyl ether) were tested using a reporter gene assay (MDA-kb2). Chemicals were combinedmore » at three mixture ratios, equivalent to single components' effect concentrations that inhibit the action of dihydrotesterone by 1%, 10% or 20%. Concentration addition (CA) and independent action were used to calculate additivity expectations. We observed complete suppression of dihydrotestosterone effects when chemicals were combined at individual concentrations eliciting 1%, 10% or 20% AR antagonistic effect. Due to the large number of mixture components, the combined AR antagonistic effects occurred at very low concentrations of individual mixture components. CA slightly underestimated the combined effects at all mixture ratios. In conclusion, large numbers of AR antagonists from a wide variety of sources and exposure routes have the ability of acting together at the receptor to produce joint effects at very low concentrations. Significant mixture effects are observed when chemicals are combined at concentrations that individually do not induce observable AR antagonistic effects. Cumulative risk assessment for AR antagonists should apply grouping criteria based on effects where data are available, rather than on criteria of chemical similarity. - Highlights: • Mixtures of AR antagonists at low individual concentrations cause complete inhibition. • Concentration addition was an appropriate prediction model for observed effects. • Risk assessment for AR antagonists should use grouping criteria based on effects.« less

Endothelium-dependent relaxation evoked by ATP and UTP in the aorta of P2Y2-deficient mice

PubMed Central

Guns, Pieter-Jan D F; Van Assche, Tim; Fransen, Paul; Robaye, Bernard; Boeynaems, Jean-Marie; Bult, Hidde

2006-01-01

Based on pharmacological criteria, we previously suggested that in the mouse aorta, endothelium-dependent relaxation by nucleotides is mediated by P2Y1 (adenosine diphosphate (ADP)), P2Y2 (adenosine triphosphate (ATP)) and P2Y6 (uridine diphosphate (UDP)) receptors. For UTP, it was unclear whether P2Y2, P2Y6 or yet another subtype was involved. Therefore, in view of the lack of selective purinergic agonists and antagonists, we used P2Y2-deficient mice to clarify the action of UTP. Thoracic aorta segments (width 2 mm) of P2Y2-deficient and wild-type (WT) mice were mounted in organ baths to measure isometric force development and intracellular calcium signalling. Relaxations evoked by ADP, UDP and acetylcholine were identical in knockout and WT mice, indicating that the receptors for these agonists function normally. P2Y2-deficient mice showed impaired ATP- and adenosine 5′[γ-thio] triphosphate (ATPγS)-evoked relaxation, suggesting that in WT mice, ATP and ATPγS activate predominantly the P2Y2 subtype. The ATP/ATPγS-evoked relaxation and calcium signals in the knockout mice were partially rescued by P2Y1, as they were sensitive to 2′-deoxy-N6-methyladenosine 3′,5′-bisphosphate (MRS2179), a P2Y1-selective antagonist. In contrast to ATP, the UTP-evoked relaxation was not different between knockout and WT mice. Moreover, the action of UTP was not sensitive to MRS2179. Therefore, the action of UTP is probably mediated mainly by a P2Y6(like) receptor subtype. In conclusion, we demonstrated that ATP-evoked relaxation of the murine aorta is mainly mediated by P2Y2. But this P2Y2 receptor has apparently no major role in UTP-evoked relaxation. The vasodilator effect of UTP is probably mediated mainly by a P2Y6(like) receptor. PMID:16415908

Pharmacologically relevant receptor binding characteristics and 5alpha-reductase inhibitory activity of free Fatty acids contained in saw palmetto extract.

PubMed

Abe, Masayuki; Ito, Yoshihiko; Oyunzul, Luvsandorj; Oki-Fujino, Tomomi; Yamada, Shizuo

2009-04-01

Saw palmetto extract (SPE), used widely for the treatment of benign prostatic hyperplasia (BPH) has been shown to bind alpha(1)-adrenergic, muscarinic and 1,4-dihydropyridine (1,4-DHP) calcium channel antagonist receptors. Major constituents of SPE are lauric acid, oleic acid, myristic acid, palmitic acid and linoleic acid. The aim of this study was to investigate binding affinities of these fatty acids for pharmacologically relevant (alpha(1)-adrenergic, muscarinic and 1,4-DHP) receptors. The fatty acids inhibited specific [(3)H]prazosin binding in rat brain in a concentration-dependent manner with IC(50) values of 23.8 to 136 microg/ml, and specific (+)-[(3)H]PN 200-110 binding with IC(50) values of 24.5 to 79.5 microg/ml. Also, lauric acid, oleic acid, myristic acid and linoleic acid inhibited specific [(3)H]N-methylscopolamine ([(3)H]NMS) binding in rat brain with IC(50) values of 56.4 to 169 microg/ml. Palmitic acid had no effect on specific [(3)H]NMS binding. The affinity of oleic acid, myristic acid and linoleic acid for each receptor was greater than the affinity of SPE. Scatchard analysis revealed that oleic acid and lauric acid caused a significant decrease in the maximal number of binding sites (B(max)) for [(3)H]prazosin, [(3)H]NMS and (+)-[(3)H]PN 200-110. The results suggest that lauric acid and oleic acid bind noncompetitively to alpha(1)-adrenergic, muscarinic and 1,4-DHP calcium channel antagonist receptors. We developed a novel and convenient method of determining 5alpha-reductase activity using LC/MS. With this method, SPE was shown to inhibit 5alpha-reductase activity in rat liver with an IC(50) of 101 microg/ml. Similarly, all the fatty acids except palmitic acid inhibited 5alpha-reductase activity, with IC(50) values of 42.1 to 67.6 microg/ml. In conclusion, lauric acid, oleic acid, myristic acid, and linoleic acid, major constituents of SPE, exerted binding activities of alpha(1)-adrenergic, muscarinic and 1,4-DHP receptors and inhibited 5alpha-reductase activity.

Metal chelators and neurotoxicity: lead, mercury, and arsenic.

PubMed

Bjørklund, Geir; Mutter, Joachim; Aaseth, Jan

2017-12-01

This article reviews the clinical use of the metal chelators sodium 2,3-dimercapto-1-propanesulfonate (DMPS), meso-2,3-dimercaptosuccinic acid (DMSA), and calcium disodium edetate (CaEDTA, calcium EDTA) in overexposure and poisonings with salts of lead (Pb), mercury (Hg), and arsenic (As). DMSA has considerably lower toxicity than the classic heavy metal antagonist BAL (2,3-dimercaptopropanol) and is also less toxic than DMPS. Because of its adverse effects, CaEDTA should be replaced by DMSA as the antidote of choice in treating moderate Pb poisoning. Combination therapy with BAL and CaEDTA was previously recommended in cases of severe acute Pb poisoning with encephalopathy. We suggest that BAL in such cases acted as a shuttling Pb transporter from the intra- to the extracellular space. The present paper discusses if a combination of the extracellularly distributed DMSA with the ionophore, Monensin may provide a less toxic combination for Pb mobilization by increasing both the efflux of intracellularly deposited Pb and the urinary Pb excretion. Anyhow, oral therapy with DMSA should be continued with several intermittent courses. DMPS and DMSA are also promising antidotes in Hg poisoning, whereas DMPS seems to be a more efficient agent against As poisoning. However, new insight indicates that a combination of low-dosed BAL plus DMPS could be a preferred antidotal therapy to obtain mobilization of the intracerebral deposits into the circulation for subsequent rapid urinary excretion.

The human mu opioid receptor: modulation of functional desensitization by calcium/calmodulin-dependent protein kinase and protein kinase C.

PubMed

Mestek, A; Hurley, J H; Bye, L S; Campbell, A D; Chen, Y; Tian, M; Liu, J; Schulman, H; Yu, L

1995-03-01

Opioids are some of the most efficacious analgesics used in humans. Prolonged administration of opioids, however, often causes the development of drug tolerance, thus limiting their effectiveness. To explore the molecular basis of those mechanisms that may contribute to opioid tolerance, we have isolated a cDNA for the human mu opioid receptor, the target of such opioid narcotics as morphine, codeine, methadone, and fentanyl. The receptor encoded by this cDNA is 400 amino acids long with 94% sequence similarity to the rat mu opioid receptor. Transient expression of this cDNA in COS-7 cells produced high-affinity binding sites to mu-selective agonists and antagonists. This receptor displays functional coupling to a recently cloned G-protein-activated K+ channel. When both proteins were expressed in Xenopus oocytes, functional desensitization developed upon repeated stimulation of the mu opioid receptor, as observed by a reduction in K+ current induced by the second mu receptor activation relative to that induced by the first. The extent of desensitization was potentiated by both the multifunctional calcium/calmodulin-dependent protein kinase and protein kinase C. These results demonstrate that kinase modulation is a molecular mechanism by which the desensitization of mu receptor signaling may be regulated at the cellular level, suggesting that this cellular mechanism may contribute to opioid tolerance in humans.

NAADP and the two-pore channel protein 1 participate in the acrosome reaction in mammalian spermatozoa

PubMed Central

Arndt, Lilli; Castonguay, Jan; Arlt, Elisabeth; Meyer, Dorke; Hassan, Sami; Borth, Heike; Zierler, Susanna; Wennemuth, Gunther; Breit, Andreas; Biel, Martin; Wahl-Schott, Christian; Gudermann, Thomas; Klugbauer, Norbert; Boekhoff, Ingrid

2014-01-01

The functional relationship between the formation of hundreds of fusion pores during the acrosome reaction in spermatozoa and the mobilization of calcium from the acrosome has been determined only partially. Hence, the second messenger NAADP, promoting efflux of calcium from lysosome-like compartments and one of its potential molecular targets, the two-pore channel 1 (TPC1), were analyzed for its involvement in triggering the acrosome reaction using a TPCN1 gene–deficient mouse strain. The present study documents that TPC1 and NAADP-binding sites showed a colocalization at the acrosomal region and that treatment of spermatozoa with NAADP resulted in a loss of the acrosomal vesicle that showed typical properties described for TPCs: Registered responses were not detectable for its chemical analogue NADP and were blocked by the NAADP antagonist trans-Ned-19. In addition, two narrow bell-shaped dose-response curves were identified with maxima in either the nanomolar or low micromolar NAADP concentration range, where TPC1 was found to be responsible for activating the low affinity pathway. Our finding that two convergent NAADP-dependent pathways are operative in driving acrosomal exocytosis supports the concept that both NAADP-gated cascades match local NAADP concentrations with the efflux of acrosomal calcium, thereby ensuring complete fusion of the large acrosomal vesicle. PMID:24451262

Calexcitin interaction with neuronal ryanodine receptors.

PubMed Central

Nelson, T J; Zhao, W Q; Yuan, S; Favit, A; Pozzo-Miller, L; Alkon, D L

1999-01-01

Calexcitin (CE), a Ca2+- and GTP-binding protein, which is phosphorylated during memory consolidation, is shown here to co-purify with ryanodine receptors (RyRs) and bind to RyRs in a calcium-dependent manner. Nanomolar concentrations of CE released up to 46% of the 45Ca label from microsomes preloaded with 45CaCl2. This release was Ca2+-dependent and was blocked by antibodies against the RyR or CE, by the RyR inhibitor dantrolene, and by a seven-amino-acid peptide fragment corresponding to positions 4689-4697 of the RyR, but not by heparin, an Ins(1,4,5)P3-receptor antagonist. Anti-CE antibodies, in the absence of added CE, also blocked Ca2+ release elicited by ryanodine, suggesting that the CE and ryanodine binding sites were in relative proximity. Calcium imaging with bis-fura-2 after loading CE into hippocampal CA1 pyramidal cells in hippocampal slices revealed slow, local calcium transients independent of membrane depolarization. Calexcitin also released Ca2+ from liposomes into which purified RyR had been incorporated, indicating that CE binding can be a proximate cause of Ca2+ release. These results indicated that CE bound to RyRs and suggest that CE may be an endogenous modulator of the neuronal RyR. PMID:10393102

Novel 1, 4-dihydropyridines for L-type calcium channel as antagonists for cadmium toxicity

PubMed Central

Saddala, Madhu Sudhana; Kandimalla, Ramesh; Adi, Pradeepkiran Jangampalli; Bhashyam, Sainath Sri; Asupatri, Usha Rani

2017-01-01

The present study, we design and synthesize the novel dihydropyridine derivatives, i.e., 3 (a-e) and 5 (a-e) and evaluated, anticonvulsant activity. Initially due to the lacuna of LCC, we modeled the protein through modeller 9.15v and evaluated through servers. Docking studies were performed with the synthesized compounds and resulted two best compounds, i.e., 5a, 5e showed the best binding energies. The activity of intracellular Ca2+ measurements was performed on two cell lines: A7r5 (rat aortic smooth muscle cells) and SH-SY5Y (human neuroblastoma cells). The 5a and 5e compounds was showing the more specific activity on L-type calcium channels, i.e. A7r5 (IC50 = 0.18 ± 0.02 and 0.25 ± 0.63 μg/ml, respectively) (containing only L-type channels) than SH-SY5Y (i.e. both L-type and T-type channels) (IC50 = 8 ± 0.23 and 10 ± 0.18 μg/ml, respectively) with intracellular calcium mobility similar to amlodipine. Finally, both in silico and in vitro results exploring two derivatives 5a and 5e succeeded to treat cadmium toxicity. PMID:28345598

TRPV1 variants impair intracellular Ca2+ signaling and may confer susceptibility to malignant hyperthermia.

PubMed

Abeele, Fabien Vanden; Lotteau, Sabine; Ducreux, Sylvie; Dubois, Charlotte; Monnier, Nicole; Hanna, Amy; Gkika, Dimitra; Romestaing, Caroline; Noyer, Lucile; Flourakis, Matthieu; Tessier, Nolwenn; Al-Mawla, Ribal; Chouabe, Christophe; Lefai, Etienne; Lunardi, Joël; Hamilton, Susan; Fauré, Julien; Van Coppenolle, Fabien; Prevarskaya, Natalia

2018-06-21

Malignant hyperthermia (MH) is a pharmacogenetic disorder arising from uncontrolled muscle calcium release due to an abnormality in the sarcoplasmic reticulum (SR) calcium-release mechanism triggered by halogenated inhalational anesthetics. However, the molecular mechanisms involved are still incomplete. We aimed to identify transient receptor potential vanilloid 1 (TRPV1) variants within the entire coding sequence in patients who developed sensitivity to MH of unknown etiology. In vitro and in vivo functional studies were performed in heterologous expression system, trpv1 -/- mice, and a murine model of human MH. We identified TRPV1 variants in two patients and their heterologous expression in muscles of trpv1 -/- mice strongly enhanced calcium release from SR upon halogenated anesthetic stimulation, suggesting they could be responsible for the MH phenotype. We confirmed the in vivo significance by using mice with a knock-in mutation (Y524S) in the type I ryanodine receptor (Ryr1), a mutation analogous to the Y522S mutation associated with MH in humans. We showed that the TRPV1 antagonist capsazepine slows the heat-induced hypermetabolic response in this model. We propose that TRPV1 contributes to MH and could represent an actionable therapeutic target for prevention of the pathology and also be responsible for MH sensitivity when mutated.

Capsaicin-induced reactivation of latent herpes simplex virus type 1 in sensory neurons in culture.

PubMed

Hunsperger, Elizabeth A; Wilcox, Christine L

2003-05-01

Herpes simplex virus type 1 (HSV-1) produces a life-long latent infection in neurons of the peripheral nervous system, primarily in the trigeminal and dorsal root ganglia. Neurons of these ganglia express high levels of the capsaicin receptor, also known as the vanilloid receptor-1 (VR-1). VR-1 is a non-selective ion channel, found on sensory neurons, that primarily fluxes Ca(2+) ions in response to various stimuli, including physiologically acidic conditions, heat greater than 45 degrees C and noxious compounds such as capsaicin. Using an in vitro neuronal model to study HSV-1 latency and reactivation, we found that agonists of the VR-1 channel - capsaicin and heat - resulted in reactivation of latent HSV-1. Capsaicin-induced reactivation of HSV-1 latently infected neurons was dose-dependent. Additionally, activation of VR-1 at its optimal temperature of 46 degrees C caused a significant increase in virus titres, which could be attenuated with the VR-1 antagonist, capsazepine. VR-1 activation that resulted in HSV-1 reactivation was calcium-dependent, since the calcium chelator BAPTA significantly reduced reactivation following treatment with caspsaicin and forskolin. Taken together, these results suggest that activation of the VR-1 channel, often associated with increases in intracellular calcium, results in HSV-1 reactivation in sensory neurons.

Stevia Nonsweetener Fraction Displays an Insulinotropic Effect Involving Neurotransmission in Pancreatic Islets.

PubMed

Piovan, Silvano; Pavanello, Audrei; Peixoto, Giuliana Maria Ledesma; Matiusso, Camila Cristina Ianoni; de Moraes, Ana Maria Praxedes; Martins, Isabela Peixoto; Malta, Ananda; Palma-Rigo, Kesia; da Silva Franco, Claudinéia Conationi; Milani, Paula Gimenez; Dacome, Antonio Sérgio; da Costa, Silvio Claudio; de Freitas Mathias, Paulo Cezar; Mareze-Costa, Cecília Edna

2018-01-01

Stevia rebaudiana (Bert.) Bertoni besides being a source of noncaloric sweeteners is also an important source of bioactive molecules. Many plant extracts, mostly obtained with ethyl acetate solvent, are rich in polyphenol compounds that present insulinotropic effects. To investigate whether the nonsweetener fraction, which is rich in phenolic compounds isolated from Stevia rebaudiana with the solvent ethyl acetate (EAF), has an insulinotropic effect, including interference at the terminals of the autonomic nervous system of the pancreatic islets of rats. Pancreatic islets were isolated from Wistar rats and incubated with EAF and inhibitory or stimulatory substances of insulin secretion, including cholinergic and adrenergic agonists and antagonists. EAF potentiates glucose-stimulated insulin secretion (GSIS) only in the presence of high glucose and calcium-dependent concentrations. EAF increased muscarinic insulinotropic effects in pancreatic islets, interfering with the muscarinic receptor subfamily M 3 . Adrenergic inhibitory effects on GSIS were attenuated in the presence of EAF, which interfered with the adrenergic α 2 receptor. Results suggest that EAF isolated from stevia leaves is a potential therapy for treating type 2 diabetes mellitus by stimulating insulin secretion only in high glucose concentrations, enhancing parasympathetic signal transduction and inhibiting sympathetic signal transduction in beta cells.

Stevia Nonsweetener Fraction Displays an Insulinotropic Effect Involving Neurotransmission in Pancreatic Islets

PubMed Central

Pavanello, Audrei; Peixoto, Giuliana Maria Ledesma; Matiusso, Camila Cristina Ianoni; de Moraes, Ana Maria Praxedes; Martins, Isabela Peixoto; Palma-Rigo, Kesia; da Silva Franco, Claudinéia Conationi; Milani, Paula Gimenez; Dacome, Antonio Sérgio; da Costa, Silvio Claudio; de Freitas Mathias, Paulo Cezar; Mareze-Costa, Cecília Edna

2018-01-01

Stevia rebaudiana (Bert.) Bertoni besides being a source of noncaloric sweeteners is also an important source of bioactive molecules. Many plant extracts, mostly obtained with ethyl acetate solvent, are rich in polyphenol compounds that present insulinotropic effects. To investigate whether the nonsweetener fraction, which is rich in phenolic compounds isolated from Stevia rebaudiana with the solvent ethyl acetate (EAF), has an insulinotropic effect, including interference at the terminals of the autonomic nervous system of the pancreatic islets of rats. Pancreatic islets were isolated from Wistar rats and incubated with EAF and inhibitory or stimulatory substances of insulin secretion, including cholinergic and adrenergic agonists and antagonists. EAF potentiates glucose-stimulated insulin secretion (GSIS) only in the presence of high glucose and calcium-dependent concentrations. EAF increased muscarinic insulinotropic effects in pancreatic islets, interfering with the muscarinic receptor subfamily M3. Adrenergic inhibitory effects on GSIS were attenuated in the presence of EAF, which interfered with the adrenergic α 2 receptor. Results suggest that EAF isolated from stevia leaves is a potential therapy for treating type 2 diabetes mellitus by stimulating insulin secretion only in high glucose concentrations, enhancing parasympathetic signal transduction and inhibiting sympathetic signal transduction in beta cells. PMID:29853880

Effects of magnolol and honokiol derived from traditional Chinese herbal remedies on gastrointestinal movement

PubMed Central

Zhang, Wei-Wei; Li, Yan; Wang, Xue-Qing; Tian, Feng; Cao, Hong; Wang, Min-Wei; Sun, Qi-Shi

2005-01-01

AIM: To study the effects of magnolol and honokiol on isolated smooth muscle of gastrointestinal tract and their relationship with Ca2+, and on the gastric emptying and the intestinal propulsive activity in mice. METHODS: Routine experimental methods using isolated gastric fundus strips of rats and isolated ileum segments of guinea pigs were adopted to measure the smooth muscle tension. The effects of magnolol 10-3, 10-4, 10-5 mol/L, and honokiol 10-4, 10-5, 10-6 mol/L on the contractility of gastric fundus strips of rats and ileum of guinea pigs induced by acetylcholine (Ach) and 5-hydroxytryptamine (5-HT) was assessed respectively. The method using nuclein and pigment methylene blue was adopted to measure the gastric retention rate of nuclein and the intestinal propulsive ratio of a nutritional semi-solid meal for assessing the effect of magnolol and honokiol (0.5, 2, 20 mg/kg) on gastric emptying and intestinal propulsion. RESULTS: Magnolol and honokiol significantly inhibited the contractility of isolated gastric fundus strips of rats treated with Ach or 5-HT and isolated ileum guinea pigs treated with Ach or CaCl2, and both of them behaved as non-competitive muscarinic antagonists. Magnolol and honokiol inhibited the contraction induced by Ach in Ca2+-free medium and extracellular Ca2+-dependent contraction induced by Ach. Each group of magnolol and honokiol experiments significantly decreased the residual rate of nuclein in the stomach and increased the intestinal propulsive ratio in mice. CONCLUSION: The inhibitory effect of magnolol and honokiol on contractility of the smooth muscles of isolated gastric fundus strips of rats and isolated ileum of guinea pigs is associated with a calcium-antagonistic effect. Magnolol and honokiol can improve the gastric emptying of a semi-solid meal and intestinal propulsive activity in mice. PMID:16038044

The Effect of Sympathetic Antagonists on the Antidepressant Action of Alprazolam

PubMed Central

Al-Tubuly, RA; Aburawi, SM; Alghzewi, EA; Gorash, ZM; Errwami, S

2008-01-01

Alprazolam is an anti-anxiety drug shown to be effective in the treatment of depression. In this study, the effect of sympathetic receptor antagonists on alprazolam–induced antidepressant action was studied using a mouse model of forced swimming behavioral despair. The interaction of three sympathetic receptor antagonists with benzodiazepines, which may impact the clinical use of alprazolam, was also studied. Behavioral despair was examined in six groups of albino mice. Drugs were administered intraperitoneally. The control group received only a single dose of 1% Tween 80. The second group received a single dose of alprazolam, and the third group received an antagonist followed by alprazolam. The fourth group was treated with imipramine, and the fifth group received an antagonist followed by imipramine. The sixth group was treated with a single dose of an antagonist alone (atenolol, a β1-selective adrenoceptor antagonist; propranolol, a non selective β-adrenoceptor antagonist; and prazocin, an α1-adrenoceptor antagonist). Results confirmed the antidepressant action of alprazolam and imipramine. Prazocin treatment alone produced depression, but it significantly potentiated the antidepressant actions of imipramine and alprazolam. Atenolol alone produced an antidepressant effect and potentiated the antidepressant action of alprazolam. Propranolol treatment alone produced depression, and antagonized the effects of alprazolam and imipramine, even producing depression in combined treatments.In conclusion, our results reveal that alprazolam may produce antidepressant effects through the release of noradrenaline, which stimulates β2 receptors to produce an antidepressant action. Imipramine may act by activating β2 receptors by blocking or down-regulating β1 receptors. PMID:21499463

Adenosinergic modulation of the discriminative-stimulus effects of methamphetamine in rats.

PubMed

Munzar, Patrik; Justinova, Zuzana; Kutkat, Scott W; Ferré, Sergi; Goldberg, Steven R

2002-06-01

A(1) and A(2A) adenosine receptors are co-localized with dopamine D(1) and D(2) receptors, respectively, and their stimulation attenuates dopaminergic functioning. To test whether adenosine antagonists with different selectivities for A(1) and A(2A) receptors mimic the discriminative-stimulus effects of dopamine releaser methamphetamine. Effects of the A(1) antagonist DPCPX, the preferential A(2A) antagonist DMPX and the non-selective adenosine antagonist caffeine were evaluated in Sprague-Dawley rats trained to discriminate 1.0 mg/kg, IP, methamphetamine from saline under a fixed-ratio 10 schedule of food presentation. The A(1) antagonist DPCPX (1.0-10.0 mg/kg) failed to substitute for methamphetamine. However, 5.6 mg/kg DPCPX shifted the methamphetamine dose-response curve to the left. The A(2A) antagonist DMPX (1.8-18.0 mg/kg) produced about 70% methamphetamine-appropriate responding and the non-selective antagonist caffeine (3.0-56.0 mg/kg) about 50% methamphetamine-appropriate responding at the highest tested doses. Both DMPX (5.6 mg/kg) and caffeine (30.0 mg/kg) shifted the methamphetamine dose-response curve to the left. Methamphetamine-like effects of DMPX were blocked fully by the D(2) antagonist spiperone (0.18 mg/kg) and partially by the D(1) antagonist SCH-23390 (0.018 mg/kg). Antagonism at A(2A) adenosine receptors directly mimics the discriminative-stimulus effects of methamphetamine through the interaction with dopamine receptors. Antagonism at A(1) adenosine receptors potentiates effects of lower methamphetamine doses and thus plays a rather indirect, modulatory role.

Opioid adjuvant strategy: improving opioid effectiveness.

PubMed

Bihel, Frédéric

2016-01-01

Opioid analgesics continue to be the mainstay of pharmacologic treatment of moderate to severe pain. Many patients, particularly those suffering from chronic pain, require chronic high-dose analgesic therapy. Achieving clinical efficacy and tolerability of such treatment regimens is hampered by the appearance of opioid-induced side effects such as tolerance, hyperalgesia and withdrawal syndrome. Among the therapeutic options to improve the opioid effectiveness, this current review focuses on strategies combining opioids to other drugs that can modulate opioid-mediated effects. We will discuss about experimental evidences reported for several potential opioid adjuvants, including N-methyl-D-aspartate receptor antagonists, 5-HT7 agonists, sigma-1 antagonists, I2-R ligands, cholecystokinin antagonists, neuropeptide FF-R antagonists and toll-like receptor 4 antagonists.

Discovery of potent peptide-mimetic antagonists for the human thrombin receptor, protease-activated receptor-1 (PAR-1).

PubMed

Maryanoff, Bruce E; Zhang, Han-Cheng; Andrade-Gordon, Patricia; Derian, Claudia K

2003-03-01

Protease-activated receptors (PARs) represent a unique family of seven-transmembrane G-protein-coupled receptors, which are enzymatically cleaved to expose a new extracellular N-terminus that acts as a tethered activating ligand. PAR-1 is cleaved and activated by the serine protease alpha-thrombin, is expressed in various tissues (e.g. platelets and vascular cells), and is involved in cellular responses associated with hemostasis, proliferation, and tissue injury. By using a de novo design approach, we have discovered a series of potent heterocycle-based peptide-miimetic antagonists of PAR-1, exemplified by advanced leads RWJ-56110 (22) and RWJ-58259 (32). These compounds are potent, selective PAR-1 antagonists, devoid of PAR-1 agonist and thrombin inhibitory activity: they bind to PAR-1, interfere with calcium mobilization and cellular functions associated with PAR-1, and do not affect PAR-2, PAR-3, or PAR-4. RWJ-56110 was determined to be a direct inhibitor of PAR-1 activation and internalization, without affecting PAR-1 N-terminal cleavage. At high concentrations of alpha-thrombin, RWJ-56110 fully blocked activation responses in human vascular cells, but not in human platelets; whereas, at high concentrations of TRAP-6, RWJ-56110 blocked activation responses in both cell types. This result is consistent with the presence of another thrombin receptor on human platelets, namely PAR-4. RWJ-56110 and RWJ-58259 clearly interrupt the binding of a tethered ligand to its receptor. RWJ-58259 demonstrated antirestenotic activity in a rat balloon angioplasty model and antithrombotic activity in a cynomolgus monkey arterial injury model. Such PAR-1 antagonists should not only serve as useful tools to delineate the physiological and pathophysiological roles of PAR-1, but also may have therapeutic potential for treating thrombosis and restenosis in humans.

High-Throughput Screening of Myometrial Calcium-Mobilization to Identify Modulators of Uterine Contractility

PubMed Central

Herington, Jennifer L.; Swale, Daniel R.; Brown, Naoko; Shelton, Elaine L.; Choi, Hyehun; Williams, Charles H.; Hong, Charles C.; Paria, Bibhash C.; Denton, Jerod S.; Reese, Jeff

2015-01-01

The uterine myometrium (UT-myo) is a therapeutic target for preterm labor, labor induction, and postpartum hemorrhage. Stimulation of intracellular Ca2+-release in UT-myo cells by oxytocin is a final pathway controlling myometrial contractions. The goal of this study was to develop a dual-addition assay for high-throughput screening of small molecular compounds, which could regulate Ca2+-mobilization in UT-myo cells, and hence, myometrial contractions. Primary murine UT-myo cells in 384-well plates were loaded with a Ca2+-sensitive fluorescent probe, and then screened for inducers of Ca2+-mobilization and inhibitors of oxytocin-induced Ca2+-mobilization. The assay exhibited robust screening statistics (Z´ = 0.73), DMSO-tolerance, and was validated for high-throughput screening against 2,727 small molecules from the Spectrum, NIH Clinical I and II collections of well-annotated compounds. The screen revealed a hit-rate of 1.80% for agonist and 1.39% for antagonist compounds. Concentration-dependent responses of hit-compounds demonstrated an EC50 less than 10μM for 21 hit-antagonist compounds, compared to only 7 hit-agonist compounds. Subsequent studies focused on hit-antagonist compounds. Based on the percent inhibition and functional annotation analyses, we selected 4 confirmed hit-antagonist compounds (benzbromarone, dipyridamole, fenoterol hydrobromide and nisoldipine) for further analysis. Using an ex vivo isometric contractility assay, each compound significantly inhibited uterine contractility, at different potencies (IC50). Overall, these results demonstrate for the first time that high-throughput small-molecules screening of myometrial Ca2+-mobilization is an ideal primary approach for discovering modulators of uterine contractility. PMID:26600013

Mangiferin, a natural xanthone, accelerates gastrointestinal transit in mice involving cholinergic mechanism

PubMed Central

Morais, Talita Cavalcante; Lopes, Synara Cavalcante; Carvalho, Karine Maria Martins Bezerra; Arruda, Bruno Rodrigues; de Souza, Francisco Thiago Correia; Trevisan, Maria Teresa Salles; Rao, Vietla Satyanarayana; Santos, Flávia Almeida

2012-01-01

AIM: To investigate the effects of mangiferin on gastrointestinal transit (GIT) in normal and constipated mice, together with the possible mechanism. METHODS: Intragastrically-administered charcoal meal was used to measure GIT in overnight starved Swiss mice. In the first experiments, mangiferin (3 mg/kg, 10 mg/kg, 30 mg/kg, and 100 mg/kg, po) or tegaserod (1 mg/kg, ip) were administered 30 min before the charcoal meal to study their effects on normal transit. In the second series, mangiferin (30 mg/kg) was tested on delayed GIT induced by several different pharmacological agonists (morphine, clonidine, capsaicin) or antagonists (ondansetron, verapamil, and atropine) whereas in the third series, mangiferin (30 mg/kg, 100 mg/kg and 300 mg/kg) or tegaserod (1 mg/kg) were tested on 6 h fecal pellets outputted by freely fed mice. The ratio of wet to dry weight was calculated and used as a marker of fecal water content. RESULTS: Mangiferin administered orally significantly (P < 0.05) accelerated GIT at 30 mg/kg and 100 mg/kg (89% and 93%, respectively), similarly to 5-hydroxytryptamine4 (5-HT4) agonist tegaserod (81%) when compared to vehicle-treated control (63%). Co-administered mangiferin (30 mg/kg) totally reversed the inhibitory effect of opioid agonist morphine, 5-HT3-receptor antagonist ondansetron and transient receptor potential vanilloid-1 receptor agonist capsaicin on GIT, but only to a partial extent with the GIT-delay induced by α2-adrenoceptor agonist clonidine, and calcium antagonist verapamil. However, co-administered atropine completely blocked the stimulant effect of mangiferin on GIT, suggesting the involvement of muscarinic acetylcholine receptor activation. Although mangiferin significantly enhanced the 6 h fecal output at higher doses (245.5 ± 10.43 mg vs 161.9 ± 10.82 mg and 227.1 ± 20.11 mg vs 161.9 ± 10.82 mg of vehicle-treated control, at 30 and 100 mg/kg, P < 0.05, respectively), the effect of tegaserod was more potent (297.4 ± 7.42 mg vs 161.9 ± 10.82 mg of vehicle-treated control, P < 0.05). Unlike tegaserod, which showed an enhanced water content in fecal pellets (59.20% ± 1.09% vs 51.44% ± 1.19% of control, P < 0.05), mangiferin evidenced no such effect, indicating that it has only a motor and not a secretomotor effect. CONCLUSION: Our data indicate the prokinetic action of mangiferin. It can stimulate the normal GIT and also overcome the drug-induced transit delay, via a cholinergic physiological mechanism. PMID:22783044

Expression of TRPV1 channels by Cajal-Retzius cells and layer-specific modulation of synaptic transmission by capsaicin in the mouse hippocampus.

PubMed

Anstötz, Max; Lee, Sun Kyong; Maccaferri, Gianmaria

2018-05-28

By taking advantage of calcium imaging and electrophysiology, we provide direct pharmacological evidence for the functional expression of TRPV1 channels in hippocampal Cajal-Retzius cells. Application of the TRPV1 activator capsaicin powerfully enhances spontaneous synaptic transmission in the hippocampal layers that are innervated by the axons of Cajal-Retzius cells. Capsaicin-triggered calcium responses and membrane currents in Cajal-Retzius cells, as well as layer-specific modulation of spontaneous synaptic transmission, are absent when the drug is applied to slices prepared from TRPV1 - / - animals. We discuss the implications of the functional expression of TRPV1 channels in Cajal-Retzius cells and of the observed TRPV1-dependent layer-specific modulation of synaptic transmission for physiological and pathological network processing. The vanilloid receptor TRPV1 forms complex polymodal channels that are expressed by sensory neurons and play a critical role in nociception. Their distribution pattern and functions in cortical circuits are, however, much less understood. Although TRPV1 reporter mice have suggested that, in the hippocampus, TRPV1 is predominantly expressed by Cajal-Retzius cells (CRs), direct functional evidence is missing. As CRs powerfully excite GABAergic interneurons of the molecular layers, TRPV1 could play important roles in the regulation of layer-specific processing. Here, we have taken advantage of calcium imaging with the genetically encoded indicator GCaMP6s and patch-clamp techniques to study the responses of hippocampal CRs to the activation of TRPV1 by capsaicin, and have compared the effect of TRPV1 stimulation on synaptic transmission in layers innervated or non-innervated by CRs. Capsaicin induced both calcium responses and membrane currents in ∼50% of the cell tested. Neither increases of intracellular calcium nor whole-cell currents were observed in the presence of the TRPV1 antagonists capsazepine/Ruthenium Red or in slices prepared from TRPV1 knockout mice. We also report a powerful TRPV1-dependent enhancement of spontaneous synaptic transmission onto interneurons with dendritic trees confined to the layers innervated by CRs. In conclusion, our work establishes that functional TRPV1 is expressed by a significant fraction of CRs and we propose that TRPV1 activity may regulate layer-specific synaptic transmission in the hippocampus. Lastly, as CR density decreases during postnatal development, we also propose that functional TRPV1 receptors may be related to mechanisms involved in CR progressive reduction by calcium-dependent toxicity/apoptosis. © 2018 The Authors. The Journal of Physiology © 2018 The Physiological Society.

Electrophysiological evidence for different release mechanism of ATP and NO as inhibitory NANC transmitters in guinea-pig colon.

PubMed

Zagorodnyuk, V; Maggi, C A

1994-08-01

1. The effect of the P2-purinoceptor antagonist, suramin, the specific N-type voltage-dependent calcium channel blocker, omega-conotoxin GVIA (omega-CgTx) and the delta-opioid receptor agonist [D-Pen2,D-Pen5] enkephalin (DPDPE) on the apamin-sensitive and apamin-resistant inhibitory junction potentials (i.j.ps) produced by electrical field stimulation (EFS) were investigated by means of a sucrose-gap technique in the circular muscle of the guinea-pig colon. 2. After incubation of muscle strips in either atropine (1 microM), guanethidine (3 microM) and NG-nitro-L-arginine (L-NOARG, 30 microM) or atropine, guanethidine and apamin (0.3 microM), the addition of the NK1 receptor antagonist, SR 140,333 (1 microM) abolished the non-adrenergic, non-cholinergic (NANC) excitatory junction potential (e.j.p.) and unmasked a pure apamin-sensitive i.j.p. (in the presence of L-NOARG) or a pure apamin-resistant i.j.p. (in the presence of apamin). Both types of i.j.p. were abolished by tetrodotoxin. 3. Suramin (30-300 microM) concentration-dependently inhibited the apamin-sensitive i.j.p., while the apamin-resistant i.j.p. was not significantly affected by suramin (up to 300 microM). L-NOARG (30 microM) markedly reduced the apamin-resistant i.j.p. 4. The delta-opioid receptor agonist, DPDPE (0.03-3 microM) concentration-dependently reduced the apamin-sensitive i.j.p., while leaving the apamin-resistant i.j.p. unaffected. Naloxone (1 microM) prevented the i.j.p. inhibition evoked by DPDPE (0.3 microM). 5. omega-CgTx (0.3 microM) markedly reduced the apamin-sensitive but not the apamin-resistant i.j.p. The application of DPDPE (3 MicroM), after development of a steady state inhibitory effect by omega-CgTx, evoked further inhibition of the apamin-sensitive ij.p., similar to the effect produced by DPDPE alone. The L-type calcium channel blocker, nifedipine (1 MicroM) did not significantly affect either the apamin-sensitive or the apamin-resistant ij.ps.6. These findings support the purinergic origin of the fast, apamin-sensitive ij.p. produced by EFS in the circular muscle of the guinea-pig colon and strongly suggest that the apamin-sensitive and the apamin-resistant components of the evoked ij.p. utilize different mechanisms for the secretion of theNANC transmitters, ATP and NO, respectively.

Cross-talk between calcium-calmodulin and nitric oxide in abscisic acid signaling in leaves of maize plants.

PubMed

Sang, Jianrong; Zhang, Aying; Lin, Fan; Tan, Mingpu; Jiang, Mingyi

2008-05-01

Using pharmacological and biochemical approaches, the signaling pathways between hydrogen peroxide (H(2)O(2)), calcium (Ca(2+))-calmodulin (CaM), and nitric oxide (NO) in abscisic acid (ABA)-induced antioxidant defense were investigated in leaves of maize (Zea mays L.) plants. Treatments with ABA, H(2)O(2), and CaCl(2) induced increases in the generation of NO in maize mesophyll cells and the activity of nitric oxide synthase (NOS) in the cytosolic and microsomal fractions of maize leaves. However, such increases were blocked by the pretreatments with Ca(2+) inhibitors and CaM antagonists. Meanwhile, pretreatments with two NOS inhibitors also suppressed the Ca(2+)-induced increase in the production of NO. On the other hand, treatments with ABA and the NO donor sodium nitroprusside (SNP) also led to increases in the concentration of cytosolic Ca(2+) in protoplasts of mesophyll cells and in the expression of calmodulin 1 (CaM1) gene and the contents of CaM in leaves of maize plants, and the increases induced by ABA were reduced by the pretreatments with a NO scavenger and a NOS inhibitor. Moreover, SNP-induced increases in the expression of the antioxidant genes superoxide dismutase 4 (SOD4), cytosolic ascorbate peroxidase (cAPX), and glutathione reductase 1 (GR1) and the activities of the chloroplastic and cytosolic antioxidant enzymes were arrested by the pretreatments with Ca(2+) inhibitors and CaM antagonists. Our results suggest that Ca(2+)-CaM functions both upstream and downstream of NO production, which is mainly from NOS, in ABA- and H(2)O(2)-induced antioxidant defense in leaves of maize plants.

Abeta oligomers induce neuronal oxidative stress through an N-methyl-D-aspartate receptor-dependent mechanism that is blocked by the Alzheimer drug memantine.

PubMed

De Felice, Fernanda G; Velasco, Pauline T; Lambert, Mary P; Viola, Kirsten; Fernandez, Sara J; Ferreira, Sergio T; Klein, William L

2007-04-13

Oxidative stress is a major aspect of Alzheimer disease (AD) pathology. We have investigated the relationship between oxidative stress and neuronal binding of Abeta oligomers (also known as ADDLs). ADDLs are known to accumulate in brain tissue of AD patients and are considered centrally related to pathogenesis. Using hippocampal neuronal cultures, we found that ADDLs stimulated excessive formation of reactive oxygen species (ROS) through a mechanism requiring N-methyl-d-aspartate receptor (NMDA-R) activation. ADDL binding to neurons was reduced and ROS formation was completely blocked by an antibody to the extracellular domain of the NR1 subunit of NMDA-Rs. In harmony with a steric inhibition of ADDL binding by NR1 antibodies, ADDLs that were bound to detergent-extracted synaptosomal membranes co-immunoprecipitated with NMDA-R subunits. The NR1 antibody did not affect ROS formation induced by NMDA, showing that NMDA-Rs themselves remained functional. Memantine, an open channel NMDA-R antagonist prescribed as a memory-preserving drug for AD patients, completely protected against ADDL-induced ROS formation, as did other NMDA-R antagonists. Memantine and the anti-NR1 antibody also attenuated a rapid ADDL-induced increase in intraneuronal calcium, which was essential for stimulated ROS formation. These results show that ADDLs bind to or in close proximity to NMDA-Rs, triggering neuronal damage through NMDA-R-dependent calcium flux. This response provides a pathologically specific mechanism for the therapeutic action of memantine, indicates a role for ROS dysregulation in ADDL-induced cognitive impairment, and supports the unifying hypothesis that ADDLs play a central role in AD pathogenesis.

Amphetamine concentrations in human urine following single-dose administration of the calcium antagonist prenylamine-studies using fluorescence polarization immunoassay (FPIA) and GC-MS.

PubMed

Kraemer, Thomas; Roditis, Susanne K; Peters, Frank T; Maurer, Hans H

2003-03-01

Prenylamine (R,S-N-(3,3-diphenylpropyl-methyl-2-phenethylamine), a World Health Organization class V calcium antagonist, is known to be metabolized to amphetamine. In this study, amphetamine concentrations after a single-dose administration of prenylamine were determined to check if they reached values that could be of analytical and/or pharmacological importance in clinical and forensic toxicology. Enantiomeric composition of amphetamine was also studied. Five volunteers received a single 120-mg oral dose of prenylamine. Urine samples were analyzed using the Abbott TDx immunoassay Amphetamine/Methamphetamine II and using our routine systematic toxicological analysis (STA) gas chromatography-mass spectrometry (GC-MS) procedure. For quantitation purposes, GC-MS was used in the selected-ion monitoring (SIM) mode (ions m/z 118, 122, 240, 244) after solid-phase extraction (Isolute Confirm HCX) and derivatization (heptafluorobutyric anhydride). Amphetamine-d5 was used as internal standard (IS). Chiral separation of the heptafluorobutyrated amphetamine enantiomers was achieved using an Astec Chiraldex G-PN column. The TDx results showed a great variability for the different volunteers. A urine sample of one volunteer showed results as high as 3200 ng/mL, whereas the urine samples of another volunteer never gave results greater than the TDx detection limit (100 ng/mL). Using the STA procedure, the presence of amphetamine could be confirmed in all urine samples with TDx results greater than the cutoff value (300 ng/mL). Using the GC-MS SIM method, amphetamine concentrations up to 1280 ng/mL were determined. Chiral analysis revealed that both enantiomers of amphetamine were present in the samples with a surplus of the S(+)-enantiomer in the early phase of excretion. Forensic implications are discussed.

Postinhibitory rebound neurons and networks are disrupted in retrovirus-induced spongiform neurodegeneration

PubMed Central

Li, Ying; Davey, Robert A.; Lynch, William P.

2014-01-01

Certain retroviruses induce progressive spongiform motor neuron disease with features resembling prion diseases and amyotrophic lateral sclerosis. With the neurovirulent murine leukemia virus (MLV) FrCasE, Env protein expression within glia leads to postsynaptic vacuolation, cellular effacement, and neuronal loss in the absence of neuroinflammation. To understand the physiological changes associated with MLV-induced spongiosis, and its neuronal specificity, we employed patch-clamp recordings and voltage-sensitive dye imaging in brain slices of the mouse inferior colliculus (IC), a midbrain nucleus that undergoes extensive spongiosis. IC neurons characterized by postinhibitory rebound firing (PIR) were selectively affected in FrCasE-infected mice. Coincident with Env expression in microglia and in glia characterized by NG2 proteoglycan expression (NG2 cells), rebound neurons (RNs) lost PIR, became hyperexcitable, and were reduced in number. PIR loss and hyperexcitability were reversed by raising internal calcium buffer concentrations in RNs. PIR-initiated rhythmic circuits were disrupted, and spontaneous synchronized bursting and prolonged depolarizations were widespread. Other IC neuron cell types and circuits within the same degenerative environment were unaffected. Antagonists of NMDA and/or AMPA receptors reduced burst firing in the IC but did not affect prolonged depolarizations. Antagonists of L-type calcium channels abolished both bursts and slow depolarizations. IC infection by the nonneurovirulent isogenic virus Friend 57E (Fr57E), whose Env protein is structurally similar to FrCasE, showed no RN hyperactivity or cell loss; however, PIR latency increased. These findings suggest that spongiform neurodegeneration arises from the unique excitability of RNs, their local regulation by glia, and the disruption of this relationship by glial expression of abnormal protein. PMID:25252336

The Role of the Lateral Habenula in Punishment

PubMed Central

Jean-Richard Dit Bressel, Philip; McNally, Gavan P.

2014-01-01

The lateral habenula (LHb) is a small epithalamic structure that projects via the fasciculus retroflexus to the midbrain. The LHb is known to modulate midbrain dopamine (DA) neurons, including inhibition of ventral tegmental area (VTA) neurons via glutamatergic excitation of the GABAergic rostromedial tegmental nucleus (RMTg). A variety of lines of evidence show activity in LHb and the LHb-RMTg pathway is correlated with, and is sufficient to support, punishment learning. However, it is not immediately clear whether LHb is necessary for punishment. Here we used a within-subjects punishment task to assess the role of LHb in the acquisition and expression of punishment as well as in aversive choice. Rats that pressed two individually presented levers for pellet rewards rapidly suppressed responding to one lever if it also caused footshock deliveries (punished lever) but continued pressing a second lever that did not cause footshock (unpunished lever). Infusions of an AMPA receptor antagonist (NBQX) into LHb had no effect on the acquisition or expression of this punishment, or on aversive choice, but did increase locomotion. Infusion of the sodium channel blocker bupivacaine likewise had no effect on expression of punishment. However, infusion of the calcium channel blocker mibefradil did affect expression of punishment by significantly decreasing the latency with which rats responded on the punished lever and significantly increasing unpunished lever-pressing. Taken together, these findings indicate that the LHb plays a limited role in punishment, influencing only latency to respond. This role is linked to calcium channel permeability and not AMPA receptor or sodium channel permeability. PMID:25365401

The role of the lateral habenula in punishment.

PubMed

Jean-Richard Dit Bressel, Philip; McNally, Gavan P

2014-01-01

The lateral habenula (LHb) is a small epithalamic structure that projects via the fasciculus retroflexus to the midbrain. The LHb is known to modulate midbrain dopamine (DA) neurons, including inhibition of ventral tegmental area (VTA) neurons via glutamatergic excitation of the GABAergic rostromedial tegmental nucleus (RMTg). A variety of lines of evidence show activity in LHb and the LHb-RMTg pathway is correlated with, and is sufficient to support, punishment learning. However, it is not immediately clear whether LHb is necessary for punishment. Here we used a within-subjects punishment task to assess the role of LHb in the acquisition and expression of punishment as well as in aversive choice. Rats that pressed two individually presented levers for pellet rewards rapidly suppressed responding to one lever if it also caused footshock deliveries (punished lever) but continued pressing a second lever that did not cause footshock (unpunished lever). Infusions of an AMPA receptor antagonist (NBQX) into LHb had no effect on the acquisition or expression of this punishment, or on aversive choice, but did increase locomotion. Infusion of the sodium channel blocker bupivacaine likewise had no effect on expression of punishment. However, infusion of the calcium channel blocker mibefradil did affect expression of punishment by significantly decreasing the latency with which rats responded on the punished lever and significantly increasing unpunished lever-pressing. Taken together, these findings indicate that the LHb plays a limited role in punishment, influencing only latency to respond. This role is linked to calcium channel permeability and not AMPA receptor or sodium channel permeability.

Activation of TRPC channels contributes to OA-NO2-induced responses in guinea-pig dorsal root ganglion neurons

PubMed Central

Zhang, Xiulin; Beckel, Jonathan M; Daugherty, Stephanie L; Wang, Ting; Woodcock, Stephen R; Freeman, Bruce A; de Groat, William C

2014-01-01

Effects of nitro-oleic acid (OA-NO2) on TRP channels were examined in guinea-pig dissociated dorsal root ganglia (DRG) neurons using calcium imaging and patch clamp techniques. OA-NO2 increased intracellular Ca2+ in 60–80% DRG neurons. 1-Oleoyl-2acetyl-sn-glycerol (OAG), a TRPC agonist, elicited responses in 36% of OA-NO2-sensitive neurons while capsaicin (TRPV1 agonist) or allyl-isothiocyanate (AITC, TRPA1 agonist) elicited responses in only 16% and 10%, respectively, of these neurons. A TRPV1 antagonist (diarylpiperazine, 5 μm) in combination with a TRPA1 antagonist (HC-030031, 30 μm) did not change the amplitude of the Ca2+ transients or percentage of neurons responding to OA-NO2; however, a reducing agent DTT (50 mm) or La3+ (50 μm) completely abolished OA-NO2 responses. OA-NO2 also induced a transient inward current associated with a membrane depolarization followed by a prolonged outward current and hyperpolarization in 80% of neurons. The reversal potentials of inward and outward currents were approximately −20 mV and −60 mV, respectively. Inward current was reduced when extracellular Na+ was absent, but unchanged by niflumic acid (100 μm), a Cl− channel blocker. Outward current was abolished in the absence of extracellular Ca2+ or a combination of two Ca2+-activated K+ channel blockers (iberiotoxin, 100 nm and apamin, 1 μm). BTP2 (1 or 10 μm), a broad spectrum TRPC antagonist, or La3+ (50 μm) completely abolished OA-NO2 currents. RT-PCR performed on mRNA extracted from DRGs revealed the expression of all seven subtypes of TRPC channels. These results support the hypothesis that OA-NO2 activates TRPC channels other than the TRPV1 and TRPA1 channels already known to be targets in rat and mouse sensory neurons and challenge the prevailing view that electrophilic compounds act specifically on TRPA1 or TRPV1 channels. The modulation of sensory neuron excitability via actions on multiple TRP channels can contribute to the anti-inflammatory effect of OA-NO2. PMID:25128576

Antagonistic versus non-antagonistic models of balancing selection: Characterizing the relative timescales and hitchhiking effects of partial selective sweeps

PubMed Central

Connallon, Tim; Clark, Andrew G.

2012-01-01

Antagonistically selected alleles -- those with opposing fitness effects between sexes, environments, or fitness components -- represent an important component of additive genetic variance in fitness-related traits, with stably balanced polymorphisms often hypothesized to contribute to observed quantitative genetic variation. Balancing selection hypotheses imply that intermediate-frequency alleles disproportionately contribute to genetic variance of life history traits and fitness. Such alleles may also associate with population genetic footprints of recent selection, including reduced genetic diversity and inflated linkage disequilibrium at linked, neutral sites. Here, we compare the evolutionary dynamics of different balancing selection models, and characterize the evolutionary timescale and hitchhiking effects of partial selective sweeps generated under antagonistic versus non-antagonistic (e.g., overdominant and frequency-dependent selection) processes. We show that that the evolutionary timescales of partial sweeps tend to be much longer, and hitchhiking effects are drastically weaker, under scenarios of antagonistic selection. These results predict an interesting mismatch between molecular population genetic and quantitative genetic patterns of variation. Balanced, antagonistically selected alleles are expected to contribute more to additive genetic variance for fitness than alleles maintained by classic, non-antagonistic mechanisms. Nevertheless, classical mechanisms of balancing selection are much more likely to generate strong population genetic signatures of recent balancing selection. PMID:23461340

Two Mechanisms Involved in Trigeminal CGRP Release: Implications for Migraine Treatment

PubMed Central

Durham, Paul L.; Masterson, Caleb G.

2012-01-01

Objective The goal of this study was to better understand the cellular mechanisms involved in proton stimulation of calcitonin gene-related peptide (CGRP) secretion from cultured trigeminal neurons by investigating the effects of two anti-migraine therapies, onabotulinumtoxin A and rizatriptan. Background Stimulated CGRP release from peripheral and central terminating processes of trigeminal ganglia neurons is implicated in migraine pathology by promoting inflammation and nociception. Based on models of migraine pathology, several inflammatory molecules including protons are thought to facilitate sensitization and activation of trigeminal nociceptive neurons and stimulate CGRP secretion. Despite the reported efficacy of triptans and onabotulinumtoxinA to treat acute and chronic migraine, respectively, a substantial number of migraneurs do not get adequate relief with these therapies. A possible explanation is that triptans and onabutulinumtoxinA are not able to block proton mediated CGRP secretion. Methods CGRP secretion from cultured primary trigeminal ganglia neurons was quantitated by radioimmunoassay while intracellular calcium and sodium levels were measured in neurons via live cell imaging using Fura2-AM and SBFI-AM, respectively. The expression of ASIC3 was determined by immunocytochemistry and western blot analysis. In addition, the involvement of ASICs in mediating proton stimulation of CGRP was investigated using the potent and selective ASIC3 inhibitor APETx2. Results While KCl caused a significant increase in CGRP secretion that was significantly repressed by treatment with EGTA, onabotulinumtoxinA, and rizatriptan, the stimulatory effect of protons (pH 5.5) was not suppressed by EGTA, onabotulinumtoxinA, or rizatriptan. In addition, while KCl caused a transient increase in intracellular calcium levels that was blocked by EGTA, no appreciable change in calcium levels was observed with proton treatment. However, protons did significantly increase the intracellular level of sodium ions. Under our culture conditions, ASIC3 was shown to be expressed in most trigeminal ganglion neurons. Importantly, proton stimulation of CGRP secretion was repressed by pretreatment with the ASIC3 inhibitor APETx2, but not the TRPV1 antagonist capsazepine. Conclusions Our findings provide evidence that proton regulated release of CGRP from trigeminal neurons utilizes a different mechanism than the calcium and SNAP-25 dependent pathways that are inhibited by the anti-migraine therapies rizatriptan and onabotulinumtoxinA. PMID:23095108

A General Population Genetic Framework for Antagonistic Selection That Accounts for Demography and Recurrent Mutation

PubMed Central

Connallon, Tim; Clark, Andrew G.

2012-01-01

Antagonistic selection—where alleles at a locus have opposing effects on male and female fitness (“sexual antagonism”) or between components of fitness (“antagonistic pleiotropy”)—might play an important role in maintaining population genetic variation and in driving phylogenetic and genomic patterns of sexual dimorphism and life-history evolution. While prior theory has thoroughly characterized the conditions necessary for antagonistic balancing selection to operate, we currently know little about the evolutionary interactions between antagonistic selection, recurrent mutation, and genetic drift, which should collectively shape empirical patterns of genetic variation. To fill this void, we developed and analyzed a series of population genetic models that simultaneously incorporate these processes. Our models identify two general properties of antagonistically selected loci. First, antagonistic selection inflates heterozygosity and fitness variance across a broad parameter range—a result that applies to alleles maintained by balancing selection and by recurrent mutation. Second, effective population size and genetic drift profoundly affect the statistical frequency distributions of antagonistically selected alleles. The “efficacy” of antagonistic selection (i.e., its tendency to dominate over genetic drift) is extremely weak relative to classical models, such as directional selection and overdominance. Alleles meeting traditional criteria for strong selection (Nes >> 1, where Ne is the effective population size, and s is a selection coefficient for a given sex or fitness component) may nevertheless evolve as if neutral. The effects of mutation and demography may generate population differences in overall levels of antagonistic fitness variation, as well as molecular population genetic signatures of balancing selection. PMID:22298707

A general population genetic framework for antagonistic selection that accounts for demography and recurrent mutation.

PubMed

Connallon, Tim; Clark, Andrew G

2012-04-01

Antagonistic selection--where alleles at a locus have opposing effects on male and female fitness ("sexual antagonism") or between components of fitness ("antagonistic pleiotropy")--might play an important role in maintaining population genetic variation and in driving phylogenetic and genomic patterns of sexual dimorphism and life-history evolution. While prior theory has thoroughly characterized the conditions necessary for antagonistic balancing selection to operate, we currently know little about the evolutionary interactions between antagonistic selection, recurrent mutation, and genetic drift, which should collectively shape empirical patterns of genetic variation. To fill this void, we developed and analyzed a series of population genetic models that simultaneously incorporate these processes. Our models identify two general properties of antagonistically selected loci. First, antagonistic selection inflates heterozygosity and fitness variance across a broad parameter range--a result that applies to alleles maintained by balancing selection and by recurrent mutation. Second, effective population size and genetic drift profoundly affect the statistical frequency distributions of antagonistically selected alleles. The "efficacy" of antagonistic selection (i.e., its tendency to dominate over genetic drift) is extremely weak relative to classical models, such as directional selection and overdominance. Alleles meeting traditional criteria for strong selection (N(e)s > 1, where N(e) is the effective population size, and s is a selection coefficient for a given sex or fitness component) may nevertheless evolve as if neutral. The effects of mutation and demography may generate population differences in overall levels of antagonistic fitness variation, as well as molecular population genetic signatures of balancing selection.

PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion.

PubMed

Li, Li; Zhao, Lei; Wang, Yang; Ma, Ke-tao; Shi, Wen-yan; Wang, Ying-zi; Si, Jun-qiang

2015-02-01

The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²⁺-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.

Antihypertensive drugs.

PubMed

Laurent, Stéphane

2017-10-01

Successful treatment of hypertension is possible with limited side effects given the availability of multiple antihypertensive drug classes. This review describes the various pharmacological classes of antihypertensive drugs, under two major aspects: their mechanisms of action and side effects. The mechanism of action is analysed through a pharmacological approach, i.e. the molecular receptor targets, the various sites along the arterial system, and the extra-arterial sites of action, in order to better understand in which type of hypertension a given pharmacological class of antihypertensive drug is most indicated. In addition, side effects are described and explained through their pharmacological mechanisms, in order to better understand their mechanism of occurrence and in which patients drugs are contra-indicated. This review does not address the effectiveness of monotherapies in large randomized clinical trials and combination therapies, since these are the matters of other articles of the present issue. Five major pharmacological classes of antihypertensive drugs are detailed here: beta-blockers, diuretics, angiotensin converting enzyme inhibitors, angiotensin II receptor antagonists, and calcium channel blockers. Four additional pharmacological classes are described in a shorter manner: renin inhibitors, alpha-adrenergic receptor blockers, centrally acting agents, and direct acting vasodilators. Copyright © 2017. Published by Elsevier Ltd.

Transient Receptor Potential Vanilloid 4 Activation-Induced Increase in Glycine-Activated Current in Mouse Hippocampal Pyramidal Neurons.

PubMed

Qi, Mengwen; Wu, Chunfeng; Wang, Zhouqing; Zhou, Li; Men, Chen; Du, Yimei; Huang, Songming; Chen, Lei; Chen, Ling

2018-01-01

Glycine plays an important role in regulating hippocampal inhibitory/ excitatory neurotransmission through activating glycine receptors (GlyRs) and acting as a co-agonist of N-methyl-d-aspartate-type glutamate receptors. Activation of transient receptor potential vanilloid 4 (TRPV4) is reported to inhibit hippocampal A-type γ-aminobutyric acid receptor, a ligand-gated chloride ion channel. GlyRs are also ligand-gated chloride ion channels and this paper aimed to explore whether activation of TRPV4 could modulate GlyRs. Whole-cell patch clamp recording was employed to record glycine-activated current (IGly) and Western blot was conducted to assess GlyRs subunits protein expression. Application of TRPV4 agonist (GSK1016790A or 5,6-EET) increased IGly in mouse hippocampal CA1 pyramidal neurons. This action was blocked by specific antagonists of TRPV4 (RN-1734 or HC-067047) and GlyR (strychnine), indicating that activation of TRPV4 increases strychnine-sensitive GlyR function in mouse hippocampal pyramidal neurons. GSK1016790A-induced increase in IGly was significantly attenuated by protein kinase C (PKC) (BIM II or D-sphingosine) or calcium/calmodulin-dependent protein kinase II (CaMKII) (KN-62 or KN-93) antagonists but was unaffected by protein kinase A or protein tyrosine kinase antagonists. Finally, hippocampal protein levels of GlyR α1 α2, α3 and β subunits were not changed by treatment with GSK1016790A for 30 min or 1 h, but GlyR α2, α3 and β subunits protein levels increased in mice that were intracerebroventricularly (icv.) injected with GSK1016790A for 5 d. Activation of TRPV4 increases GlyR function and expression, and PKC and CaMKII signaling pathways are involved in TRPV4 activation-induced increase in IGly. This study indicates that GlyRs may be effective targets for TRPV4-induced modulation of hippocampal inhibitory neurotransmission. © 2018 The Author(s). Published by S. Karger AG, Basel.

Structure-guided design of a high-affinity platelet integrin αIIbβ3 receptor antagonist that disrupts Mg²⁺ binding to the MIDAS.

PubMed

Zhu, Jieqing; Choi, Won-Seok; McCoy, Joshua G; Negri, Ana; Zhu, Jianghai; Naini, Sarasija; Li, Jihong; Shen, Min; Huang, Wenwei; Bougie, Daniel; Rasmussen, Mark; Aster, Richard; Thomas, Craig J; Filizola, Marta; Springer, Timothy A; Coller, Barry S

2012-03-14

An integrin found on platelets, α(IIb)β(3) mediates platelet aggregation, and α(IIb)β(3) antagonists are effective antithrombotic agents in the clinic. Ligands bind to integrins in part by coordinating a magnesium ion (Mg(2+)) located in the β subunit metal ion-dependent adhesion site (MIDAS). Drugs patterned on the integrin ligand sequence Arg-Gly-Asp have a basic moiety that binds the α(IIb) subunit and a carboxyl group that coordinates the MIDAS Mg(2+) in the β(3) subunits. They induce conformational changes in the β(3) subunit that may have negative consequences such as exposing previously hidden epitopes and inducing the active conformation of the receptor. We recently reported an inhibitor of α(IIb)β(3) (RUC-1) that binds exclusively to the α(IIb) subunit; here, we report the structure-based design and synthesis of RUC-2, a RUC-1 derivative with a ~100-fold higher affinity. RUC-2 does not induce major conformational changes in β(3) as judged by monoclonal antibody binding, light scattering, gel chromatography, electron microscopy, and a receptor priming assay. X-ray crystallography of the RUC-2-α(IIb)β(3) headpiece complex in 1 mM calcium ion (Ca(2+))/5 mM Mg(2+) at 2.6 Å revealed that RUC-2 binds to α(IIb) the way RUC-1 does, but in addition, it binds to the β(3) MIDAS residue glutamic acid 220, thus displacing Mg(2+) from the MIDAS. When the Mg(2+) concentration was increased to 20 mM, however, Mg(2+) was identified in the MIDAS and RUC-2 was absent. RUC-2's ability to inhibit ligand binding and platelet aggregation was diminished by increasing the Mg(2+) concentration. Thus, RUC-2 inhibits ligand binding by a mechanism different from that of all other α(IIb)β(3) antagonists and may offer advantages as a therapeutic agent.

Synergistic Antiproliferative Effects of Combined γ-Tocotrienol and PPARγ Antagonist Treatment Are Mediated through PPARγ-Independent Mechanisms in Breast Cancer Cells

PubMed Central

Sylvester, Paul W.

2014-01-01

Previous findings showed that the anticancer effects of combined γ-tocotrienol and peroxisome proliferator activated receptor γ (PPARγ) antagonist treatment caused a large reduction in PPARγ expression. However, other studies suggest that the antiproliferative effects of γ-tocotrienol and/or PPARγ antagonists are mediated, at least in part, through PPARγ-independent mechanism(s). Studies were conducted to characterize the role of PPARγ in mediating the effects of combined treatment of γ-tocotrienol with PPARγ agonists or antagonists on the growth of PPARγ negative +SA mammary cells and PPARγ-positive and PPARγ-silenced MCF-7 and MDA-MB-231 breast cancer cells. Combined treatment of γ-tocotrienol with PPARγ antagonist decreased, while combined treatment of γ-tocotrienol with PPARγ agonist increased, growth of all cancer cells. However, treatment with high doses of 15d-PGJ2, an endogenous natural ligand for PPARγ, had no effect on cancer cell growth. Western blot and qRT-PCR studies showed that the growth inhibitory effects of combined γ-tocotrienol and PPARγ antagonist treatment decreased cyclooxygenase (COX-2), prostaglandin synthase (PGDS), and prostaglandin D2 (PGD2) synthesis. In conclusion, the anticancer effects of combined γ-tocotrienol and PPARγ antagonists treatment in PPARγ negative/silenced breast cancer cells are mediated through PPARγ-independent mechanisms that are associated with a downregulation in COX-2, PGDS, and PGD2 synthesis. PMID:24729783

The Influence of Glutamate on Axonal Compound Action Potential In Vitro.

PubMed

Abouelela, Ahmed; Wieraszko, Andrzej

2016-01-01

Background  Our previous experiments demonstrated modulation of the amplitude of the axonal compound action potential (CAP) by electrical stimulation. To verify assumption that glutamate released from axons could be involved in this phenomenon, the modification of the axonal CAP induced by glutamate was investigated. Objectives  The major objective of this research is to verify the hypothesis that axonal activity would trigger the release of glutamate, which in turn would interact with specific axonal receptors modifying the amplitude of the action potential. Methods  Segments of the sciatic nerve were exposed to exogenous glutamate in vitro, and CAP was recorded before and after glutamate application. In some experiments, the release of radioactive glutamate analog from the sciatic nerve exposed to exogenous glutamate was also evaluated. Results  The glutamate-induced increase in CAP was blocked by different glutamate receptor antagonists. The effect of glutamate was not observed in Ca-free medium, and was blocked by antagonists of calcium channels. Exogenous glutamate, applied to the segments of sciatic nerve, induced the release of radioactive glutamate analog, demonstrating glutamate-induced glutamate release. Immunohistochemical examination revealed that axolemma contains components necessary for glutamatergic neurotransmission. Conclusion  The proteins of the axonal membrane can under the influence of electrical stimulation or exogenous glutamate change membrane permeability and ionic conductance, leading to a change in the amplitude of CAP. We suggest that increased axonal activity leads to the release of glutamate that results in changes in the amplitude of CAPs.

Ketamine alters cortical integration of GABAergic interneurons and induces long-term sex-dependent impairments in transgenic Gad67-GFP mice.

PubMed

Aligny, C; Roux, C; Dourmap, N; Ramdani, Y; Do-Rego, J-C; Jégou, S; Leroux, P; Leroux-Nicollet, I; Marret, S; Gonzalez, B J

2014-07-03

Ketamine, a non-competitive N-methyl-D-aspartate (NMDA) antagonist, widely used as an anesthetic in neonatal pediatrics, is also an illicit drug named Super K or KitKat consumed by teens and young adults. In the immature brain, despite several studies indicating that NMDA antagonists are neuroprotective against excitotoxic injuries, there is more and more evidence indicating that these molecules exert a deleterious effect by suppressing a trophic function of glutamate. In the present study, we show using Gad67-GFP mice that prenatal exposure to ketamine during a time-window in which GABAergic precursors are migrating results in (i) strong apoptotic death in the ganglionic eminences and along the migratory routes of GABAergic interneurons; (ii) long-term deficits in interneuron density, dendrite numbers and spine morphology; (iii) a sex-dependent deregulation of γ-aminobutyric acid (GABA) levels and GABA transporter expression; (iv) sex-dependent changes in the response to glutamate-induced calcium mobilization; and (v) the long-term sex-dependent behavioral impairment of locomotor activity. In conclusion, using a preclinical approach, the present study shows that ketamine exposure during cortical maturation durably affects the integration of GABAergic interneurons by reducing their survival and differentiation. The resulting molecular, morphological and functional modifications are associated with sex-specific behavioral deficits in adults. In light of the present data, it appears that in humans, ketamine could be deleterious for the development of the brain of preterm neonates and fetuses of addicted pregnant women.

Proton pump inhibitors are associated with lower gastrointestinal tract bleeding in low-dose aspirin users with ischaemic heart disease.

PubMed

Miyake, Kazumasa; Akimoto, Teppei; Hanada, Yuriko; Nagoya, Hiroyuki; Kodaka, Yasuhiro; Ueki, Nobue; Kusunoki, Masafumi; Kawagoe, Tetsuro; Futagami, Seiji; Takahashi, Yasuhiro; Takano, Hitoshi; Sakamoto, Choitsu

2015-09-01

Impact of acid suppressants on lower gastrointestinal bleeding remains unclear in low-dose aspirin users; we aimed to investigate this relationship. Retrospective cohort study of low-dose aspirin users who underwent coronary angiography for ischaemic heart disease in our institution between October 2005 and December 2006; patients were evaluated for upper or lower gastrointestinal bleedings within 3 years post-angiography. 538 patients were enrolled (males, 74.4%; mean age 67.4±10.6 years). Risk for upper gastrointestinal bleeding decreased with concomitant use of statins (HR, 0.37; 95% CI, 0.15-0.89), calcium channel blockers (HR, 0.29; 95% CI, 0.10-0.85), and histamine-2 receptor antagonists (HR, 0.26; 95% CI, 0.08-0.89). Concomitant use of proton pump inhibitors tended to decrease risk of upper gastrointestinal bleeding (HR, 0.27; 95% CI, 0.06-1.18). Risk for lower gastrointestinal bleeding increased with both concomitant use of warfarin (HR, 15.68; 95% CI, 4.43-55.53) and proton pump inhibitors (HR, 6.55; 95% CI, 2.01-21.32), but not with histamine-2 receptor antagonists. Hyperuricemia lowered risk for lower gastrointestinal bleeding (HR, 0.12; 95% CI, 0.02-0.88). In low-dose aspirin users, concomitant use of proton pump inhibitors increased lower gastrointestinal bleeding risk, independent from effects on upper gastrointestinal bleeding. Copyright © 2015 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.

Wnt and the Wnt signaling pathway in bone development and disease

PubMed Central

Wang, Yiping; Li, Yi-Ping; Paulson, Christie; Shao, Jian-Zhong; Zhang, Xiaoling; Wu, Mengrui; Chen, Wei

2014-01-01

Wnt signaling affects both bone modeling, which occurs during development, and bone remodeling, which is a lifelong process involving tissue renewal. Wnt signals are especially known to affect the differentiation of osteoblasts. In this review, we summarize recent advances in understanding the mechanisms of Wnt signaling, which is divided into two major branches: the canonical pathway and the noncanonical pathway. The canonical pathway is also called the Wnt/β-catenin pathway. There are two major noncanonical pathways: the Wnt-planar cell polarity pathway (Wnt-PCP pathway) and the Wnt-calcium pathway (Wnt-Ca2+ pathway). This review also discusses how Wnt ligands, receptors, intracellular effectors, transcription factors, and antagonists affect both the bone modeling and bone remodeling processes. We also review the role of Wnt ligands, receptors, intracellular effectors, transcription factors, and antagonists in bone as demonstrated in mouse models. Disrupted Wnt signaling is linked to several bone diseases, including osteoporosis, van Buchem disease, and sclerosteosis. Studying the mechanism of Wnt signaling and its interactions with other signaling pathways in bone will provide potential therapeutic targets to treat these bone diseases. PMID:24389191

Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP)-mediated Calcium Signaling and Arrhythmias in the Heart Evoked by β-Adrenergic Stimulation*♦

PubMed Central

Nebel, Merle; Schwoerer, Alexander P.; Warszta, Dominik; Siebrands, Cornelia C.; Limbrock, Ann-Christin; Swarbrick, Joanna M.; Fliegert, Ralf; Weber, Karin; Bruhn, Sören; Hohenegger, Martin; Geisler, Anne; Herich, Lena; Schlegel, Susan; Carrier, Lucie; Eschenhagen, Thomas; Potter, Barry V. L.; Ehmke, Heimo; Guse, Andreas H.

2013-01-01

Nicotinic acid adenine dinucleotide phosphate (NAADP) is the most potent Ca2+-releasing second messenger known to date. Here, we report a new role for NAADP in arrhythmogenic Ca2+ release in cardiac myocytes evoked by β-adrenergic stimulation. Infusion of NAADP into intact cardiac myocytes induced global Ca2+ signals sensitive to inhibitors of both acidic Ca2+ stores and ryanodine receptors and to NAADP antagonist BZ194. Furthermore, in electrically paced cardiac myocytes BZ194 blocked spontaneous diastolic Ca2+ transients caused by high concentrations of the β-adrenergic agonist isoproterenol. Ca2+ transients were recorded both as increases of the free cytosolic Ca2+ concentration and as decreases of the sarcoplasmic luminal Ca2+ concentration. Importantly, NAADP antagonist BZ194 largely ameliorated isoproterenol-induced arrhythmias in awake mice. We provide strong evidence that NAADP-mediated modulation of couplon activity plays a role for triggering spontaneous diastolic Ca2+ transients in isolated cardiac myocytes and arrhythmias in the intact animal. Thus, NAADP signaling appears an attractive novel target for antiarrhythmic therapy. PMID:23564460

NMDA receptor antagonists inhibit catalepsy induced by either dopamine D1 or D2 receptor antagonists.

PubMed

Moore, N A; Blackman, A; Awere, S; Leander, J D

1993-06-11

In the present study, we investigated the ability of NMDA receptor antagonists to inhibit catalepsy induced by haloperidol, or SCH23390 and clebopride, selective dopamine D1 and D2 receptor antagonists respectively. Catalepsy was measured by recording the time the animal remained with its forepaws placed over a rod 6 cm above the bench. Pretreatment with either the non-competitive NMDA receptor antagonist, MK-801 (0.25-0.5 mg/kg i.p.) or the competitive antagonist, LY274614 (10-20 mg/kg i.p.) reduced the cataleptic response produced by haloperidol (10 mg/kg), SCH23390 (2.5-10 mg/kp i.p.) or clebopride (5-20 mg/kg i.p.). This demonstrates that NMDA receptor antagonists will reduce both dopamine D1 and D2 receptor antagonist-induced catalepsy. Muscle relaxant doses of chlordiazepoxide (10 mg/kg i.p.) failed to reduce the catalepsy induced by haloperidol, suggesting that the anticataleptic effect of the NMDA receptor antagonists was not due to a non-specific action. These results support the hypothesis that NMDA receptor antagonists may have beneficial effects in disorders involving reduced dopaminergic function, such as Parkinson's disease.

Acute exposure to progesterone attenuates cardiac contraction by modifying myofilament calcium sensitivity in the female mouse heart

PubMed Central

Feridooni, Hirad A.; MacDonald, Jennifer K.; Ghimire, Anjali; Pyle, W. Glen

2017-01-01

Acute application of progesterone attenuates cardiac contraction, although the underlying mechanisms are unclear. We investigated whether progesterone modified contraction in isolated ventricular myocytes and identified the Ca2+ handling mechanisms involved in female C57BL/6 mice (6–9 mo; sodium pentobarbital anesthesia). Cells were field-stimulated (4 Hz; 37°C) and exposed to progesterone (0.001–10.0 μM) or vehicle (35 min). Ca2+ transients (fura-2) and cell shortening were recorded simultaneously. Maximal concentrations of progesterone inhibited peak contraction by 71.4% (IC50 = 160 ± 50 nM; n = 12) and slowed relaxation by 75.4%. By contrast, progesterone had no effect on amplitudes or time courses of underlying Ca2+ transients. Progesterone (1 µM) also abbreviated action potential duration. When the duration of depolarization was controlled by voltage-clamp, progesterone attenuated contraction and slowed relaxation but did not affect Ca2+ currents, Ca2+ transients, sarcoplasmic reticulum (SR) content, or fractional release of SR Ca2+. Actomyosin MgATPase activity was assayed in myofilaments from hearts perfused with progesterone (1 μM) or vehicle (35 min). While maximal responses to Ca2+ were not affected by progesterone, myofilament Ca2+ sensitivity was reduced (EC50 = 0.94 ± 0.01 µM for control, n = 7 vs. 1.13 ± 0.05 μM for progesterone, n = 6; P < 0.05) and progesterone increased phosphorylation of myosin binding protein C. The effects on contraction were inhibited by lonaprisan (progesterone receptor antagonist) and levosimendan (Ca2+ sensitizer). Unlike results in females, progesterone had no effect on contraction or myofilament Ca2+ sensitivity in age-matched male mice. These data indicate that progesterone reduces myofilament Ca2+ sensitivity in female hearts, which may exacerbate manifestations of cardiovascular disease late in pregnancy when progesterone levels are high. NEW & NOTEWORTHY We investigated myocardial effects of acute application of progesterone. In females, but not males, progesterone attenuates and slows cardiomyocyte contraction with no effect on calcium transients. Progesterone also reduces myofilament calcium sensitivity in female hearts. This may adversely affect heart function, especially when serum progesterone levels are high in pregnancy. Listen to this article’s corresponding podcast at https://ajpheart.podbean.com/e/acute-progesterone-modifies-cardiac-contraction/. PMID:27793852