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Sample records for mediate mechanical hyperalgesia

  1. SPINAL CORD MECHANISMS MEDIATING BEHAVIORAL HYPERALGESIA INDUCED BY NEUROKININ-1 TACHYKININ RECEPTOR ACTIVATION IN THE ROSTRAL VENTROMEDIAL MEDULLA

    PubMed Central

    Lagraize, S. C.; Guo, W.; Yang, K.; Wei, F.; Ren, K.; Dubner, R.

    2010-01-01

    Hyperalgesia in animal injury models is linked to activation of descending raphespinal modulatory circuits originating in the rostral ventromedial medulla (RVM). A neurokinin-1 (NK-1) receptor antagonist microinjected into the RVM before or after inflammation produced by complete Freund’s adjuvant (CFA) resulted in an attenuation of thermal hyperalgesia. A transient (acute) or a continuous infusion of Substance P (SP) microinjected into the RVM of non-inflamed animals led to similar pain hypersensitivity. Intrathecal pretreatment or post-treatment of a 5-HT3 receptor antagonist (Y-25130 or ondansetron) blocked the SP-induced hyperalgesia. The SP-induced hyperalgesia was both GABAA and NMDA receptor-dependent after pre- and post-treatment with selective antagonists at the spinal level. A microinjection of SP into the RVM also led to increased NMDA NR1 receptor subunit phosphorylation in spinal cord tissue. The GABAA receptor-mediated hyperalgesia involved a shift in the anionic gradient in dorsal horn nociceptive neurons and an increase in phosphorylated NKCC1 protein (isoform of the Na-K-Cl cotransporter). Following a low dose of SP infused into the RVM, intrathecal muscimol (GABAA agonist) increased SP-induced thermal hyperalgesia, phosphorylated NKCC1 protein expression, and NMDA NR1 subunit phosphorylation in the spinal cord. The thermal hyperalgesia was blocked by intrathecal gabazine, the GABAA receptor antagonist, and MK-801, the NMDA receptor channel blocker. These findings indicate that NK-1 receptors in the RVM are involved in SP-induced thermal hyperalgesia, this hyperalgesia is 5-HT3-receptor dependent at the spinal level, and involves the functional interaction of spinal GABAA and NMDA receptors. PMID:20888891

  2. Mechanical Stimulation Enhances Endothelin-1 Hyperalgesia

    PubMed Central

    Joseph, Elizabeth K.; Gear, Robert W.; Levine, Jon D.

    2011-01-01

    When comparing a cumulative dose-response curve for endothelin-1 (ET-1)-induced mechanical hyperalgesia to the effect of individual doses (1 ng, 10 ng, 100 ng and 1 µg) administered in separate groups of rats, a marked difference was observed in the peak magnitude of hyperalgesia. Hyperalgesia was measured as decrease in the threshold for mechanically-induced withdrawal of the hind paw. The cumulative dosing protocol produced markedly greater maximum hyperalgesia. To determine whether this was due to the cumulative dosing protocol or to the repeated exposure to the mechanical test stimulus, we evaluated the impact of repeated testing on ET-1-induced mechanical hyperalgesia. While ET-1-induced mechanical hyperalgesia was dose- and time-dependent, repeated testing of nociceptive threshold, at 5 minute intervals, following a single dose of ET-1, produced further decrease in nociceptive threshold. This mechanical stimulation-induced enhancement of ET-1 hyperalgesia lasted only 3–4 hrs, while the hyperalgesia lasted in excess of 5 days. The stimulation-enhanced hyperalgesia also occurred after a second injection of ET-1, administered 24 hours after the initial dose. That this phenomenon is unique to ET-1 is suggested by the observation that while five additional, direct-acting hyperalgesic agents — PGE2, NGF, GDNF, IL-6 and TNFα — induced robust mechanical hyperalgesia, none produced mechanical stimulation-enhanced hyperalgesia. PMID:21277948

  3. Vascular Endothelial Cells Mediate Mechanical Stimulation-Induced Enhancement of Endothelin Hyperalgesia via Activation of P2X2/3 Receptors on Nociceptors

    PubMed Central

    Joseph, Elizabeth K.; Green, Paul G.; Bogen, Oliver; Alvarez, Pedro; Levine, Jon D.

    2013-01-01

    Endothelin-1 (ET-1) is unique amongst a broad range of hyperalgesic agents in that it induces hyperalgesia in rats that is markedly enhanced by repeated mechanical stimulation at the site of administration. Antagonists to the ET-1 receptors, ETA and ETB, attenuated both initial as well as stimulation-induced enhancement of hyperalgesia (SIEH) by endothelin. However, administering antisense oligodeoxynucleotide to attenuate ETA receptor expression on nociceptors attenuated ET-1 hyperalgesia, but had no effect on SIEH suggesting that this is mediated via a non-neuronal cell. Since vascular endothelial cells are both stretch-sensitive and express ETA and ETB receptors, we tested the hypothesis that SIEH is dependent on endothelial cells by impairing vascular endothelial function with octoxynol-9 administration; this procedure eliminated SIEH without attenuating ET-1 hyperalgesia. A role for protein kinase C epsilon (PKCε), a second messenger implicated in the induction and maintenance of chronic pain, was explored. Intrathecal antisense for PKCε did not inhibit either ET-1 hyperalgesia or SIEH, suggesting no role for neuronal PKCε; however, administration of a PKCε inhibitor at the site of testing selectively attenuated SIEH. Compatible with endothelial cells releasing ATP in response to mechanical stimulation, P2X2/3 receptor antagonists eliminated SIEH. The endothelium also appears to contribute to hyperalgesia in two ergonomic pain models (eccentric exercise and hind limb vibration) and in model of endometriosis. We propose that SIEH is produced by an effect of ET-1 on vascular endothelial cells, sensitizing its release of ATP in response to mechanical stimulation; ATP in turn acts at the nociceptor P2X2/3 receptor. PMID:23407944

  4. The transient receptor potential ankyrin-1 mediates mechanical hyperalgesia induced by the activation of B1 receptor in mice.

    PubMed

    Meotti, Flavia Carla; Figueiredo, Cláudia Pinto; Manjavachi, Marianne; Calixto, João B

    2017-02-01

    The kinin receptor B1 and the transient receptor potential ankyrin 1 (TRPA1) work as initiators and gatekeepers of nociception and inflammation. This study reports that the nociceptive transmission induced by activation of B1 receptor is dependent on TRPA1 ion channel. The mechanical hyperalgesia was induced by intrathecal (i.t.) injection of B1 agonist des-Arginine(9)-bradykinin (DABK) or TRPA1 agonist cinnamaldehyde and was evaluated by the withdrawal response after von Frey Hair application in the hind paw. After behavioral experiments, lumbar spinal cord and dorsal root ganglia (DRG) were harvested to assess protein expression and mRNA by immunohistochemistry and real time-PCR, respectively. The pharmacological antagonism (HC030031) or the down-regulation of TRPA1 greatly inhibited the mechanical hyperalgesia induced by DABK. Intrathecal injection of DABK up regulated the ionized calcium binding adaptor molecule (Iba-1) in lumbar spinal cord (L5-L6); TRPA1 protein and mRNA in lumbar spinal cord; and B1 receptor mRNA in both lumbar spinal cord and DRG. The knockdown of TRPA1 prevented microglia activation induced by DABK. Furthermore, the mechanical hyperalgesia induced by either DABK or by cinnamaldehyde was significantly reduced by inhibition of cyclooxygenase (COX), protein kinase C (PKC) or phospholipase C (PLC). In summary, this study revealed that TRPA1 positively modulates the mechanical hyperalgesia induced by B1 receptor activation in the spinal cord and that the classical GPCR downstream molecules PLC, diacylglycerol (DAG), 3,4,5-inositide phosphate (IP3) and PKC are involved in the nociceptive transmission triggered by these two receptors.

  5. Peripheral Receptor Mechanisms Underlying Orofacial Muscle Pain and Hyperalgesia

    NASA Astrophysics Data System (ADS)

    Saloman, Jami L.

    Musculoskeletal pain conditions, particularly those associated with temporomandibular joint and muscle disorders (TMD) are severely debilitating and affect approximately 12% of the population. Identifying peripheral nociceptive mechanisms underlying mechanical hyperalgesia, a prominent feature of persistent muscle pain, could contribute to the development of new treatment strategies for the management of TMD and other muscle pain conditions. This study provides evidence of functional interactions between ligand-gated channels, P2X3 and TRPV1/TRPA1, in trigeminal sensory neurons, and proposes that these interactions underlie the development of mechanical hyperalgesia. In the masseter muscle, direct P2X3 activation, via the selective agonist αβmeATP, induced a dose- and time-dependent hyperalgesia. Importantly, the αβmeATP-induced hyperalgesia was prevented by pretreatment of the muscle with a TRPV1 antagonist, AMG9810, or the TRPA1 antagonist, AP18. P2X3 was co-expressed with both TRPV1 and TRPA1 in masseter muscle afferents confirming the possibility for intracellular interactions. Moreover, in a subpopulation of P2X3 /TRPV1 positive neurons, capsaicin-induced Ca2+ transients were significantly potentiated following P2X3 activation. Inhibition of Ca2+-dependent kinases, PKC and CaMKII, prevented P2X3-mechanical hyperalgesia whereas blockade of Ca2+-independent PKA did not. Finally, activation of P2X3 induced phosphorylation of serine, but not threonine, residues in TRPV1 in trigeminal sensory neurons. Significant phosphorylation was observed at 15 minutes, the time point at which behavioral hyperalgesia was prominent. Similar data were obtained regarding another nonselective cation channel, the NMDA receptor (NMDAR). Our data propose P2X3 and NMDARs interact with TRPV1 in a facilitatory manner, which could contribute to the peripheral sensitization underlying masseter hyperalgesia. This study offers novel mechanisms by which individual pro-nociceptive ligand

  6. TRPA1, NMDA receptors and nitric oxide mediate mechanical hyperalgesia induced by local injection of magnesium sulfate into the rat hind paw.

    PubMed

    Srebro, Dragana P; Vučković, Sonja M; Savić Vujović, Katarina R; Prostran, Milica Š

    2015-02-01

    Previous studies have shown that while magnesium, an antagonist of the glutamate subtype of N-methyl-D-aspartate receptors, possesses analgesic properties, it can induce writhing in rodents. The aim of this study was to determine the effect and mechanism of action of local (intraplantar) administration of magnesium sulfate (MS) on the paw withdrawal threshold (PWT) to mechanical stimuli. The PWT was evaluated by the electronic von Frey test in male Wistar rats. Tested drugs were either co-administered intraplantarly (i.pl.) with MS or given into the contralateral paw to exclude systemic effects. MS at doses of 0.5, 1.5, 3 and 6.2 mg/paw (i.pl.) induced a statistically significant (as compared to 0.9% NaCl) and dose-dependent mechanical hyperalgesia. Only isotonic MS (250 mmol/l or 6.2% or 6.2 mg/paw) induced mechanical hyperalgesia that lasted at least six hours. Isotonic MS-induced mechanical hyperalgesia was reduced in a dose-dependent manner by co-injection of camphor, a non-selective TRPA1 antagonist (0.3, 1 and 2.5 μg/paw), MK-801, a NMDA receptor antagonist (0.001, 0.025 and 0.1 μg/paw), L-NAME, a non-selective nitric oxide (NO) synthase inhibitor (20, 50 and 100 μg/paw), ARL 17477, a selective neuronal NOS inhibitor (5.7 and 17 μg/paw), SMT, a selective inducible NOS inhibitor (1 and 2.78 μg/paw), and methylene blue, a guanylate cyclase inhibitor (5, 20 and 125 μg/paw). Drugs injected into the contralateral hind paw did not produce significant effects. These results suggest that an i.pl. injection of MS produces local peripheral mechanical hyperalgesia via activation of peripheral TRPA1 and NMDA receptors and peripheral production of NO.

  7. GRK2 in sensory neurons regulates epinephrine-induced signalling and duration of mechanical hyperalgesia.

    PubMed

    Wang, Huijing; Heijnen, Cobi J; Eijkelkamp, Niels; Garza Carbajal, Anibal; Schedlowski, Manfred; Kelley, Keith W; Dantzer, Robert; Kavelaars, Annemieke

    2011-07-01

    Epinephrine (EPI) contributes to hyperalgesia in inflammatory and stress conditions. EPI signals via adrenoceptors, which are regulated by G protein-coupled receptor kinase 2 (GRK2). We previously reported that GRK2 is decreased in nociceptors during chronic inflammation. Herein, we investigated whether GRK2 modulates EPI-induced mechanical and thermal hyperalgesia by using GRK2(+/-) mice, which express 50% of the GRK2 protein. We demonstrate for the first time that EPI-induced mechanical as well as thermal hyperalgesia is prolonged to approximately 21 days in GRK2(+/-) mice, whereas it lasts only 3 to 4 days in wild-type mice. Using cell- specific GRK2-deficient mice, we further show that a low level of GRK2 in primary sensory neurons is critical for this prolongation of EPI-induced hyperalgesia. Low GRK2 in microglia had only a small effect on EPI-induced hyperalgesia. Low GRK2 in astrocytes did not alter EPI-induced hyperalgesia. EPI-induced hyperalgesia was prolonged similarly in mice with tamoxifen-induced homozygous or heterozygous deletion of GRK2. In terms of EPI signalling pathways, the protein kinase A (PKA) inhibitor H-89 inhibited EPI-induced mechanical hyperalgesia in wild-type mice, whereas H-89 had no effect in mice with low GRK2 in sensory neurons (SNS-GRK2(+/-) mice). Conversely, intraplantar injection of the protein kinase Cε PKCε inhibitor TAT-PKC(εv1-2) inhibited hyperalgesia in sensory neuron specific (SNS)-GRK2(+/-) mice and not in wild-type mice. These results indicate that low GRK2 in primary sensory neurons switches EPI-induced signalling from a protein kinase A-dependent toward a PKCε-dependent pathway that ultimately mediates prolonged EPI-induced hyperalgesia.

  8. Pharmacological modulation of secondary mediator systems--cyclic AMP and cyclic GMP--on inflammatory hyperalgesia.

    PubMed

    Cunha, F Q; Teixeira, M M; Ferreira, S H

    1999-06-01

    1. The objective of the present paper was to evaluate the relevance of neuronal balance of cyclic AMP and cyclic GMP concentration for functional regulation of nociceptor sensitivity during inflammation. 2. Injection of PGE2 (10-100 ng paw-1) evoked a dose-dependent hyperalgesic effect which was mediated via a cyclic AMP-activated protein kinase (PKA) inasmuch as hyperalgesia was blocked by the PKA inhibitor H89. 3. The PDE4 inhibitor rolipram and RP73401, but not PDE3 and PDE5 inhibitors potentiated the hyperalgesic effects of PGE2. The hyperalgesic effect of dopamine was also enhanced by rolipram. Moreover, rolipram significantly potentiated hyperalgesia induced by carrageenan, bradykinin, TNF alpha, IL-1 beta, IL-6 and IL-8. This suggests that neuronal cyclic AMP mediates the prostanoid and sympathetic components of mechanical hyperalgesia. Moreover, in the neuron cyclic AMP is mainly metabolized by PDE4. 4. To examine the role of the NO/cyclic GMP pathway in modulating mechanical hyperalgesia, we tested the effects of the soluble guanylate cyclase inhibitor, ODQ. This substance counteracts the inhibitory effects of the NO donor, SNAP, on the hyperalgesia induced by PGE2. 5. The ODQ potentiated hyperalgesia induced by carrageenan, bradykinin, TNF alpha, IL-1 beta, IL-6 and IL-8. In contrast, ODQ had no significant effect on the hyperalgesia induced by PGE2 and dopamine. This indicates that the hyperalgesic cytokines may activate soluble guanylate cyclase, which down-regulate the ability of these substances to cause hyperalgesia. This event appears not to be mediated by prostaglandin or dopamine. 6. In conclusion, the results presented in this paper confirm an association between (i) hyperalgesia and elevated levels of cyclic AMP as well as (ii) antinociception and elevated levels of cyclic GMP. The intracellular levels of cyclic AMP that enhance hyperalgesia are controlled by the PDE4 isoform and appear to result in activation of protein kinase A whereas the

  9. Activation of TRPV1 and TRPA1 leads to muscle nociception and mechanical hyperalgesia.

    PubMed

    Ro, Jin Y; Lee, Jong-Seok; Zhang, Youping

    2009-08-01

    The involvement of TRPV1 and TRPA1 in mediating craniofacial muscle nociception and mechanical hyperalgesia was investigated in male Sprague-Dawley rats. First, we confirmed the expression of TRPV1 in masseter afferents in rat trigeminal ganglia (TG), and provided new data that TRPA1 is also expressed in primary afferents innervating masticatory muscles in double-labeling immunohistochemistry experiments. We then examined whether the activation of each TRP channel in the masseter muscle evokes acute nocifensive responses and leads to the development of masseter hypersensitivity to mechanical stimulation using the behavioral models that have been specifically designed and validated for the craniofacial system. Intramuscular injections with specific agonists for TRPV1 and TRPA1, capsaicin and mustard oil (MO), respectively, produced immediate nocifensive hindpaw responses followed by prolonged mechanical hyperalgesia in a concentration-dependent manner. Pretreatment of the muscle with a TRPV1 antagonist, capsazepine, effectively attenuated the capsaicin-induced muscle nociception and mechanical hyperalgesia. Similarly, pretreatment of the muscle with a selective TRPA1 antagonist, AP18, significantly blocked the MO-induced muscle nociception and mechanical hyperalgesia. We confirmed these data with another set of selective antagonist for TRPV1 and TRPA1, AMG9810 and HC030031, respectively. Collectively, these results provide compelling evidence that TRPV1 and TRPA1 can functionally contribute to muscle nociception and hyperalgesia, and suggest that TRP channels expressed in muscle afferents can engage in the development of pathologic muscle pain conditions.

  10. Electroacupuncture suppresses capsaicin-induced secondary hyperalgesia through an endogenous spinal opioid mechanism

    PubMed Central

    Kim, Hee Young; Wang, Jigong; Lee, Inhyung; Kim, Hee Kee; Chung, Kyungsoon; Chung, Jin Mo

    2009-01-01

    Central sensitization, caused either by tissue inflammation or peripheral nerve injury, plays an important role in persistent pain. An animal model of capsaicin-induced pain has well-defined peripheral and central sensitization components, thus is useful for studying the analgesic effect on two separate components. The focus of this study is to examine the analgesic effects of electroacupuncture (EA) on capsaicin-induced secondary hyperalgesia, which represents central sensitization. Capsaicin (0.5%, 10 μl) was injected into the plantar side of the left hind paw, and foot withdrawal thresholds in response to von Frey stimuli (mechanical sensitivity) were determined for both primary and secondary hyperalgesia in rats. EA (2 Hz, 3 mA) was applied to various pairs of acupoints, GB30-GB34, BL40-BL60, GV2-GV6, LI3-LI6 and SI3-TE8, for 30 min under isofluraine anesthesia and then the effect of EA on mechanical sensitivity of paw was determined. EA applied to the ipsilateral SI3-TE8, but none the other acupoints, significantly reduced capsaicin-induced secondary hyperalgesia but not primary hyperalgesia. EA analgesic effect was inhibited by a systemic non-specific opioid receptor (OR) antagonist or an intrathecal μ- or δ-OR antagonist. EA analgesic effect was not affected by an intrathecal κ-OR antagonist or systemic adrenergic receptor antagonist. This study demonstrates that EA produces a stimulation point specific analgesic effect on capsaicin-induced secondary hyperalgesia (central sensitization), mediated by activating endogenous spinal μ and δ opioid receptors. PMID:19646817

  11. Soluble epoxide hydrolase limits mechanical hyperalgesia during inflammation

    PubMed Central

    2011-01-01

    Background Cytochrome-P450 (CYP450) epoxygenases metabolise arachidonic acid (AA) into four different biologically active epoxyeicosatrienoic acid (EET) regioisomers. Three of the EETs (i.e., 8,9-, 11,12- and 14,15-EET) are rapidly hydrolysed by the enzyme soluble epoxide hydrolase (sEH). Here, we investigated the role of sEH in nociceptive processing during peripheral inflammation. Results In dorsal root ganglia (DRG), we found that sEH is expressed in medium and large diameter neurofilament 200-positive neurons. Isolated DRG-neurons from sEH-/- mice showed higher EET and lower DHET levels. Upon AA stimulation, the largest changes in EET levels occurred in culture media, indicating both that cell associated EET concentrations quickly reach saturation and EET-hydrolyzing activity mostly effects extracellular EET signaling. In vivo, DRGs from sEH-deficient mice exhibited elevated 8,9-, 11,12- and 14,15-EET-levels. Interestingly, EET levels did not increase at the site of zymosan-induced inflammation. Cellular imaging experiments revealed direct calcium flux responses to 8,9-EET in a subpopulation of nociceptors. In addition, 8,9-EET sensitized AITC-induced calcium increases in DRG neurons and AITC-induced calcitonin gene related peptide (CGRP) release from sciatic nerve axons, indicating that 8,9-EET sensitizes TRPA1-expressing neurons, which are known to contribute to mechanical hyperalgesia. Supporting this, sEH-/- mice showed increased nociceptive responses to mechanical stimulation during zymosan-induced inflammation and 8,9-EET injection reduced mechanical thresholds in naive mice. Conclusion Our results show that the sEH can regulate mechanical hyperalgesia during inflammation by inactivating 8,9-EET, which sensitizes TRPA1-expressing nociceptors. Therefore we suggest that influencing the CYP450 pathway, which is actually highly considered to treat cardiovascular diseases, may cause pain side effects. PMID:21970373

  12. Epac–protein kinase C alpha signaling in purinergic P2X3R-mediated hyperalgesia after inflammation

    PubMed Central

    Gu, Yanping; Li, Guangwen; Chen, Yong; Huang, Li-Yen Mae

    2016-01-01

    Abstract Sensitization of purinergic P2X3 receptors (P2X3Rs) is a major mechanism contributing to injury-induced exaggerated pain responses. We showed in a previous study that cyclic adenosine monophosphate (cAMP)–dependent guanine nucleotide exchange factor 1 (Epac1) in rat sensory dorsal root ganglia (DRGs) is upregulated after inflammatory injury, and it plays a critical role in P2X3R sensitization by activating protein kinase C epsilon (PKCε) inside the cells. protein kinase C epsilon has been established as the major PKC isoform mediating injury-induced hyperalgesic responses. On the other hand, the role of PKCα in receptor sensitization was seldom considered. Here, we studied the participation of PKCα in Epac signaling in P2X3R-mediated hyperalgesia. The expression of both Epac1 and Epac2 and the level of cAMP in DRGs are greatly enhanced after complete Freund adjuvant (CFA)–induced inflammation. The expression of phosphorylated PKCα is also upregulated. Complete Freund adjuvant (CFA)–induced P2X3R-mediated hyperalgesia is not only blocked by Epac antagonists but also by the classical PKC isoform inhibitors, Go6976, and PKCα-siRNA. These CFA effects are mimicked by the application of the Epac agonist, 8-(4-chlorophenylthio)-2 -O-methyl-cAMP (CPT), in control rats, further confirming the involvement of Epacs. Because the application of Go6976 prior to CPT still reduces CPT-induced hyperalgesia, PKCα is downstream of Epacs to mediate the enhancement of P2X3R responses in DRGs. The pattern of translocation of PKCα inside DRG neurons in response to CPT or CFA stimulation is distinct from that of PKCε. Thus, in contrast to prevalent view, PKCα also plays an essential role in producing complex inflammation-induced receptor-mediated hyperalgesia. PMID:26963850

  13. Enhanced cytokine-induced mechanical hyperalgesia in skeletal muscle produced by a novel mechanism in rats exposed to unpredictable sound stress.

    PubMed

    Dina, Olayinka A; Levine, Jon D; Green, Paul G

    2011-09-01

    Stress exacerbates both experimental and clinical pain, most well-characterized in irritable bowel and fibromyalgia syndromes. Since it has been hypothesized that cytokines play an etiopathogenic role in fibromyalgia and other chronic widespread pain conditions, we investigated the relationship between stress and cytokines in a model of stress-induced chronic somatic pain. A series of experiments were performed to evaluate the impact of stress on the hyperalgesia-induced by endotoxin (lipopolysaccharide, LPS) and the role of two pro-inflammatory cytokines, interleukin-6 (IL-6) and tumor necrosis α (TNFα). Fourteen days after exposure to a 4-day protocol of unpredictable sound stress, the ability of systemic LPS (100 μg/kg, i.p) to elicit cytokine-mediated mechanical hyperalgesia was measured in gastrocnemius muscle. LPS-induced hyperalgesia was significantly greater in stressed rats, but when rats were treated intrathecally with antisense oligodeoxynucleotide (ODN), to decrease either the gp130 subunit of the IL-6 receptor or the TNFα receptor, in nociceptors, skeletal muscle hyperalgesia in sound stressed, but not control, rats was prevented. These data suggest that chronic stress alters signaling in the primary afferent nociceptor for the hyperalgesia induced by endogenously produced pro-inflammatory cytokines.

  14. HCN2 channels account for mechanical (but not heat) hyperalgesia during long-standing inflammation.

    PubMed

    Schnorr, Sabine; Eberhardt, Mirjam; Kistner, Katrin; Rajab, Hamsa; Käßer, Johannes; Hess, Andreas; Reeh, Peter; Ludwig, Andreas; Herrmann, Stefan

    2014-06-01

    There is emerging evidence that hyperpolarization-activated cation (HCN) channels are involved in the development of pathological pain, including allodynia and hyperalgesia. Mice lacking the HCN isoform 2 display reduced heat but unchanged mechanical pain behavior, as recently shown in preclinical models of acute inflammatory pain. However, the impact of HCN2 to chronic pain conditions is less clear and has not been examined so far. In this report, we study the role of HCN2 in the complete Freund's adjuvant inflammation model reflecting chronic pain conditions. We used sensory neuron-specific as well as inducible global HCN2 mutants analyzing pain behavior in persistent inflammation and complemented this by region-specific administration of an HCN channel blocker. Our results demonstrate that the absence of HCN2 in primary sensory neurons reduces tactile hypersensitivity in chronic inflammatory conditions but leaves heat hypersensitivity unaffected. This result is in remarkable contrast to the recently described role of HCN2 in acute inflammatory conditions. We show that chronic inflammation results in an increased expression of HCN2 and causes sensitization in peripheral and spinal terminals of the pain transduction pathway. The contribution of HCN2 to peripheral sensitization mechanisms was further supported by single-fiber recordings from isolated skin-nerve preparations and by conduction velocity measurements of saphenous nerve preparations. Global HCN2 mutants revealed that heat hypersensitivity-unaffected in peripheral HCN2 mutants-was diminished by the additional disruption of central HCN2 channels, suggesting that thermal hyperalgesia under chronic inflammatory conditions is mediated by HCN2 channels beyond primary sensory afferents.

  15. Synthesis of Lipid Mediators during UVB-Induced Inflammatory Hyperalgesia in Rats and Mice

    PubMed Central

    Sisignano, Marco; Angioni, Carlo; Ferreiros, Nerea; Schuh, Claus-Dieter; Suo, Jing; Schreiber, Yannick; Dawes, John M.; Antunes-Martins, Ana; Bennett, David L. H.; McMahon, Stephen B.; Geisslinger, Gerd; Scholich, Klaus

    2013-01-01

    Peripheral sensitization during inflammatory pain is mediated by a variety of endogenous proalgesic mediators including a number of oxidized lipids, some of which serve endogenous modulators of sensory TRP-channels. These lipids are eicosanoids of the arachidonic acid and linoleic acid pathway, as well as lysophophatidic acids (LPAs). However, their regulation pattern during inflammatory pain and their contribution to peripheral sensitization is still unclear. Here, we used the UVB-model for inflammatory pain to investigate alterations of lipid concentrations at the site of inflammation, the dorsal root ganglia (DRGs) as well as the spinal dorsal horn and quantified 21 lipid species from five different lipid families at the peak of inflammation 48 hours post irradiation. We found that known proinflammatory lipids as well as lipids with unknown roles in inflammatory pain to be strongly increased in the skin, whereas surprisingly little changes of lipid levels were seen in DRGs or the dorsal horn. Importantly, although there are profound differences between the number of cytochrome (CYP) genes between mice and rats, CYP-derived lipids were regulated similarly in both species. Since TRPV1 agonists such as LPA 18∶1, 9- and 13-HODE, 5- and 12-HETE were elevated in the skin, they may contribute to thermal hyperalgesia and mechanical allodynia during UVB-induced inflammatory pain. These results may explain why some studies show relatively weak analgesic effects of cyclooxygenase inhibitors in UVB-induced skin inflammation, as they do not inhibit synthesis of other proalgesic lipids such as LPA 18∶1, 9-and 13-HODE and HETEs. PMID:24349046

  16. The dorsomedial hypothalamus mediates stress-induced hyperalgesia and is the source of the pronociceptive peptide cholecystokinin in the rostral ventromedial medulla.

    PubMed

    Wagner, K M; Roeder, Z; Desrochers, K; Buhler, A V; Heinricher, M M; Cleary, D R

    2013-05-15

    While intense or highly arousing stressors have long been known to suppress pain, relatively mild or chronic stress can enhance pain. The mechanisms underlying stress-induced hyperalgesia (SIH) are only now being defined. The physiological and neuroendocrine effects of mild stress are mediated by the dorsomedial hypothalamus (DMH), which has documented connections with the rostral ventromedial medulla (RVM), a brainstem region capable of facilitating nociception. We hypothesized that stress engages both the DMH and the RVM to produce hyperalgesia. Direct pharmacological activation of the DMH increased sensitivity to mechanical stimulation in awake animals, confirming that the DMH can mediate behavioral hyperalgesia. A behavioral model of mild stress also produced mechanical hyperalgesia, which was blocked by inactivation of either the DMH or the RVM. The neuropeptide cholecystokinin (CCK) acts in the RVM to enhance nociception and is abundant in the DMH. Using a retrograde tracer and immunohistochemical labeling, we determined that CCK-expressing neurons in the DMH are the only significant supraspinal source of CCK in the RVM. However, not all neurons projecting from the DMH to the RVM contained CCK, and microinjection of the CCK2 receptor antagonist YM022 in the RVM did not interfere with SIH, suggesting that transmitters in addition to CCK play a significant role in this connection during acute stress. While the RVM has a well-established role in facilitation of nociception, the DMH, with its well-documented role in stress, may also be engaged in a number of chronic or abnormal pain states. Taken as a whole, these findings establish an anatomical and functional connection between the DMH and RVM by which stress can facilitate pain.

  17. Spinal fMRI Reveals Decreased Descending Inhibition during Secondary Mechanical Hyperalgesia

    PubMed Central

    Rempe, Torge; Wolff, Stephan; Riedel, Christian; Baron, Ralf; Stroman, Patrick W.; Jansen, Olav; Gierthmühlen, Janne

    2014-01-01

    Mechanical hyperalgesia is one distressing symptom of neuropathic pain which is explained by central sensitization of the nociceptive system. This sensitization can be induced experimentally with the heat/capsaicin sensitization model. The aim was to investigate and compare spinal and supraspinal activation patterns of identical mechanical stimulation before and after sensitization using functional spinal magnetic resonance imaging (spinal fMRI). Sixteen healthy subjects (6 female, 10 male, mean age 27.2±4.0 years) were investigated with mechanical stimulation of the C6 dermatome of the right forearm during spinal fMRI. Testing was always performed in the area outside of capsaicin application (i.e. area of secondary mechanical hyperalgesia). During slightly noxious mechanical stimulation before sensitization, activity was observed in ipsilateral dorsolateral pontine tegmentum (DLPT) which correlated with activity in ipsilateral spinal cord dorsal gray matter (dGM) suggesting activation of descending nociceptive inhibition. During secondary mechanical hyperalgesia, decreased activity was observed in bilateral DLPT, ipsilateral/midline rostral ventromedial medulla (RVM), and contralateral subnucleus reticularis dorsalis, which correlated with activity in ipsilateral dGM. Comparison of voxel-based activation patterns during mechanical stimulation before/after sensitization showed deactivations in RVM and activations in superficial ipsilateral dGM. This study revealed increased spinal activity and decreased activity in supraspinal centers involved in pain modulation (SRD, RVM, DLPT) during secondary mechanical hyperalgesia suggesting facilitation of nociception via decreased endogenous inhibition. Results should help prioritize approaches for further in vivo studies on pain processing and modulation in humans. PMID:25372292

  18. Dynamic mechanical assessment of muscle hyperalgesia in humans: The dynamic algometer

    PubMed Central

    Finocchietti, Sara; Graven-Nielsen, Thomas; Arendt-Nielsen, Lars

    2015-01-01

    BACKGROUND: Musculoskeletal pain is often associated with a nonhomogeneous distribution of mechanical hyperalgesia. Consequently, new methods able to detect this distribution are needed. OBJECTIVE: To develop and test a new method for assessing muscle hyperalgesia with high temporal and spatial resolution that provides complementary information compared with information obtained by traditional static pressure algometry. METHODS: The dynamic pressure algometer was tested bilaterally on the tibialis anterior muscle in 15 healthy subjects and compared with static pressure algometry. The device consisted of a wheel that was rolled over the muscle tissue with a fixed velocity and different predefined forces. The pain threshold force was determined and pain intensity to a fixed-force stimulation was continuously rated on a visual analogue scale while the wheel was rolling over the muscle. The pressure pain sensitivity was evaluated before, during, and after muscle pain and hyperalgesia induced unilaterally by either injection of hypertonic saline (0.5 mL, 6%) into the tibialis anterior or eccentric exercise evoking delayed-onset muscle soreness (DOMS). RESULTS: The intraclass correlation coefficient was >0.88 for the dynamic thresholds; thus, the method was reliable. Compared with baseline, both techniques detected hyperalgesia at the saline injection site and during DOMS (P<0.05). The dynamic algometer also detected the widespread, patchy distribution of sensitive loci during DOMS, which was difficult to evaluate using static pressure algometry. DISCUSSION AND CONCLUSION: The present study showed that dynamic pressure algometry is a reliable tool for evaluating muscle hyperalgesia (threshold and pain rating) with high temporal and spatial resolution. It can be applied as a simple clinical bed-side test and as a quantitative tool in pharmacological profiling studies. PMID:25664539

  19. Activation of the cAMP transduction cascade contributes to the mechanical hyperalgesia and allodynia induced by intradermal injection of capsaicin.

    PubMed

    Sluka, K A

    1997-11-01

    cord level results in mechanical hyperalgesia and allodynia and that the secondary mechanical hyperalgesia and allodynia following intradermal injection of capsaicin is mediated by this same transduction cascade.

  20. Behavioral evidence of thermal hyperalgesia and mechanical allodynia induced by intradermal cinnamaldehyde in rats

    PubMed Central

    Tsagareli, Merab G.; Tsiklauri, Nana; Zanotto, Karen L.; Carstens, Mirela Iodi; Klein, Amanda H.; Sawyer, Carolyn M.; Gurtskaia, Gulnazi; Abzianidze, Elene; Carstens, E.

    2010-01-01

    TRPA1 agonists cinnamaldehyde (CA) and mustard oil (allyl isothiocyanate= AITC) induce heat hyperalgesia and mechanical allodynia in human skin, and sensitize responses of spinal and trigeminal dorsal horn neurons to noxious skin heating in rats. TRPA1 is also implicated in cold nociception. We presently used behavioral methods to investigate if CA affects sensitivity to thermal and mechanical stimuli in rats. Unilateral intraplantar injection of CA (5-20%) induced a significant, concentration-dependent reduction in latency for ipsilateral paw withdrawal from a noxious heat stimulus, peaking (61.7% of pre-injection baseline) by 30 min with partial recovery at 120 min. The highest dose of CA also significantly reduced the contralateral paw withdrawal latency. CA significantly reduced mechanical withdrawal thresholds of the injected paw that peaked sooner (3 min) and was more profound (44.4% of baseline), with no effect contralaterally. Bilateral intraplantar injections of CA resulted in a significant cold hyperalgesia (cold-plate test) and a weak enhancement of innocuous cold avoidance (thermal preference test). The data are consistent with roles for TRPA1 in thermal (hot and cold) hyperalgesia and mechanical allodynia. PMID:20219630

  1. Melatonin Alters the Mechanical and Thermal Hyperalgesia Induced by Orofacial Pain Model in Rats.

    PubMed

    Scarabelot, Vanessa Leal; Medeiros, Liciane Fernandes; de Oliveira, Carla; Adachi, Lauren Naomi Spezia; de Macedo, Isabel Cristina; Cioato, Stefania Giotti; de Freitas, Joice S; de Souza, Andressa; Quevedo, Alexandre; Caumo, Wolnei; Torres, Iraci Lucena da Silva

    2016-10-01

    Melatonin is a neuroendocrine hormone that presents a wide range of physiological functions including regulating circadian rhythms and sleep, enhancing immune function, sleep improvement, and antioxidant effects. In addition, melatonin has received special attention in pain treatment since it is effective and presents few adverse effects. In this study, we evaluated the effect of acute dose of melatonin upon hyperalgesia induced by complete Freund's adjuvant in a chronic orofacial pain model in Sprague-Dawley rats. Nociceptive behavior was assessed by facial Von Frey and the hot plate tests at baseline and thereafter 30, 60, and 120 min, 24 h, and 7 days after melatonin treatment. We demonstrated that acute melatonin administration alters mechanical and thermal hyperalgesia induced by an orofacial pain model (TMD), highlighting that the melatonin effect upon mechanical hyperalgesia remained until 7 days after its administration. Besides, we observed specific tissue profiles of neuroimmunomodulators linked to pain conditions and/or melatonin effect (brain-derived neurotrophic factor, nerve growth factor, and interleukins 6 and 10) in the brainstem levels, and its effects were state-dependent of the baseline of these animals.

  2. Thiamine, pyridoxine, cyanocobalamin and their combination inhibit thermal, but not mechanical hyperalgesia in rats with primary sensory neuron injury.

    PubMed

    Wang, Zheng-Bei; Gan, Qiang; Rupert, Ronald L; Zeng, Yin-Ming; Song, Xue-Jun

    2005-03-01

    Neuropathic pain after nerve injury is severe and intractable, and current drugs and nondrug therapies offer substantial pain relief to no more than half of affected patients. The present study investigated the analgesic roles of the B vitamins thiamine (B1), pyridoxine (B6) and cyanocobalamin (B12) in rats with neuropathic pain caused by spinal ganglia compression (CCD) or loose ligation of the sciatic nerve (CCI). Thermal hyperalgesia was determined by a significantly shortened latency of foot withdrawal to radiant heat, and mechanical hyperalgesia was determined by a significantly decreased threshold of foot withdrawal to von Frey filaments stimulation of the plantar surface of hindpaw. Results showed that (1) intraperitoneal injection of B1 (5, 10, 33 and 100 mg/kg), B6 (33 and 100 mg/kg) or B12 (0.5 and 2 mg/kg) significantly reduced thermal hyperalgesia; (2) the combination of B1, B6 and B12 synergistically inhibited thermal hyperalgesia; (3) repetitive administration of vitamin B complex (containing B1/B6/B12 33/33/0.5 mg/kg, for 1 and 2 wk) produced long-term inhibition of thermal hyperalgesia; and (4) B vitamins did not affect mechanical hyperalgesia or normal pain sensation, and exhibited similar effects on CCD and CCI induced-hyperalgesia. The present studies demonstrate effects of B vitamins on pain and hyperalgesia following primary sensory neurons injury, and suggest the possible clinical utility of B vitamins in the treatment of neuropathic painful conditions following injury, inflammation, degeneration or other disorders in the nervous systems in human beings.

  3. Anti-Nociceptive Effect of Resveratrol During Inflammatory Hyperalgesia via Differential Regulation of pro-Inflammatory Mediators.

    PubMed

    Singh, Ajeet Kumar; Vinayak, Manjula

    2016-07-01

    Sensitization of nociceptive neurons by inflammatory mediators leads to hypersensitivity for normal painful stimuli which is termed hyperalgesia. Oxidative stress is an essential factor in pathological pain; therefore, antioxidants qualify as potential anti-hyperalgesic agents. The present study examines the efficacy of the natural antioxidant resveratrol in complete Freund's adjuvant (CFA) induced hyperalgesic rats. Thermal hyperalgesia was measured at different time points by paw withdrawal latency test and confirmed by c-Fos expression in spinal dorsal horn. The impact of resveratrol treatment on inflammatory mediators at peripheral (paw skin) and central (spinal cord) sites was determined during early (6 h) as well as late phase (48 h) of hyperalgesia. Intraplanter injection of CFA increased the level of cytokines IL-1β, TNF-α and IL-6 as well as inflammatory enzymes COX-2 and iNOS in paw skin in both phases. In case of spinal cord, the level of COX-2 was found to be elevated in both phases, whereas iNOS could not be detected. The cytokines were found to be elevated only in late phase in spinal cord. Administration of resveratrol (20 mg/kg) shifted the level of all inflammatory mediators towards normal, except cytokines in paw skin. The present study suggests that the anti-nociceptive effect of resveratrol is implicated at both peripheral and central sites in a tissue specific manner. Copyright © 2016 John Wiley & Sons, Ltd.

  4. Phosphatidylinositol 3-kinase activates ERK in primary sensory neurons and mediates inflammatory heat hyperalgesia through TRPV1 sensitization.

    PubMed

    Zhuang, Zhi-Ye; Xu, Haoxing; Clapham, David E; Ji, Ru-Rong

    2004-09-22

    Although the PI3K (phosphatidylinositol 3-kinase) pathway typically regulates cell growth and survival, increasing evidence indicates the involvement of this pathway in neural plasticity. It is unknown whether the PI3K pathway can mediate pain hypersensitivity. Intradermal injection of capsaicin and NGF produce heat hyperalgesia by activating their respective TRPV1 (transient receptor potential vanilloid receptor-1) and TrkA receptors on nociceptor sensory nerve terminals. We examined the activation of PI3K in primary sensory DRG neurons by these inflammatory agents and the contribution of PI3K activation to inflammatory pain. We further investigated the correlation between the PI3K and the ERK (extracellular signal-regulated protein kinase) pathway. Capsaicin and NGF induce phosphorylation of the PI3K downstream target AKT (protein kinase B), which is blocked by the PI3K inhibitors LY294002 and wortmannin, indicative of the activation of PI3K by both agents. ERK activation by capsaicin and NGF was also blocked by PI3K inhibitors. Similarly, intradermal capsaicin in rats activated PI3K and ERK in C-fiber DRG neurons and epidermal nerve fibers. Injection of PI3K or MEK (ERK kinase) inhibitors into the hindpaw attenuated capsaicin- and NGF-evoked heat hyperalgesia but did not change basal heat sensitivity. Furthermore, PI3K, but not ERK, inhibition blocked early induction of hyperalgesia. In acutely dissociated DRG neurons, the capsaicin-induced TRPV1 current was strikingly potentiated by NGF, and this potentiation was completely blocked by PI3K inhibitors and primarily suppressed by MEK inhibitors. Therefore, PI3K induces heat hyperalgesia, possibly by regulating TRPV1 activity, in an ERK-dependent manner. The PI3K pathway also appears to play a role that is distinct from ERK by regulating the early onset of inflammatory pain.

  5. TRPA1 mediates bladder hyperalgesia in a mouse model of cystitis.

    PubMed

    DeBerry, Jennifer J; Schwartz, Erica S; Davis, Brian M

    2014-07-01

    Urinary bladder pain is a primary symptom associated with interstitial cystitis/painful bladder syndrome. We used systemic injections of cyclophosphamide (CYP), an alkylating antineoplastic agent, to induce cystitis and examine the roles of 2 channels previously demonstrated to be required for inflammatory visceral hyperalgesia: transient receptor potential vanilloid-1 (TRPV1) and ankyrin-1 (TRPA1). Injection of CYP (100 mg/kg, i.p.) every other day for 5 days was accompanied by bladder edema and urothelial ulceration, but without significant plasma extravasation or infiltration of neutrophils. Toluidine blue staining showed a significant increase in the number of degranulated bladder mast cells after CYP treatment. Despite this mild pathology, CYP-treated mice exhibited bladder hyperalgesia 1 day after the final injection that persisted 7 days later. Although many previous studies of visceral hyperalgesia have reported changes in dorsal root ganglion neuron TRPV1 expression and/or function, we found no change in bladder afferent TRPV1 expression or sensitivity on the basis of the percentage of bladder afferents responsive to capsaicin, including at submaximal concentrations. In contrast, the percentage of bladder afferents expressing functional TRPA1 protein (i.e., those responsive to mustard oil) increased ∼2.5-fold 1 day after CYP treatment, and remained significantly elevated 7 days later. Moreover, bladder hyperalgesia was reversed by acute treatment with the TRPA1 antagonist HC-030031 (300 mg/kg, i.p.). Our results indicate that CYP-induced bladder hyperalgesia can be induced without robust inflammation or changes in primary afferent TRPV1. However, significant changes were observed in TRPA1 expression, and blockade of TRPA1 alleviated CYP-induced bladder hyperalgesia.

  6. Reactive Oxygen Species Donors Increase the Responsiveness of Dorsal Horn Neurons and Induce Mechanical Hyperalgesia in Rats

    PubMed Central

    Kim, Hee Young; Lee, Inhyung; Chun, Sang Woo; Kim, Hee Kee

    2015-01-01

    Our previous studies suggest that reactive oxygen species (ROS) scavengers have analgesic effect on neuropathic pain through spinal mechanisms in the rat. The studies suggest that superoxide in spinal cord is one of important mediators of persistent pain. To test the hypothesis that increase of superoxide-derived intermediates leads to central sensitization and pain, the effects of an intrathecal injection of chemical ROS donors releasing either OH∙, OCl−, or H2O2 were examined on pain behaviors. Following treatment with t-BOOH (OH∙ donor), dorsal horn neuron responses to mechanical stimuli in normal rats and the changes of neuronal excitability were explored on substantia gelatinosa (SG) neurons using whole-cell patch clamping recordings. Intrathecal administration of t-BOOH or NaOCl (OCl− donor), but not H2O2, significantly decreased mechanical thresholds of hind paws. The responses of wide dynamic range neurons to mechanical stimuli increased after a local application of t-BOOH. The t-BOOH increased the frequency and the amplitude of excitatory postsynaptic potentials, depolarized membrane potential in SG neurons, and increased the frequency of action potentials evoked by depolarizing current pulses. These results suggest that elevated ROS, especially OH∙, in the spinal cord sensitized dorsal horn neurons and produced hyperalgesia in normal rats. PMID:26457204

  7. Mechanical and thermal hyperalgesia and ectopic neuronal discharge after chronic compression of dorsal root ganglia.

    PubMed

    Song, X J; Hu, S J; Greenquist, K W; Zhang, J M; LaMotte, R H

    1999-12-01

    Chronic compression of the dorsal root ganglion (CCD) was produced in adult rats by implanting a stainless steel rod unilaterally into the intervertebral foramen, one rod at L(4) and another at L(5). Two additional groups of rats received either a sham surgery or an acute injury consisting of a transient compression of the ganglion. Withdrawal of the hindpaw was used as evidence of a nocifensive response to mechanical and thermal stimulation of the plantar surface. In addition, extracellular electrophysiological recordings of spontaneous discharges were obtained from dorsal root fibers of formerly compressed ganglia using an in vitro nerve-DRG-dorsal root preparation. The mean threshold force of punctate indentation and the mean threshold temperature of heating required to elicit a 50% incidence of foot withdrawal ipsilateral to the CCD were significantly lower than preoperative values throughout the 35 days of postoperative testing. The number of foot withdrawals ipsilateral to the CCD during a 20-min contact with a temperature-controlled floor was significantly increased over preoperative values throughout postoperative testing when the floor was 4 degrees C (hyperalgesia) and, to a lesser extent, when it was 30 degrees C (spontaneous pain). Stroking the foot with a cotton wisp never elicited a reflex withdrawal before surgery but did so in most rats tested ipsilateral to the CCD during the first 2 postoperative weeks. In contrast, the CCD produced no changes in responses to mechanical or thermal stimuli on the contralateral foot. The sham operation and acute injury produced no change in behavior other than slight, mechanical hyperalgesia for approximately 1 day, ipsilateral to the acute injury. Ectopic spontaneous discharges generated within the chronically compressed ganglion and, occurring in the absence of blood-borne chemicals and without an intact sympathetic nervous system, were recorded from neurons with intact, conducting, myelinated or unmyelinated

  8. Tetrodotoxin suppresses thermal hyperalgesia and mechanical allodynia in a rat full thickness thermal injury pain model.

    PubMed

    Salas, Margaux M; McIntyre, Matthew K; Petz, Lawrence N; Korz, Walter; Wong, Donald; Clifford, John L

    2015-10-21

    Burn injuries have been identified as the primary cause of injury in 5% of U.S. military personnel evacuated from Operations Iraqi Freedom and Enduring Freedom. Severe burn-associated pain is typically treated with opioids such as fentanyl, morphine, and methadone. Side effects of opioids include respiratory depression, cardiac depression, decrease in motor and cognitive function, as well as the development of hyperalgesia, tolerance and dependence. These effects have led us to search for novel analgesics for the treatment of burn-associated pain in wounded combat service members. Tetrodotoxin (TTX) is a selective voltage-gated sodium channel blocker currently in clinical trials as an analgesic. A phase 3 clinical trial for cancer-related pain has been completed and phase 3 clinical trials on chemotherapy-induced neuropathic pain are planned. It has also been shown in mice to inhibit the development of chemotherapy-induced neuropathic pain. TTX was originally identified as a neurotoxin in marine animals but has now been shown to be safe in humans at therapeutic doses. The antinociceptive effects of TTX are thought to be due to inhibition of Na(+) ion influx required for initiation and conduction of nociceptive impulses. One TTX sensitive sodium channel, Nav1.7, has been shown to be essential in lowering the heat pain threshold after burn injuries. To date, the analgesic effect of TTX has not been tested in burn-associated pain. Male Sprague-Dawley rats were subjected to a full thickness thermal injury on the right hind paw. TTX (8 μg/kg) was administered once a day systemically by subcutaneous injection beginning 3 days post thermal injury and continued through 7 days post thermal injury. Thermal hyperalgesia and mechanical allodynia were assessed 60 and 120 min post injection on each day of TTX treatment. TTX significantly reduced thermal hyperalgesia at all days tested and had a less robust, but statistically significant suppressive effect on mechanical

  9. CX3CR1 Mediates Nicotine Withdrawal-Induced Hyperalgesia via Microglial P38 MAPK Signaling.

    PubMed

    Ding, Yonghong; Shi, Wenhui; Xie, Guannan; Yu, Ailan; Wang, Qinghe; Zhang, Zongwang

    2015-11-01

    Previously, we reported that nicotine withdrawal (NT) significantly increased pain sensitivity in rats. Recent reports suggest that fractalkine is involved in the spinal cord neuron-to-microglia activation via CX3CR1 signaling. However, its contribution to NT-induced hyperalgesia and the underlying mechanisms have yet to be elucidated. In the present study, a rat model of NT was used to test the changes in CX3CR1 expression in the spinal cord. We also evaluated the effect of the CX3CR1 neutralizing antibody on spinal microglial activity, the expression of phosphorylated p38-mitogen-activated protein kinase (p-p38-MAPK) and heat-induced pain responses. We established a NT model via subcutaneous injection of pure nicotine (3 mg/kg), three times daily for 7 days. The expression of CX3CR1 was studied by Western blot and immunofluorescence staining. Following NT, the rats received daily intrathecal injections of CX3CR1 neutralizing antibody for 3 days. The change in paw withdrawal latency (PWL) was observed. The activation of microglia and the expression of p-p38-MAPK were investigated by Western blot and immunofluorescence staining. The expression of CX3CR1 was significantly increased after NT and co-localized with IBA-1. NT rats treated with CX3CR1 neutralizing antibody showed significantly increased PWL on day 4 after NT. Furthermore, the activation of microglia and the expression of p-p38-MAPK in the spinal cord were suppressed. These results indicate that microglial CX3CR1/p38MAPK pathway is critical for the development of pain hypersensitivity after NT.

  10. The effect of intrathecal gabapentin on mechanical and thermal hyperalgesia in neuropathic rats induced by spinal nerve ligation.

    PubMed Central

    Cho, Hyun Sung; Kim, Myung Hee; Choi, Duck Hwan; Lee, Jung Il; Gwak, Mi Sook; Hahm, Tae Soo

    2002-01-01

    Gabapentin decreases the level of glutamate and elevates that of alpha-amino-butyric acid in the central nervous system. Gabapentin was shown to have antinociceptive effects in several facilitated pain models. Intrathecal gabapentin was also known to be effective in reducing mechanical allodynia in animals with neuropathic pain. In this study, we investigated to see whether intrathecal gabapentin produces antihyperalgesic effects on thermal and mechanical hyperalgesia in neuropathic rats and whether its effects are associated with motor impairment. To induce neuropathic pain in Sprague-Dawley rats, left L5 and L6 spinal nerves were ligated. After a week, lumbar catheterization into subarachnoid space was performed. Then, paw withdrawal times to thermal stimuli and vocalization thresholds to paw pressure were determined before and up to 2 hr after intrathecal injection of gabapentin. Also, motor functions including performance times on rota-rod were determined. Intrathecal gabapentin attenuated significantly thermal and mechanical hyperalgesia in neuropathic rats, but did not block thermal and mechanical nociception in sham-operated rats. Intrathecal gabapentin of antihyperalgesic doses inhibited motor coordination performance without evident ambulatory dysfunction. This study demonstrates that intrathecal gabapentin is effective against thermal and mechanical hyperalgesia, in spite of moderate impairment of motor coordination. PMID:11961308

  11. Kinin Receptors Sensitize TRPV4 Channel and Induce Mechanical Hyperalgesia: Relevance to Paclitaxel-Induced Peripheral Neuropathy in Mice.

    PubMed

    Costa, Robson; Bicca, Maíra A; Manjavachi, Marianne N; Segat, Gabriela C; Dias, Fabiana Chaves; Fernandes, Elizabeth S; Calixto, João B

    2017-03-10

    Kinin B1 (B1R) and B2 receptors (B2R) and the transient receptor potential vanilloid 4 (TRPV4) channel are known to play a critical role in the peripheral neuropathy induced by paclitaxel (PTX) in rodents. However, the downstream pathways activated by kinin receptors as well as the sensitizers of the TRPV4 channel involved in this process remain unknown. Herein, we investigated whether kinins sensitize TRPV4 channels in order to maintain PTX-induced peripheral neuropathy in mice. The mechanical hyperalgesia induced by bradykinin (BK, a B2R agonist) or des-Arg(9)-BK (DABK, a B1R agonist) was inhibited by the selective TRPV4 antagonist HC-067047. Additionally, BK was able to sensitize TRPV4, thus contributing to mechanical hyperalgesia. This response was dependent on phospholipase C/protein kinase C (PKC) activation. The selective kinin B1R (des-Arg(9)-[Leu(8)]-bradykinin) and B2R (HOE 140) antagonists reduced the mechanical hyperalgesia induced by PTX, with efficacies and time response profiles similar to those observed for the TRPV4 antagonist (HC-067047). Additionally, both kinin receptor antagonists inhibited the overt nociception induced by hypotonic solution in PTX-injected animals. The same animals presented lower PKCε levels in skin and dorsal root ganglion samples. The selective PKCε inhibitor (εV1-2) reduced the hypotonicity-induced overt nociception in PTX-treated mice with the same magnitude observed for the kinin receptor antagonists. These findings suggest that B1R or B2R agonists sensitize TRPV4 channels to induce mechanical hyperalgesia in mice. This mechanism of interaction may contribute to PTX-induced peripheral neuropathy through the activation of PKCε. We suggest these targets represent new opportunities for the development of effective analgesics to treat chronic pain.

  12. Involvement of spinal glutamate transporter-1 in the development of mechanical allodynia and hyperalgesia associated with type 2 diabetes

    PubMed Central

    Shi, Jinshan; Jiang, Ke; Li, Zhaoduan

    2016-01-01

    Little is known about the effects of the development of type 2 diabetes on glutamate homeostasis in the spinal cord. Therefore, we quantified the extracellular levels of glutamate in the spinal cord of Zucker diabetic fatty (ZDF) rats using in vivo microdialysis. In addition, protein levels of glutamate transporter-1 (GLT-1) in the spinal cord of ZDF rats were measured using Western blot. Finally, the effects of repeated intrathecal injections of ceftriaxone, which was previously shown to enhance GLT-1 expression, on the development of mechanical allodynia and hyperalgesia as well as on basal extracellular level of glutamate and the expression of GLT-1 in the spinal cord of ZDF rats were evaluated. It was found that ZDF rats developed mechanical hyperalgesia and allodynia, which were associated with increased basal extracellular levels of glutamate and attenuated levels of GLT-1 expression in the spinal cord, particularly in the dorsal horn. Furthermore, repeated intrathecal administrations of ceftriaxone dose-dependently prevented the development of mechanical hyperalgesia and allodynia in ZDF rats, which were correlated with enhanced GLT-1 expression without altering the basal glutamate levels in the spinal cord of ZDF rats. Overall, the results suggested that impaired glutamate reuptake in the spinal cord may contribute to the development of neuropathic pains in type 2 diabetes. PMID:27932896

  13. Assessment of Morphine-induced Hyperalgesia and Analgesic Tolerance in Mice Using Thermal and Mechanical Nociceptive Modalities

    PubMed Central

    Elhabazi, Khadija; Ayachi, Safia; Ilien, Brigitte; Simonin, Frédéric

    2014-01-01

    Opioid-induced hyperalgesia and tolerance severely impact the clinical efficacy of opiates as pain relievers in animals and humans. The molecular mechanisms underlying both phenomena are not well understood and their elucidation should benefit from the study of animal models and from the design of appropriate experimental protocols. We describe here a methodological approach for inducing, recording and quantifying morphine-induced hyperalgesia as well as for evidencing analgesic tolerance, using the tail-immersion and tail pressure tests in wild-type mice. As shown in the video, the protocol is divided into five sequential steps. Handling and habituation phases allow a safe determination of the basal nociceptive response of the animals. Chronic morphine administration induces significant hyperalgesia as shown by an increase in both thermal and mechanical sensitivity, whereas the comparison of analgesia time-courses after acute or repeated morphine treatment clearly indicates the development of tolerance manifested by a decline in analgesic response amplitude. This protocol may be similarly adapted to genetically modified mice in order to evaluate the role of individual genes in the modulation of nociception and morphine analgesia. It also provides a model system to investigate the effectiveness of potential therapeutic agents to improve opiate analgesic efficacy. PMID:25145878

  14. Nerve growth factor and associated nerve sprouting contribute to local mechanical hyperalgesia in a rat model of bone injury.

    PubMed

    Yasui, M; Shiraishi, Y; Ozaki, N; Hayashi, K; Hori, K; Ichiyanagi, M; Sugiura, Y

    2012-08-01

    To clarify the mechanism of tenderness after bone injury, we investigated changes in the withdrawal threshold to mechanical stimuli, nerve distribution and nerve growth factor (NGF)-expression in a rat model of bone injury without immobilization for bone injury healing. Rats were divided into three groups as follows: (1) rats incised in the skin and periosteum, followed by drilling a hole in the tibia [bone lesion group (BLG)]; (2) those incised in the skin and periosteum without bone drilling [periosteum lesion group (PLG)]; and (3) those incised in the skin [skin lesion group (SLG)]. Mechanical hyperalgesia continued for 28 days at a lesion in the BLG, 21 days in PLG and 5 days in SLG after treatments, respectively. Endochondral ossification was observed on days 5-28 in BLG and on days 5-21 in PLG. Nerve growth appeared in deep connective tissue (DCT) at day 28 in BLG. Nerve fibres increased in both cutaneous tissue and DCT at day 7 in PLG, but they were not found at day 28. Mechanical hyperalgesia accompanied with endochondral ossification and nerve fibres increasing at the lesion in both BLG and PLG. NGF was expressed in bone-regenerating cells during the bone injury healing. Anti-NGF and trk inhibitor K252a inhibited hyperalgesia in the different time course. This study shows that localized tenderness coincides with the bone healing and involves NGF expression and nerve sprouting after bone injury. The findings present underlying mechanisms and provide pathophysiological relevance of local tenderness to determination of bone fracture and its healing.

  15. The TrkA receptor mediates experimental thermal hyperalgesia produced by nerve growth factor: Modulation by the p75 neurotrophin receptor.

    PubMed

    Khodorova, Alla; Nicol, Grant D; Strichartz, Gary

    2017-01-06

    The p75 neurotrophin receptor (p75(NTR)) and its activation of the sphingomyelin signaling cascade are essential for mechanical hypersensitivity resulting from locally injected nerve growth factor (NGF). Here the roles of the same effectors, and of the tropomyosin receptor kinase A (TrkA) receptor, are evaluated for thermal hyperalgesia from NGF. Sensitivity of rat hind paw plantar skin to thermal stimulation after local sub-cutaneous injection of NGF (500ng) was measured by the latency for paw withdrawal (PWL) from a radiant heat source. PWL was reduced from baseline values at 0.5-22h by ∼40% from that in naïve or vehicle-injected rats, and recovered to pre-injection levels by 48h. Local pre-injection with a p75(NTR) blocking antibody did not affect the acute thermal hyperalgesia (0.5-3.5h) but hastened its recovery so that it had reversed to baseline by 22h. In addition, GW4869 (2mM), an inhibitor of the neutral sphingomyelinase (nSMase) that is an enzyme in the p75(NTR) pathway, also failed to prevent thermal hyperalgesia. However, C2-ceramide, an analog of the ceramide produced by sphingomyelinase, did cause thermal hyperalgesia. Injection of an anti-TrkA antibody known to promote dimerization and activation of that receptor, independent of NGF, also caused thermal hyperalgesia, and prevented the further reduction of PWL from subsequently injected NGF. A non-specific inhibitor of tropomyosin receptor kinases, K252a, prevented thermal hyperalgesia from NGF, but not that from the anti-TrkA antibody. These findings suggest that the TrkA receptor has a predominant role in thermal hypersensitivity induced by NGF, while p75(NTR) and its pathway intermediates serve a modulatory role.

  16. The relationship of bone-tumor-induced spinal cord astrocyte activation and aromatase expression to mechanical hyperalgesia and cold hypersensitivity in intact female and ovariectomized mice.

    PubMed

    Smeester, B A; O'Brien, E E; Michlitsch, K S; Lee, J-H; Beitz, A J

    2016-06-02

    Recently, our group established a relationship between tumor-induced spinal cord astrocyte activation and aromatase expression and the development of bone tumor nociception in male mice. As an extension of this work, we now report on the association of tumor-induced mechanical hyperalgesia and cold hypersensitivity to changes in spinal cord dorsal horn GFAP and aromatase expression in intact (INT) female mice and the effect of ovariectomy on these parameters. Implantation of fibrosarcoma cells produced robust mechanical hyperalgesia in INT animals, while ovariectomized (OVX) females had significantly less mechanical hyperalgesia. Cold hypersensitivity was apparent by post-implantation day 7 in INT and OVX females compared to their saline-injected controls and increased throughout the experiment. The decrease in mechanical hyperalgesia in OVX females was mirrored by significant decreases in spinal astrocyte activity in laminae I-II, III-IV, V-VI and X and aromatase expression in laminae V-VI and X in the dorsal horn of tumor-bearing animals. Administration of the aromatase inhibitor letrozole reduced tumor-induced hyperalgesia in INT females only suggesting that the tumor-induced increase in aromatase expression and its associated increase in spinal estrogen play a role in the development of bone tumor-induced hyperalgesia. Finally, intrathecal (i.t.) administration of 17β-estradiol caused a significant increase in tumor-induced hyperalgesia in INT tumor-bearing females. Since i.t. 17β-estradiol increases tumor pain and ovariectomy significantly decreases tumor pain, as well as spinal aromatase, estrogen may play a critical role in the spinal cord response to the changing tumor environment and the development of tumor-induced nociception.

  17. Heat hyperalgesia and mechanical hypersensitivity induced by calcitonin gene-related peptide in a mouse model of neurofibromatosis.

    PubMed

    White, Stephanie; Marquez de Prado, Blanca; Russo, Andrew F; Hammond, Donna L

    2014-01-01

    This study examined whether mice with a deficiency of neurofibromin, a Ras GTPase activating protein, exhibit a nociceptive phenotype and probed a possible contribution by calcitonin gene-related peptide. In the absence of inflammation, Nf1+/- mice (B6.129S6 Nf1/J) and wild type littermates responded comparably to heat or mechanical stimuli, except for a subtle enhanced mechanical sensitivity in female Nf1+/- mice. Nociceptive phenotype was also examined after inflammation induced by capsaicin and formalin, which release endogenous calcitonin gene-related peptide. Intraplantar injection of capsaicin evoked comparable heat hyperalgesia and mechanical hypersensitivity in Nf1+/- and wild type mice of both genders. Formalin injection caused a similar duration of licking in male Nf1+/- and wild type mice. Female Nf1+/- mice licked less than wild type mice, but displayed other nociceptive behaviors. In contrast, intraplantar injection of CGRP caused greater heat hyperalgesia in Nf1+/- mice of both genders compared to wild type mice. Male Nf1+/- mice also exhibited greater mechanical hypersensitivity; however, female Nf1+/- mice exhibited less mechanical hypersensitivity than their wild type littermates. Transcripts for calcitonin gene-related peptide were similar in the dorsal root ganglia of both genotypes and genders. Transcripts for receptor activity-modifying protein-1, which is rate-limiting for the calcitonin gene-related peptide receptor, in the spinal cord were comparable for both genotypes and genders. The increased responsiveness to intraplantar calcitonin gene-related peptide suggests that the peripheral actions of calcitonin gene-related peptide are enhanced as a result of the neurofibromin deficit. The analgesic efficacy of calcitonin gene-related peptide receptor antagonists may therefore merit investigation in neurofibromatosis patients.

  18. Inflammation enhances Y1 receptor signaling, neuropeptide Y-mediated inhibition of hyperalgesia, and substance P release from primary afferent neurons

    PubMed Central

    Taylor, Bradley K.; Fu, Weisi; Kuphal, Karen E.; Stiller, Carl-Olav; Winter, Michelle K.; Chen, Wenling; Corder, Gregory F.; Urban, Janice H.; McCarson, Kenneth E.; Marvizon, Juan Carlos

    2014-01-01

    Neuropeptide Y (NPY) is present in the superficial laminae of the dorsal horn and inhibits spinal nociceptive processing, but the mechanisms underlying its anti-hyperalgesic actions are unclear. We hypothesized that NPY acts at neuropeptide Y1 receptors in dorsal horn to decrease nociception by inhibiting substance P (SP) release, and that these effects are enhanced by inflammation. To evaluate SP release, we used microdialysis and neurokinin 1 receptor (NK1R) internalization in rat. NPY decreased capsaicin-evoked SP-like immunoreactivity in microdialysate of the dorsal horn. NPY also decreased non-noxious stimulus (paw brush)-evoked NK1R internalization (as well as mechanical hyperalgesia and mechanical and cold allodynia) after intraplantar injection of carrageenan. Similarly, in rat spinal cord slices with dorsal root attached, [Leu31, Pro34]-NPY inhibited dorsal root stimulus-evoked NK1R internalization. In rat dorsal root ganglion neurons, Y1 receptors colocalized extensively with calcitonin gene-related peptide (CGRP). In dorsal horn neurons, Y1 receptors were extensively expressed and this may have masked detection of terminal co-localization with CGRP or SP. To determine whether the pain inhibitory actions of Y1 receptors are enhanced by inflammation, we administered [Leu31, Pro34]-NPY after intraplantar injection of complete Freund's adjuvant (CFA) in rat. We found that [Leu31, Pro34]-NPY reduced paw clamp-induced NK1R internalization in CFA rats but not uninjured controls. To determine the contribution of increased Y1 receptor-G protein coupling, we measured [35S]GTPγS binding simulated by [Leu31, Pro34]-NPY in mouse dorsal horn. CFA inflammation increased the affinity of Y1 receptor G-protein coupling. We conclude that Y1 receptors contribute to the anti-hyperalgesic effects of NPY by mediating inhibition of SP release, and that Y1 receptor signaling in the dorsal horn is enhanced during inflammatory nociception. PMID:24184981

  19. Phosphorylation of CREB and mechanical hyperalgesia is reversed by blockade of the cAMP pathway in a time-dependent manner after repeated intramuscular acid injections.

    PubMed

    Hoeger-Bement, Marie K; Sluka, Kathleen A

    2003-07-02

    Spinal activation of the cAMP pathway produces mechanical hyperalgesia, sensitizes nociceptive spinal neurons, and phosphorylates the transcription factor cAMP-responsive element binding protein (CREB), which initiates gene transcription. This study examined the role of the cAMP pathway in a model of chronic muscle pain by assessing associated behavioral changes and phosphorylation of CREB. Bilateral mechanical hyperalgesia of the paw was induced by administering two injections of acidic saline, 5 d apart, into the gastrocnemius muscle of male Sprague Dawley rats. Interestingly, the increases in immunoreactivity for CREB and phosphorylated CREB (p-CREB) in the spinal dorsal horn occur 24 hr, but not 1 week, after the second injection of acidic saline compared with pH 7.2 intramuscular injections. Spinal blockade of adenylate cyclase prevents the expected increase in p-CREB that occurs after intramuscular acid injection. The reversal of mechanical hyperalgesia by adenylate cyclase or protein kinase A inhibitors spinally follows a similar pattern with reversal at 24 hr, but not 1 week, compared with the vehicle controls. The p-CREB immunoreactivity in the superficial dorsal horn correlates with the mechanical withdrawal threshold such that increases in p-CREB are associated with decreases in threshold. Therefore, activation of the cAMP pathway in the spinal cord phosphorylates CREB and produces mechanical hyperalgesia associated with intramuscular acid injections. The mechanical hyperalgesia and phosphorylation of CREB depend on early activation of the cAMP pathway during the first 24 hr but are independent of the cAMP pathway by 1 week after intramuscular injection of acid.

  20. The critical role of leukotriene B4 in antigen-induced mechanical hyperalgesia in immunised rats

    PubMed Central

    Cunha, Joice Maria; Sachs, Daniela; Canetti, Claudio Azevedo; Poole, Stephen; Ferreira, Sérgio Henrique; Cunha, Fernando Queiroz

    2003-01-01

    We investigated the mediators responsible for mechanical hypersensitivity induced by antigen challenge in rats immunised with ovalbumin (OVA). Challenge with OVA (12.5–100 μg, intraplantar) caused a dose- and time-dependent mechanical hypersensitivity, which peaked 3 h after, decreased thereafter and reached control levels 24 h later. Levels of TNFα, IL-1β and cytokine-induced neutrophil chemoattractant 1 (CINC-1) were increased in paw skin after antigen challenge. OVA-evoked hypersensitivity was partially inhibited (about 51%) by pretreatment with anti-TNFα , IL-1β and IL-8 sera or with IL-1 receptor antagonist (IL-1ra), but not anti-NGF serum. Pretreatment with thalidomide (45 mg kg−1) or pentoxifylline (100 mg kg−1) also partially inhibited the hypersensitivity at 1–3 h after challenge. Pretreatment with indomethacin (5 mg kg−1) or atenolol (1 mg kg−1) reduced the OVA-induced hypersensitivity at 1 and 3 h, but not at 5 h after challenge, while the combination of B1 and B2 bradykinin receptor antagonists was ineffective over the same times. Pretreatment with MK886 (5-lipoxygenase-activating protein inhibitor, 3 mg kg−1), CP 105696 (LTB4 receptor antagonist; 3 mg kg−1) or dexamethasone (0.5 mg kg−1) inhibited the hypersensitivity from 1 to 5 h. Furthermore, LTB4 levels were increased in the paw skin of challenged rats. In conclusion, our results suggest that the TNFα-, IL-1β- and CINC-1-driven release of prostaglandins, sympathetic amines and LTB4 mediates the first 3 h of mechanical hypersensitivity induced by antigen challenge in rats. At 5 h after OVA administration, although TNFα has some role, LTB4 is the critical nociceptive mediator. PMID:12871832

  1. Analgesic Effect of Photobiomodulation on Bothrops Moojeni Venom-Induced Hyperalgesia: A Mechanism Dependent on Neuronal Inhibition, Cytokines and Kinin Receptors Modulation

    PubMed Central

    Oliveira, Victoria Regina da Silva; Toniolo, Elaine Flamia; Feliciano, Regiane dos Santos; da Silva Jr., José Antonio; Zamuner, Stella Regina

    2016-01-01

    Background Envenoming induced by Bothrops snakebites is characterized by drastic local tissue damage that involves an intense inflammatory reaction and local hyperalgesia which are not neutralized by conventional antivenom treatment. Herein, the effectiveness of photobiomodulation to reduce inflammatory hyperalgesia induced by Bothrops moojeni venom (Bmv), as well as the mechanisms involved was investigated. Methodology/Principal Findings Bmv (1 μg) was injected through the intraplantar route in the right hind paw of mice. Mechanical hyperalgesia and allodynia were evaluated by von Frey filaments at different time points after venom injection. Low level laser therapy (LLLT) was applied at the site of Bmv injection at wavelength of red 685 nm with energy density of 2.2 J/cm2 at 30 min and 3 h after venom inoculation. Neuronal activation in the dorsal horn spinal cord was determined by immunohistochemistry of Fos protein and the mRNA expression of IL-6, TNF-α, IL-10, B1 and B2 kinin receptors were evaluated by Real time-PCR 6 h after venom injection. Photobiomodulation reversed Bmv-induced mechanical hyperalgesia and allodynia and decreased Fos expression, induced by Bmv as well as the mRNA levels of IL-6, TNF-α and B1 and B2 kinin receptors. Finally, an increase on IL-10, was observed following LLLT. Conclusion/Significance These data demonstrate that LLLT interferes with mechanisms involved in nociception and hyperalgesia and modulates Bmv-induced nociceptive signal. The use of photobiomodulation in reducing local pain induced by Bothropic venoms should be considered as a novel therapeutic tool for the treatment of local symptoms induced after bothropic snakebites. PMID:27749899

  2. Effects of non-steroidal anti-inflammatory drugs on the hyperalgesia to noxious mechanical stimulation induced by the application of a tourniquet to a forelimb of sheep.

    PubMed

    Welsh, E M; Nolan, A M

    1994-11-01

    A tourniquet was used in conjunction with a mechanical threshold testing device to investigate the suitability of the technique for the investigation of analgesic drugs in sheep. The changes to the mechanical thresholds to noxious stimulation during and after the inflation of a pneumatic tourniquet on a limb were recorded, and the influence of pre-treatment with two non-steroidal anti-inflammatory drugs was studied. Fentanyl, an opioid agonist with known analgesic properties in sheep, was used as a positive control. The tourniquet significantly reduced the mechanical thresholds on the ipsi- but not the contralateral limb. Pretreatment with either flunixin meglumine or carprofen attenuated the development of mechanical hyperalgesia, and fentanyl initially caused a significant anti-nociceptive effect. The time to aversion was not significantly different between the treatments. These results suggest that hyperalgesia induced by a tourniquet may be a useful technique for the investigation of the anti-nociceptive effects of analgesic drugs in sheep.

  3. Pulsed radiofrequency reduced complete Freund's adjuvant-induced mechanical hyperalgesia via the spinal c-Jun N-terminal kinase pathway.

    PubMed

    Chen, Kuan-Hung; Yang, Chien-Hui; Juang, Sin-Ei; Huang, Hui-Wen; Cheng, Jen-Kun; Sheen-Chen, Shyr-Ming; Cheng, Jiin-Tsuey; Lin, Chung-Ren

    2014-03-01

    Pulsed radiofrequency (PRF) treatment involves the pulsed application of a radiofrequency electric field to a nerve. The technology offers pain relief for patients suffering from chronic pain who do not respond well to conventional treatments. We tested whether PRF treatment attenuated complete Freund's adjuvant (CFA) induced inflammatory pain. The profile of spinal c-Jun N-terminal kinases (JNKs) phosphorylation was evaluated to elucidate the potential mechanism. Injection of CFA into the unilateral hind paw of rats induced mechanical hyperalgesia in both the ipsilateral and contralateral hind paws. We administered 500-kHz PRF treatment in 20-ms pulses, at a rate of 2 Hz (2 pulses per second) either to the sciatic nerve in the mid-thigh, or to the L4 anterior primary ramus just distal to the intervertebral foramen in both the CFA group and no-PRF group rats. Tissue samples were examined at 1, 3, 7, and 14 days following PRF treatments. Behavioral studies showed that PRF applied close to the dorsal root ganglion (DRG) significantly attenuated CFA-induced mechanical hyperalgesia compared to no-PRF group (P < .05). And western blotting revealed significant attenuation of the activation of JNK in the spinal dorsal horn compared to no-PRF group animals (P < .05). Application of PRF close to DRG provides an effective treatment for CFA-induced persistent mechanical hyperalgesia by attenuating JNK activation in the spinal dorsal horn.

  4. Activation of transient receptor potential vanilloid 2-expressing primary afferents stimulates synaptic transmission in the deep dorsal horn of the rat spinal cord and elicits mechanical hyperalgesia.

    PubMed

    Petitjean, Hugues; Hugel, Sylvain; Barthas, Florent; Bohren, Yohann; Barrot, Michel; Yalcin, Ipek; Schlichter, Rémy

    2014-10-01

    Probenecid, an agonist of transient receptor vanilloid (TRPV) type 2, was used to evaluate the effects of TRPV2 activation on excitatory and inhibitory synaptic transmission in the dorsal horn (DH) of the rat spinal cord and on nociceptive reflexes induced by thermal heat and mechanical stimuli. The effects of probenecid were compared with those of capsaicin, a TRPV1 agonist. Calcium imaging experiments on rat dorsal root ganglion (DRG) and DH cultures indicated that functional TRPV2 and TRPV1 were expressed by essentially non-overlapping subpopulations of DRG neurons, but were absent from DH neurons and DH and DRG glial cells. Pretreatment of DRG cultures with small interfering RNAs against TRPV2 suppressed the responses to probenecid. Patch-clamp recordings from spinal cord slices showed that probenecid and capsaicin increased the frequencies of spontaneous excitatory postsynaptic currents (sEPSCs) and spontaneous inhibitory postsynaptic currents in a subset of laminae III-V neurons. In contrast to capsaicin, probenecid failed to stimulate synaptic transmission in lamina II. Intrathecal or intraplantar injections of probenecid induced mechanical hyperalgesia/allodynia without affecting nociceptive heat responses. Capsaicin induced both mechanical hyperalgesia/allodynia and heat hyperalgesia. Activation of TRPV1 or TRPV2 in distinct sets of primary afferents increased the sEPSC frequencies in a largely common population of DH neurons in laminae III-V, and might underlie the development of mechanical hypersensitivity following probenecid or capsaicin treatment. However, only TRPV1-expressing afferents facilitated excitatory and/or inhibitory transmission in a subpopulation of lamina II neurons, and this phenomenon might be correlated with the induction of thermal heat hyperalgesia.

  5. Effects of Distal Nerve Injuries on Dorsal-Horn Neurons and Glia: Relationships Between Lesion Size and Mechanical Hyperalgesia

    PubMed Central

    Lee, J. W.; Siegel, S. M.; Oaklander, A. L.

    2008-01-01

    Penetrating limb injuries are common and usually heal without long-lasting effects, even when nerves are cut. However, rare nerve-injury patients develop prolonged and disabling chronic pain (neuralgia). When pain severity is disproportionate to severity of the inciting injury, physicians and insurers may suspect exaggeration and limit care or benefits, although the nature of the relationship between lesion-size and the development and persistence of neuralgia remains largely unknown. We compared cellular changes in the spinal dorsal-horn (the initial CNS pain-processing area) after partial or total tibial-nerve axotomies in male Sprague–Dawley rats to determine if these changes are proportional to the numbers of peripheral axons cut. Unoperated rats provided controls. Plantar hind-paw responses to touch, pin, and cold were quantitated bilaterally to identify hyperalgesic rats. We also compared data from nerve-injured rats with or without hyperalgesic responses to mechanical hind-paw stimulation to evaluate concordance between pain behaviors and dorsal-horn cellular changes. Hyperalgesia was no less prevalent or severe after partial than after total axotomy. L5 spinal-cord sections from rats killed 7 days postoperatively were labeled for markers of primary afferents (substance P calcitonin gene-related peptide isolectin B4, gamma aminobutyric acid, and glial fibrillary acidic protein), then labeled cells were stereologically quantitated in somatotopically defined dorsal-horn regions. Total axotomy reduced markers of primary afferents more than partial axotomy. In contrast, GABA-immunoreactive profiles were similarly reduced after both lesions, and in rats with sensory loss versus hyperalgesia. Numbers of GFAP-immunoreactive astrocytes increased independently of lesion size and pain status. Small nerve injuries can thus have magnified and disproportionate effects on dorsal-horn neurons and glia, perhaps providing a biological correlate for the disproportionate

  6. Galanin-Mediated Behavioural Hyperalgesia from the Dorsomedial Nucleus of the Hypothalamus Involves Two Independent Descending Pronociceptive Pathways.

    PubMed

    Amorim, Diana; Viisanen, Hanna; Wei, Hong; Almeida, Armando; Pertovaara, Antti; Pinto-Ribeiro, Filipa

    2015-01-01

    Activation of the dorsomedial nucleus of the hypothalamus (DMH) by galanin (GAL) induces behavioural hyperalgesia. Since DMH neurones do not project directly to the spinal cord, we hypothesized that the medullary dorsal reticular nucleus (DRt), a pronociceptive region projecting to the spinal dorsal horn (SDH) and/or the serotoninergic raphe-spinal pathway acting on the spinal 5-HT3 receptor (5HT3R) could relay descending nociceptive facilitation induced by GAL in the DMH. Heat-evoked paw-withdrawal latency (PWL) and activity of SDH neurones were assessed in monoarthritic (ARTH) and control (SHAM) animals after pharmacological manipulations of the DMH, DRt and spinal cord. The results showed that GAL in the DMH and glutamate in the DRt lead to behavioural hyperalgesia in both SHAM and ARTH animals, which is accompanied particularly by an increase in heat-evoked responses of wide-dynamic range neurons, a group of nociceptive SDH neurones. Facilitation of pain behaviour induced by GAL in the DMH was reversed by lidocaine in the DRt and by ondansetron, a 5HT3R antagonist, in the spinal cord. However, the hyperalgesia induced by glutamate in the DRt was not blocked by spinal ondansetron. In addition, in ARTH but not SHAM animals PWL was increased after lidocaine in the DRt and ondansetron in the spinal cord. Our data demonstrate that GAL in the DMH activates two independent descending facilitatory pathways: (i) one relays in the DRt and (ii) the other one involves 5-HT neurones acting on spinal 5HT3Rs. In experimental ARTH, the tonic pain-facilitatory action is increased in both of these descending pathways.

  7. Motor cortex stimulation reduces hyperalgesia in an animal model of central pain.

    PubMed

    Lucas, Jessica M; Ji, Yadong; Masri, Radi

    2011-06-01

    Electrical stimulation of the primary motor cortex has been used since 1991 to treat chronic neuropathic pain. Since its inception, motor cortex stimulation (MCS) treatment has had varied clinical outcomes. Until this point, there has not been a systematic study of the stimulation parameters that most effectively treat chronic pain, or of the mechanisms by which MCS relieves pain. Here, using a rodent model of central pain, we perform a systematic study of stimulation parameters used for MCS and investigate the mechanisms by which MCS reduces hyperalgesia. Specifically, we study the role of the inhibitory nucleus zona incerta (ZI) in mediating the analgesic effects of MCS. In animals with mechanical and thermal hyperalgesia, we find that stimulation at 50 μA, 50 Hz, and 300 μs square pulses for 30 minutes is sufficient to reverse mechanical and thermal hyperalgesia. We also find that stimulation of the ZI mimics the effects of MCS and that reversible inactivation of ZI blocks the effects of MCS. These findings suggest that the reduction of hyperalgesia may be due to MCS effects on ZI. In an animal model of central pain syndrome, motor cortex stimulation reduces hyperalgesia by activating zona incerta and therefore restoring inhibition in the thalamus.

  8. Nuclear factor-kappa B decoy suppresses nerve injury and improves mechanical allodynia and thermal hyperalgesia in a rat lumbar disc herniation model

    PubMed Central

    Suzuki, Munetaka; Inoue, Gen; Gemba, Takefumi; Watanabe, Tomoko; Ito, Toshinori; Koshi, Takana; Yamauchi, Kazuyo; Yamashita, Masaomi; Orita, Sumihisa; Eguchi, Yawara; Ochiai, Nobuyasu; Kishida, Shunji; Takaso, Masashi; Aoki, Yasuchika; Takahashi, Kazuhisa

    2009-01-01

    Nuclear factor-kappa B (NF-κB) is a gene transcriptional regulator of inflammatory cytokines. We investigated the transduction efficiency of NF-κB decoy to dorsal root ganglion (DRG), as well as the decrease in nerve injury, mechanical allodynia, and thermal hyperalgesia in a rat lumbar disc herniation model. Forty rats were used in this study. NF-κB decoy–fluorescein isothiocyanate (FITC) was injected intrathecally at the L5 level in five rats, and its transduction efficiency into DRG measured. In another 30 rats, mechanical pressure was placed on the DRG at the L5 level and nucleus pulposus harvested from the rat coccygeal disc was transplanted on the DRG. Rats were classified into three groups of ten animals each: a herniation + decoy group, a herniation + oligo group, and a herniation only group. For behavioral testing, mechanical allodynia and thermal hyperalgesia were evaluated. In 15 of the herniation rats, their left L5 DRGs were resected, and the expression of activating transcription factor 3 (ATF-3) and calcitonin gene-related peptide (CGRP) was evaluated immunohistochemically compared to five controls. The total transduction efficiency of NF-κB decoy–FITC in DRG neurons was 10.8% in vivo. The expression of CGRP and ATF-3 was significantly lower in the herniation + decoy group than in the other herniation groups. Mechanical allodynia and thermal hyperalgesia were significantly suppressed in the herniation + decoy group. NF-κB decoy was transduced into DRGs in vivo. NF-κB decoy may be useful as a target for clarifying the mechanism of sciatica caused by lumbar disc herniation. PMID:19308465

  9. Nuclear factor-kappa B decoy suppresses nerve injury and improves mechanical allodynia and thermal hyperalgesia in a rat lumbar disc herniation model.

    PubMed

    Suzuki, Munetaka; Inoue, Gen; Gemba, Takefumi; Watanabe, Tomoko; Ito, Toshinori; Koshi, Takana; Yamauchi, Kazuyo; Yamashita, Masaomi; Orita, Sumihisa; Eguchi, Yawara; Ochiai, Nobuyasu; Kishida, Shunji; Takaso, Masashi; Aoki, Yasuchika; Takahashi, Kazuhisa; Ohtori, Seiji

    2009-07-01

    Nuclear factor-kappa B (NF-kappaB) is a gene transcriptional regulator of inflammatory cytokines. We investigated the transduction efficiency of NF-kappaB decoy to dorsal root ganglion (DRG), as well as the decrease in nerve injury, mechanical allodynia, and thermal hyperalgesia in a rat lumbar disc herniation model. Forty rats were used in this study. NF-kappaB decoy-fluorescein isothiocyanate (FITC) was injected intrathecally at the L5 level in five rats, and its transduction efficiency into DRG measured. In another 30 rats, mechanical pressure was placed on the DRG at the L5 level and nucleus pulposus harvested from the rat coccygeal disc was transplanted on the DRG. Rats were classified into three groups of ten animals each: a herniation + decoy group, a herniation + oligo group, and a herniation only group. For behavioral testing, mechanical allodynia and thermal hyperalgesia were evaluated. In 15 of the herniation rats, their left L5 DRGs were resected, and the expression of activating transcription factor 3 (ATF-3) and calcitonin gene-related peptide (CGRP) was evaluated immunohistochemically compared to five controls. The total transduction efficiency of NF-kappaB decoy-FITC in DRG neurons was 10.8% in vivo. The expression of CGRP and ATF-3 was significantly lower in the herniation + decoy group than in the other herniation groups. Mechanical allodynia and thermal hyperalgesia were significantly suppressed in the herniation + decoy group. NF-kappaB decoy was transduced into DRGs in vivo. NF-kappaB decoy may be useful as a target for clarifying the mechanism of sciatica caused by lumbar disc herniation.

  10. Tempol Ameliorates and Prevents Mechanical Hyperalgesia in a Rat Model of Chemotherapy-Induced Neuropathic Pain

    PubMed Central

    Kim, Hee Kee; Hwang, Seon-Hee; Abdi, Salahadin

    2017-01-01

    Chemotherapy-induced neuropathic pain is difficult to treat and prevent. Tempol decreases cellular superoxide radical levels and oxidative stress. The aims of our study were to investigate the analgesic and preventive effects of tempol on paclitaxel-induced neuropathic pain in rats and to identify the associated mechanisms of action. Neuropathic pain was induced with intraperitoneally injected paclitaxel on four alternate days in male Sprague–Dawley rats. Tempol was administered systemically as a single injection and a continuous infusion before or after the injection of paclitaxel. The mechanical threshold for allodynia, protein levels, and free radical levels were measured using von Frey filaments, Western blotting, and live cell imaging, respectively. After the rats developed neuropathic pain behavior, a single intraperitoneal injection and continuous infusion of tempol ameliorated paclitaxel-induced mechanical allodynia. Systemic infusion of tempol in the early phase of the development of pain behavior prevented the development of paclitaxel-induced pain behavior. Paclitaxel increased the levels of phosphorylated protein kinase C, phosphorylated nuclear factor κB, phosphodiesterase 4D (PDE4D), IL-1β, and monocyte chemoattractant protein-1 in the lumbar dorsal root ganglia; however, tempol decreased these levels. Paclitaxel also increased superoxide levels in a culture of primary dorsal root ganglion cells and tempol decreased these levels. In conclusion, tempol alleviates and prevents chemotherapy-induced neuropathic pain in rats by reducing the levels of inflammatory cytokines and free radicals in dorsal root ganglia. PMID:28138318

  11. Inhibition of endogenous NGF degradation induces mechanical allodynia and thermal hyperalgesia in rats

    PubMed Central

    2013-01-01

    Background We have previously shown a sprouting of sympathetic fibers into the upper dermis of the skin following subcutaneous injection of complete Freund’s adjuvant (CFA) into the hindpaw. This sprouting correlated with an increase in pain-related sensitivity. We hypothesized that this sprouting and pain-related behavior were caused by an increase in nerve growth factor (NGF) levels. In this study, we investigated whether the inhibition of mature NGF degradation, using a matrix metalloproteinase 2 and 9 (MMP-2/9) inhibitor, was sufficient to reproduce a similar phenotype. Results Behavioral tests performed on male Sprague–Dawley rats at 1, 3, 7 and 14 days after intra-plantar MMP-2/9 inhibitor administration demonstrated that acute and chronic injections of the MMP-2/9 inhibitor induced sensitization, in a dose dependent manner, to mechanical, hot and cold stimuli as measured by von Frey filaments, Hargreaves and acetone tests, respectively. Moreover, the protein levels of mature NGF (mNGF) were increased, whereas the levels and enzymatic activity of matrix metalloproteinase 9 were reduced in the glabrous skin of the hind paw. MMP-2/9 inhibition also led to a robust sprouting of sympathetic fibers into the upper dermis but there were no changes in the density of peptidergic nociceptive afferents. Conclusions These findings indicate that localized MMP-2/9 inhibition provokes a pattern of sensitization and fiber sprouting comparable to that previously obtained following CFA injection. Accordingly, the modulation of endogenous NGF levels should be considered as a potential therapeutic target for the management of inflammatory pain associated with arthritis. PMID:23889761

  12. Colocalization of aromatase in spinal cord astrocytes: Differences in expression and relationship to mechanical and thermal hyperalgesia in murine models of a painful and a non-painful bone tumor

    PubMed Central

    O’Brien, Elaine E; Smeester, Branden A; Michlitsch, Kyle S; Lee, Jang-Hern; Beitz, Alvin J

    2015-01-01

    While spinal cord astrocytes play a key role in the generation of cancer pain, there have been no studies that have examined the relationship of tumor-induced astrocyte activation and aromatase expression during the development of cancer pain. Here, we examined tumor-induced mechanical hyperalgesia and cold allodynia, and changes in GFAP and aromatase expression in murine models of painful and non-painful bone cancer. We demonstrate that implantation of fibrosarcoma cells, but not melanoma cells, produces robust mechanical hyperalgesia and cold allodynia in tumor-bearing mice compared to saline-injected controls. Secondly, this increase in mechanical hyperalgesia and cold allodynia is mirrored by significant increases in both spinal astrocyte activity and aromatase expression in the dorsal horn of fibrosarcoma-bearing mice. Importantly, we show that aromatase is only found within a subset of astrocytes and not in neurons in the lumbar spinal cord. Finally, administration of an aromatase inhibitor reduced tumor-induced hyperalgesia in fibrosarcoma-bearing animals. We conclude that a painful fibrosarcoma tumor induces a significant increase in spinal astrocyte activation and aromatase expression and that the up-regulation of aromatase plays a role in the development of bone tumor-induced hyperalgesia. Since spinal aromatase is also upregulated, but to a lesser extent, in non-painful melanoma bone tumors, it may also be neuroprotective and responsive to the changing tumor environment. PMID:26071956

  13. Eccentric Muscle Contraction and Stretching Evoke Mechanical Hyperalgesia and Modulate CGRP and P2X3 Expression in a Functionally Relevant Manner

    PubMed Central

    Dessem, Dean; Ambalavanar, Ranjinidevi; Evancho, Melena; Moutanni, Aicha; Yallampalli, Chandrasekhar; Bai, Guang

    2010-01-01

    Non-invasive, movement-based models were used to investigate muscle pain. In rats, the masseter muscle was rapidly stretched or electrically stimulated during forced lengthening to produce eccentric muscle contractions (EC). Both EC and stretching disrupted scattered myofibers and produced intramuscular plasma extravasation. Pro-inflammatory cytokines (IL-1β, TNF-α, IL-6) and vascular endothelial growth factor (VEGF) were elevated in the masseter 24h following EC. At 48h, neutrophils increased and ED1 macrophages infiltrated myofibers while ED2 macrophages were abundant at 4d. Mechanical hyperalgesia was evident in the ipsilateral head 4h-4d after a single bout of EC and for 7d following multiple bouts (1 bout/d for 4d). Calcitonin gene-related peptide (CGRP) mRNA increased in the trigeminal ganglion 24h following EC while immunoreactive CGRP decreased. By 2d, CGRP-muscle afferent numbers equaled naive numbers implying that CGRP is released following EC and replenished within 2d. EC elevated P2X3 mRNA and increased P2X3-muscle afferent neuron number for 12d while electrical stimulation without muscle contraction altered neither CGRP nor P2X3 mRNA levels. Muscle stretching produced hyperalgesia for 2d whereas contraction alone produced no hyperalgesia. Stretching increased CGRP mRNA at 24h but not CGRP-muscle afferent number at 2–12d. In contrast, stretching significantly increased the number of P2X3-muscle afferent neurons for 12d. The sustained, elevated P2X3 expression evoked by EC and stretching may enhance nociceptor responsiveness to ATP released during subsequent myofiber damage. Movement-based actions such as EC and muscle stretching produce unique tissue responses and modulate neuropeptide and nociceptive receptor expression in a manner particularly relevant to repeated muscle damage. PMID:20207080

  14. The Relationship between Mechanical Hyperalgesia Assessed by Manual Tender Point Examination and Disease Severity in Patients with Chronic Widespread Pain: A Cross-Sectional Study.

    PubMed

    Amris, Kirstine; Wæhrens, Eva Ejlersen; Jespersen, Anders; Stockmarr, Anders; Bennett, Robert; Bliddal, Henning; Danneskiold-Samsøe, Bente

    2014-01-01

    The clinical utility of tender point (TP) examination in patients reporting chronic widespread pain (CWP) is the subject of contemporary debate. The objective of this study was to assess the relationship between mechanical hyperalgesia assessed by manual TP examination and clinical disease severity. 271 women with CWP were recruited from a clinical setting. Data collection included patient-reported symptoms, health-related quality of life variables, and observation-based measures of functional ability, muscle strength, 6-minute walk, and pressure pain thresholds measured by cuff algometry. TP examination was conducted according to ACR-guidelines. Relationships between disease variables and TP count (TPC) were analyzed with logistic regression in a continuum model, allowing the TPC to depend on the included disease variables and two regression models carried out for a TPC threshold level, varying between 1 and 17. The threshold analyses indicated a TPC threshold at 8, above which a large number of disease variables became consistently significant explanatory factors, whereas none of the disease variables reached a significance level in the continuum model. These results support the premise that the presence of mechanical hyperalgesia influences symptomatology in CWP and that the severity of clinical expression is related to a threshold of TPs, rather than being part of a continuum.

  15. Distinct Terminal and Cell Body Mechanisms in the Nociceptor Mediate Hyperalgesic Priming

    PubMed Central

    Ferrari, Luiz F.; Araldi, Dioneia

    2015-01-01

    Hyperalgesic priming, a form of neuroplasticity in nociceptors, is a model of the transition from acute to chronic pain in the rat, which involves signaling from the site of an acute tissue insult in the vicinity of the peripheral terminal of a nociceptor to its cell body that, in turn, induces a signal that travels back to the terminal to mediate a marked prolongation of prostaglandin E2-induced hyperalgesia. In the present experiments, we studied the underlying mechanisms in the cell body and compared them to the mechanisms in the nerve terminal. Injection of a cell-permeant cAMP analog, 8-bromo cAMP, into the dorsal root ganglion induced mechanical hyperalgesia and priming with an onset more rapid than when induced at the peripheral terminal. Priming induced by intraganglion 8-bromo cAMP was prevented by an oligodeoxynucleotide antisense to mRNA for a transcription factor, cAMP response element-binding protein (CREB), and by an inhibitor of importin, which is required for activated CREB to get into the nucleus. While peripheral administration of 8-bromo cAMP also produced hyperalgesia, it did not produce priming. Conversely, interventions administered in the vicinity of the peripheral terminal of the nociceptor that induces priming—PKCε activator, NGF, and TNF-α—when injected into the ganglion produce hyperalgesia but not priming. The protein translation inhibitor cordycepin, injected at the peripheral terminal but not into the ganglion, reverses priming induced at either the ganglion or peripheral terminal of the nociceptor. These data implicate different mechanisms in the soma and terminal in the transition to chronic pain. PMID:25878283

  16. Distinct terminal and cell body mechanisms in the nociceptor mediate hyperalgesic priming.

    PubMed

    Ferrari, Luiz F; Araldi, Dioneia; Levine, Jon D

    2015-04-15

    Hyperalgesic priming, a form of neuroplasticity in nociceptors, is a model of the transition from acute to chronic pain in the rat, which involves signaling from the site of an acute tissue insult in the vicinity of the peripheral terminal of a nociceptor to its cell body that, in turn, induces a signal that travels back to the terminal to mediate a marked prolongation of prostaglandin E2-induced hyperalgesia. In the present experiments, we studied the underlying mechanisms in the cell body and compared them to the mechanisms in the nerve terminal. Injection of a cell-permeant cAMP analog, 8-bromo cAMP, into the dorsal root ganglion induced mechanical hyperalgesia and priming with an onset more rapid than when induced at the peripheral terminal. Priming induced by intraganglion 8-bromo cAMP was prevented by an oligodeoxynucleotide antisense to mRNA for a transcription factor, cAMP response element-binding protein (CREB), and by an inhibitor of importin, which is required for activated CREB to get into the nucleus. While peripheral administration of 8-bromo cAMP also produced hyperalgesia, it did not produce priming. Conversely, interventions administered in the vicinity of the peripheral terminal of the nociceptor that induces priming-PKCε activator, NGF, and TNF-α-when injected into the ganglion produce hyperalgesia but not priming. The protein translation inhibitor cordycepin, injected at the peripheral terminal but not into the ganglion, reverses priming induced at either the ganglion or peripheral terminal of the nociceptor. These data implicate different mechanisms in the soma and terminal in the transition to chronic pain.

  17. The Glt1 glutamate receptor mediates the establishment and perpetuation of chronic visceral pain in an animal model of stress-induced bladder hyperalgesia.

    PubMed

    Ackerman, A Lenore; Jellison, Forrest C; Lee, Una J; Bradesi, Sylvie; Rodríguez, Larissa V

    2016-04-01

    Psychological stress exacerbates interstitial cystitis/bladder pain syndrome (IC/BPS), a lower urinary tract pain disorder characterized by increased urinary frequency and bladder pain. Glutamate (Glu) is the primary excitatory neurotransmitter modulating nociceptive networks. Glt1, an astrocytic transporter responsible for Glu clearance, is critical in pain signaling termination. We sought to examine the role of Glt1 in stress-induced bladder hyperalgesia and urinary frequency. In a model of stress-induced bladder hyperalgesia with high construct validity to human IC/BPS, female Wistar-Kyoto (WKY) rats were subjected to 10-day water avoidance stress (WAS). Referred hyperalgesia and tactile allodynia were assessed after WAS with von Frey filaments. After behavioral testing, we assessed Glt1 expression in the spinal cord by immunoblotting. We also examined the influence of dihydrokainate (DHK) and ceftriaxone (CTX), which downregulate and upregulate Glt1, respectively, on pain development. Rats exposed to WAS demonstrated increased voiding frequency, increased colonic motility, anxiety-like behaviors, and enhanced visceral hyperalgesia and tactile allodynia. This behavioral phenotype correlated with decreases in spinal Glt1 expression. Exogenous Glt1 downregulation by DHK resulted in hyperalgesia similar to that following WAS. Exogenous Glt1 upregulation via intraperitoneal CTX injection inhibited the development of and reversed preexisting pain and voiding dysfunction induced by WAS. Repeated psychological stress results in voiding dysfunction and hyperalgesia that correlate with altered central nervous system glutamate processing. Manipulation of Glu handling altered the allodynia developing after psychological stress, implicating Glu neurotransmission in the pathophysiology of bladder hyperalgesia in the WAS model of IC/BPS.

  18. Pre-injury administration of morphine prevents development of neuropathic hyperalgesia through activation of descending monoaminergic mechanisms in the spinal cord in mice

    PubMed Central

    Rashid, Md Harunor; Ueda, Hiroshi

    2005-01-01

    The present study examined whether pre-injury administration of morphine can prevent partial sciatic nerve injury-induced neuropathic pain in mice. We observed that pre-injury administration of subcutaneous (s.c.) and intracerebroventricular (i.c.v.) morphine dose-dependently prevented the development of both thermal and mechanical hyperalgesia at 7 days following nerve injury in mice. The pre-injury morphine (s.c.)-induced analgesia was significantly blocked by pretreatment with naloxone injected s.c. or i.c.v., but not i.t., suggesting that systemic morphine produced the pre-emptying effects mainly by acting at the supra-spinal sites. Since it is believed that activation of descending monoaminergic mechanisms in spinal cord largely contributes to the supra-spinal analgesic effects of morphine, we investigated the involvement of serotonergic and noradrenergic mechanisms in spinal cord in the pre-injury morphine-induced analgesic effects. We found that pre-injury s.c. morphine-induced analgesic effect was significantly blocked by i.t. pretreatment with serotonergic antagonist, methysergide and noradrenergic antagonist, phentolamine. In addition, pre-injury i.t. injection of serotonin uptake inhibitor, fluoxetine and α2-adrenergic agonist, clonidine significantly prevented the neuropathic hyperalgesia. We next examined whether pre-injury morphine prevented the expression of neuronal hyperactivity markers such as c-Fos and protein kinase C γ (PKCγ) in the spinal dorsal horn. We found that pre-injury administration of s.c. morphine prevented increased expressions of both c-Fos and PKCγ observed following nerve injury. Similar results were obtained with i.t. fluoxetine and clonidine. Altogether these results suggest that pre-injury administration of morphine might prevent the development of neuropathic pain through activation of descending monoaminergic pain inhibitory pathways. PMID:15932652

  19. Cold allodynia and hyperalgesia in neuropathic pain: the effect of N-methyl-D-aspartate (NMDA) receptor antagonist ketamine--a double-blind, cross-over comparison with alfentanil and placebo.

    PubMed

    Jørum, E; Warncke, T; Stubhaug, A

    2003-02-01

    Cold allodynia and hyperalgesia are frequent clinical findings in patients with neuropathic pain. While there have been several clinical studies showing the involvement of central sensitization mechanisms and N-methyl-D-aspartate (NMDA) receptor activation in mechanical allodynia/hyperalgesia and ongoing pain, the mechanisms of thermal allodynia and hyperalgesia have received less attention. The aim of the present study was to examine the effect of the NMDA-receptor antagonist ketamine on thermal allodynia/hyperalgesia, ongoing pain and mechanical allodynia/hyperalgesia in patients with neuropathic pain (11 patients with post-traumatic neuralgia and one patient with post-herpetic neuralgia). All the patients were known to suffer from severe cold allodynia (cold pain detection threshold (CPDT): 23.8 degrees C, median value). The mu-opioid agonist alfentanil was used as an active control. The study design was double-blind and placebo-controlled and the drugs were administered i.v. (bolus dose and infusion). CPDT in the asymptomatic contralateral area was found to be significantly decreased (cold allodynia) compared to CPDT in site- and age-matched normal controls. Heat pain detection thresholds were found to be normal and no consistent heat hyperalgesia occurred. Alfentanil significantly reduced cold allodynia (by increasing CPDT) in symptomatic area (P=0.0076). Ketamine did not significantly increase the threshold. Significant and marked reductions of hyperalgesia to cold (visual analogue score at threshold value) were seen following both alfentanil (4.5 before, 1.4 after, median value) and ketamine (6.8 before, 0.4 after, median value). Alfentanil and ketamine also significantly reduced ongoing pain and mechanical hyperalgesia. It is concluded that NMDA-receptor mediated central sensitization is involved in cold hyperalgesia, but since CPDT remained unaltered, it is likely that other mechanisms are present.

  20. PHARMACOLOGIC TREATMENT OF HYPERALGESIA EXPERIMENTALLY INDUCED BY NUCLEUS PULPOSUS

    PubMed Central

    de Souza Grava, André Luiz; Ferrari, Luiz Fernando; Parada, Carlos Amílcar; Defino, Helton Luiz Aparecido

    2015-01-01

    Objective: To evaluate the effect of anti-inflammatory drugs (dexamethasone, indomethacin, atenolol and indomethacin plus atenolol) and analgesic drugs (morphine) on hyperalgesia experimentally induced by the nucleus pulposus (NP) in contact with the L5 dorsal root ganglion (DRG). Methods: Thirty male Wistar rats of weights ranging from 220 to 250 g were used in the study. Hyperalgesia was induced by means of a fragment of NP removed from the sacrococcygeal region that was placed in contact with the L5 dorsal root ganglion. The 30 animals were divided into experimental groups according to the drug used. The drugs were administered for two weeks after the surgical procedure to induce hyperalgesia. Mechanical and thermal hyperalgesia was evaluated using the paw pressure test, von Frey electronic test and Hargreaves test, over a seven-week period. Results: The greatest reduction of hyperalgesia was observed in the group of animals treated with morphine, followed by dexamethasone, indomethacin and atenolol. Reductions in hyperalgesia were observed after drug administration ceased, except for the group of animals treated with morphine, in which there was an increase in hyperalgesia after discontinuation of the treatment. Conclusion: Hyperalgesia induced by NP contact with the DRG can be reduced through administration of anti-inflammatory and analgesic drugs, but a greater reduction was observed with the administration of dexamethasone. PMID:27026966

  1. Characterization of nociceptin hyperalgesia and allodynia in conscious mice

    PubMed Central

    Hara, Naoki; Minami, Toshiaki; Okuda-Ashitaka, Emiko; Sugimoto, Tetsuo; Sakai, Masato; Onaka, Masahiko; Mori, Hidemaro; Imanishi, Toshihiro; Shingu, Koh; Ito, Seiji

    1997-01-01

    demonstrate that, whereas the mechanisms of the nociceptin-induced allodynia and hyperalgesia are evidently distinct, they involve a common neurochemical event beginning with the disinhibition of the inhibitory glycinergic response. Morphine may induce allodynia through a pathway common to nociceptin, but the nociceptin receptor does not mediate the action of high doses of morphine. PMID:9179380

  2. A comparison of hyperalgesia and neurogenic inflammation induced by melittin and capsaicin in humans.

    PubMed

    Sumikura, H; Andersen, O K; Drewes, A M; Arendt-Nielsen, L

    2003-02-13

    Melittin (a main compound of bee venom) and capsaicin were injected intradermally in healthy human volunteers: (1) to study secondary mechanical hyperalgesia (static hyperalgesia and dynamic hyperalgesia) around the injection site; and (2) to correlate the sensory changes to the neurogenic inflammation assessed by laser-doppler blood flowmetry. Melittin 50 microg and capsaicin 10 microg induced comparable spontaneous pain and increased blood flow (neurogenic inflammation). Intradermal injection of melittin induced regions of secondary mechanical hyperalgesia around the injection site, however, they were not as large as the hyperalgesia induced by capsaicin. This is the first report studying mechanical hyperalgesia induced by melittin in humans, and the results were in agreement with the previous observations in rats. Melittin seems to be a valuable model to study a possible contribution of neurogenic inflammation to hyperalgesia in humans.

  3. IB4(+) nociceptors mediate persistent muscle pain induced by GDNF

    PubMed Central

    Alvarez, Pedro; Chen, Xiaojie; Bogen, Oliver; Green, Paul G.

    2012-01-01

    Skeletal muscle is a well-known source of glial cell line-derived neurotrophic factor (GDNF), which can produce mechanical hyperalgesia. Since some neuromuscular diseases are associated with both increased release of GDNF and intense muscle pain, we explored the role of GDNF as an endogenous mediator in muscle pain. Intramuscularly injected GDNF induced a dose-dependent (0.1–10 ng/20 μl) persistent (up to 3 wk) mechanical hyperalgesia in the rat. Once hyperalgesia subsided, injection of prostaglandin E2 at the site induced a prolonged mechanical hyperalgesia (>72 h) compared with naïve rats (<4 h; hyperalgesic priming). Selective neurotoxic destruction of IB4(+) nociceptors attenuated both GDNF hyperalgesia and hyperalgesic priming. Ergonomic muscular injury induced by eccentric exercise or mechanical vibration increased muscle GDNF levels at 24 h, a time point where rats also exhibited marked muscle hyperalgesia. Intrathecal antisense oligodeoxynucleotides to mRNA encoding GFRα1, the canonical binding receptor for GDNF, reversibly inhibited eccentric exercise- and mechanical vibration-induced muscle hyperalgesia. Finally, electrophysiological recordings from nociceptors innervating the gastrocnemius muscle in anesthetized rats, revealed significant increase in response to sustained mechanical stimulation after local GDNF injection. In conclusion, these data indicate that GDNF plays a role as an endogenous mediator in acute and induction of chronic muscle pain, an effect likely to be produced by GDNF action at GFRα1 receptors located in IB4(+) nociceptors. PMID:22914655

  4. Gabapentin enhances anti-nociceptive effects of morphine on heat, cold, and mechanical hyperalgesia in a rat model of neuropathic pain

    PubMed Central

    Hamidi, Gholam Ali; Jafari-Sabet, Majid; Abed, Alireza; Mesdaghinia, Azam; Mahlooji, Mohadeseh; Banafshe, Hamid Reza

    2014-01-01

    Objective(s): Neuropathic pain is caused by lesions or diseases affecting the somatosensory system and often responds poorly to typical medications. In this study, we evaluated anti-nociceptive effects of morphine, gabapentin and their combination on heat hyperalgesia, cold and mechanical allodynia in chronic constriction injury (CCI) model of neuropathic pain in rats. Materials and Methods: Morphine (2, 4 and 8 mg/kg) and gabapentin (5, 10 and 20 mg/kg) were administered either alone or in combination (morphine 2 mg/kg and gabapentin 5 mg/kg). Results: Our results showed that morphine and gabapentin alone produce anti-nociceptive effects at higher doses (morphine 4 and 8 mg/kg and gabapentin 10 and 20 mg/kg) whereas their combination resulted in better analgesia at lower doses as compared to other treatment groups (morphine 2 mg/kg or gabapentin 5 mg/kg). Conclusion: These findings suggest that gabapentin potentiates the analgesic effects of morphine in the chronic constriction injury (CCI) model of neuropathic pain and combination of these drugs may be considered as a beneficial treatment for neuropathic pain. PMID:25729543

  5. Spinal prostaglandin E receptors of the EP2 subtype and the glycine receptor alpha3 subunit, which mediate central inflammatory hyperalgesia, do not contribute to pain after peripheral nerve injury or formalin injection.

    PubMed

    Hösl, Katharina; Reinold, Heiko; Harvey, Robert J; Müller, Ulrike; Narumiya, Shuh; Zeilhofer, Hanns Ulrich

    2006-12-15

    Inflammation, peripheral nerve injury and chemical irritants can cause central sensitization in pain pathways. Prostaglandins produced in the CNS induce central sensitization during inflammation mainly by relieving nociceptive neurons from glycinergic inhibition. We have recently identified spinal prostaglandin E receptors of the EP2 subtype (EP2 receptors) and the glycine receptor alpha3 subunit (GlyR alpha3) as signal transduction elements involved in the generation of central inflammatory hyperalgesia. It is however still unknown to what extent inhibition of glycine receptors by PGE2 contributes to neuropathic or chemically induced pain. To address this question, we have analyzed mice deficient in the EP2 receptor (EP2-/- mice) or in the GlyR alpha3 subunit (GlyR alpha3-/- mice) using the chronic constriction injury (CCI) model of neuropathic pain and the formalin test. We found that EP2-/- mice and GlyR alpha3-/- mice develop thermal and mechanical hyperalgesia in the CCI model indistinguishable from that seen in wild-type mice. In the formalin test, EP2-/- mice, but not GlyR alpha3-/- mice, exhibited reduced nocifensive behavior. The lack of a phenotype in GlyR alpha3-/- mice together with the absence of a facilitating effect of intrathecal PGE2 on formalin-induced nociception in wild-type mice suggests that peripheral rather than spinal EP2 receptors are involved. These results indicate that inhibition of glycinergic neurotransmission by EP2 receptor activation does not contribute to pain following peripheral nerve injury or chemical irritation with formalin. Our results thus provide further evidence that inflammatory hyperalgesia and neuropathic pain involve different mechanisms of central sensitization.

  6. Differential coding of hyperalgesia in the human brain: a functional MRI study.

    PubMed

    Maihöfner, Christian; Handwerker, Hermann O

    2005-12-01

    Neuropathic pain can be both ongoing or stimulus-induced. Stimulus-induced pain, also known as hyperalgesia, can be differentiated into primary and secondary hyperalgesia. The former results from sensitization of peripheral nociceptive structures, the latter involves sensitization processes within the central nervous system (CNS). Hypersensitivity towards heat stimuli, i.e. thermal hyperalgesia, is a key feature of primary hyperalgesia, whereas secondary hyperalgesia is characterized by hypersensitivity towards mechanical (e.g. pin-prick) stimulation. Using functional magnetic resonance imaging (fMRI), we investigated if brain activation patterns associated with primary and secondary hyperalgesia might differ. Thermal and pin-prick hyperalgesia were induced on the left forearm in 12 healthy subjects by topical capsaicin (2.5%, 30 min) application. Equal pain intensities of both hyperalgesia types were applied during fMRI experiments, based on previous quantitative sensory testing. Simultaneously, subjects had to rate the unpleasantness of stimulus-related pain. Pin-prick hyperalgesia (i.e. subtraction of brain activations during pin-prick stimulation before and after capsaicin exposure) led to activations of primary and secondary somatosensory cortices (S1 and S2), associative-somatosensory cortices, insula and superior and inferior frontal cortices (SFC, IFC). Brain areas activated during thermal hyperalgesia (i.e. subtraction of brain activations during thermal stimulation before and after capsaicin exposure) were S1 and S2, insula, associative-somatosensory cortices, cingulate cortex (GC), SFC, middle frontal cortex (MFC) and IFC. When compared to pin-prick hyperalgesia, thermal hyperalgesia led to an increased activation of bilateral anterior insular cortices, MFC, GC (Brodmann area 24' and 32') and contralateral SFC and IFC, despite equal pain intensities. Interestingly, stronger activations of GC, contralateral MFC and anterior insula significantly correlated

  7. Phosphorylation of TRPV1 by cyclin-dependent kinase 5 promotes TRPV1 surface localization, leading to inflammatory thermal hyperalgesia.

    PubMed

    Liu, Jiao; Du, Junxie; Yang, Yanrui; Wang, Yun

    2015-11-01

    Cyclin-dependent kinase 5 (Cdk5) is an important serine/threonine kinase that plays critical roles in many physiological processes. Recently, Cdk5 has been reported to phosphorylate TRPV1 at threonine 407 (Thr-407) in humans (Thr-406 in rats), which enhances the function of TRPV1 channel and promotes thermal hyperalgesia in the complete Freund's adjuvant (CFA)-induced inflammatory pain rats. However, the underlying mechanisms are still unknown. Here, we demonstrate that Cdk5 phosphorylates TRPV1 at Threonine 406 and promotes the surface localization of TRPV1, leading to inflammatory thermal hyperalgesia. The mutation of Thr-406 of TRPV1 to alanine reduced the interaction of TRPV1 with the cytoskeletal elements and decreased the binding of TRPV1 with the motor protein KIF13B, which led to reduced surface distribution of TRPV1. Disrupting the phosphorylation of TRPV1 at Thr-406 dramatically reduced the surface level of TRPV1 in HEK 293 cells after transient expression and the channel function in cultured dorsal root ganglion (DRG) neurons. Notably, intrathecal administration of the interfering peptide against the phosphorylation of Thr-406 alleviated heat hyperalgesia and reduced the surface level of TRPV1 in inflammatory pain rats. Together, these results demonstrate that Cdk5-mediated phosphorylation of TRPV1 at Thr-406 increases the surface level and the function of TRPV1, while the TAT-T406 peptide can effectively attenuate thermal hyperalgesia. Our studies provide a potential therapy for inflammatory pain.

  8. Sustained Morphine Administration Induces TRPM8-Dependent Cold Hyperalgesia.

    PubMed

    Gong, Kerui; Jasmin, Luc

    2017-02-01

    It is not uncommon for patients chronically treated with opioids to exhibit opioid-induced hyperalgesia, and this has been widely reported clinically and experimentally. The molecular substrate for this hyperalgesia is multifaceted, and associated with a complex neural reorganization even in the periphery. For instance, we have recently shown that chronic morphine-induced heat hyperalgesia is associated with an increased expression of GluN2B containing N-methyl-D-aspartate receptors, as well as of the neuronal excitatory amino acid transporter 3/excitatory amino acid carrier 1, in small-diameter primary sensory neurons only. Cold allodynia is also a common complaint of patients chronically treated with opioids, yet its molecular mechanisms remain to be understood. Here we present evidence that the cold sensor TRPM8 channel is involved in opioid-induced hyperalgesia. After 7 days of morphine administration, we observed an upregulation of TRPM8 channels using patch clamp recording on sensory neurons and Western blot analysis on dorsal root ganglia. The selective TRPM8 antagonist RQ-00203078 blocked cold hyperalgesia in morphine-treated rats. Also, TRPM8 knockout mice failed to develop cold hyperalgesia after chronic administration of morphine. Our results show that chronic morphine upregulates TRPM8 channels, which is in contrast with the previous finding that acute morphine triggers TRPM8 internalization.

  9. Sustained Suppression of Hyperalgesia during Latent Sensitization by μ-, δ-, and κ-opioid receptors and α2A Adrenergic Receptors: Role of Constitutive Activity

    PubMed Central

    Walwyn, Wendy M.; Chen, Wenling; Kim, Hyeyoung; Minasyan, Ani; Ennes, Helena S.; McRoberts, James A.

    2016-01-01

    Many chronic pain disorders alternate between bouts of pain and periods of remission. The latent sensitization model reproduces this in rodents by showing that the apparent recovery (“remission”) from inflammatory or neuropathic pain can be reversed by opioid antagonists. Therefore, this remission represents an opioid receptor-mediated suppression of a sustained hyperalgesic state. To identify the receptors involved, we induced latent sensitization in mice and rats by injecting complete Freund's adjuvant (CFA) in the hindpaw. In WT mice, responses to mechanical stimulation returned to baseline 3 weeks after CFA. In μ-opioid receptor (MOR) knock-out (KO) mice, responses did not return to baseline but partially recovered from peak hyperalgesia. Antagonists of α2A-adrenergic and δ-opioid receptors reinstated hyperalgesia in WT mice and abolished the partial recovery from hyperalgesia in MOR KO mice. In rats, antagonists of α2A adrenergic and μ-, δ-, and κ-opioid receptors reinstated hyperalgesia during remission from CFA-induced hyperalgesia. Therefore, these four receptors suppress hyperalgesia in latent sensitization. We further demonstrated that suppression of hyperalgesia by MORs was due to their constitutive activity because of the following: (1) CFA-induced hyperalgesia was reinstated by the MOR inverse agonist naltrexone (NTX), but not by its neutral antagonist 6β-naltrexol; (2) pro-enkephalin, pro-opiomelanocortin, and pro-dynorphin KO mice showed recovery from hyperalgesia and reinstatement by NTX; (3) there was no MOR internalization during remission; (4) MORs immunoprecipitated from the spinal cord during remission had increased Ser375 phosphorylation; and (5) electrophysiology recordings from dorsal root ganglion neurons collected during remission showed constitutive MOR inhibition of calcium channels. SIGNIFICANCE STATEMENT Chronic pain causes extreme suffering to millions of people, but its mechanisms remain to be unraveled. Latent

  10. Low doses of cyclic AMP-phosphodiesterase inhibitors rapidly evoke opioid receptor-mediated thermal hyperalgesia in naïve mice which is converted to prominent analgesia by cotreatment with ultra-low-dose naltrexone.

    PubMed

    Crain, Stanley M; Shen, Ke-Fei

    2008-09-22

    Systemic (s.c.) injection in naïve mice of cyclic AMP-phosphodiesterase (cAMP-PDE) inhibitors, e.g. 3-isobutyl-1-methylxanthine [(IBMX) or caffeine, 10 mg/kg] or the more specific cAMP-PDE inhibitor, rolipram (1 mug/kg), rapidly evokes thermal hyperalgesia (lasting >5 h). These effects appear to be mediated by enhanced excitatory opioid receptor signaling, as occurs during withdrawal in opioid-dependent mice. Cotreatment of these mice with ultra-low-dose naltrexone (NTX, 0.1 ng/kg-1 pg/kg, s.c.) results in prominent opioid analgesia (lasting >4 h) even when the dose of rolipram is reduced to 1 pg/kg. Cotreatment of these cAMP-PDE inhibitors in naïve mice with an ultra-low-dose (0.1 ng/kg) of the kappa-opioid receptor antagonist, nor-binaltorphimine (nor-BNI) or the mu-opioid receptor antagonist, beta-funaltrexamine (beta-FNA) also results in opioid analgesia. These excitatory effects of cAMP-PDE inhibitors in naïve mice may be mediated by enhanced release of small amounts of endogenous bimodally-acting (excitatory/inhibitory) opioid agonists by neurons in nociceptive networks. Ultra-low-dose NTX, nor-BNI or beta-FNA selectively antagonizes high-efficacy excitatory (hyperalgesic) Gs-coupled opioid receptor-mediated signaling in naïve mice and results in rapid conversion to inhibitory (analgesic) Gi/Go-coupled opioid receptor-mediated signaling which normally requires activation by much higher doses of opioid agonists. Cotreatment with a low subanalgesic dose of kelatorphan, an inhibitor of multiple endogenous opioid peptide-degrading enzymes, stabilizes endogenous opioid agonists released by cAMP-PDE inhibitors, resulting in conversion of the hyperalgesia to analgesia without requiring selective blockade of excitatory opioid receptor signaling. The present study provides a novel pharmacologic paradigm that may facilitate development of valuable non-narcotic clinical analgesics utilizing cotreatment with ultra-low-dose rolipram plus ultra-low-dose NTX or related

  11. The Complement System Component C5a Produces Thermal Hyperalgesia via Macrophage-to-Nociceptor Signaling That Requires NGF and TRPV1

    PubMed Central

    Shutov, Leonid P.; Warwick, Charles A.; Shi, Xiaoyu; Gnanasekaran, Aswini; Shepherd, Andrew J.; Mohapatra, Durga P.; Woodruff, Trent M.; Clark, J. David

    2016-01-01

    The complement cascade is a principal component of innate immunity. Recent studies have underscored the importance of C5a and other components of the complement system in inflammatory and neuropathic pain, although the underlying mechanisms are largely unknown. In particular, it is unclear how the complement system communicates with nociceptors and which ion channels and receptors are involved. Here we demonstrate that inflammatory thermal and mechanical hyperalgesia induced by complete Freund's adjuvant was accompanied by C5a upregulation and was markedly reduced by C5a receptor (C5aR1) knock-out or treatment with the C5aR1 antagonist PMX53. Direct administration of C5a into the mouse hindpaw produced strong thermal hyperalgesia, an effect that was absent in TRPV1 knock-out mice, and was blocked by the TRPV1 antagonist AMG9810. Immunohistochemistry of mouse plantar skin showed prominent expression of C5aR1 in macrophages. Additionally, C5a evoked strong Ca2+ mobilization in macrophages. Macrophage depletion in transgenic macrophage Fas-induced apoptosis mice abolished C5a-dependent thermal hyperalgesia. Examination of inflammatory mediators following C5a injection revealed a rapid upregulation of NGF, a mediator known to sensitize TRPV1. Preinjection of an NGF-neutralizing antibody or Trk inhibitor GNF-5837 prevented C5a-induced thermal hyperalgesia. Notably, NGF-induced thermal hyperalgesia was unaffected by macrophage depletion. Collectively, these results suggest that complement fragment C5a induces thermal hyperalgesia by triggering macrophage-dependent signaling that involves mobilization of NGF and NGF-dependent sensitization of TRPV1. Our findings highlight the importance of macrophage-to-neuron signaling in pain processing and identify C5a, NGF, and TRPV1 as key players in this cross-cellular communication. SIGNIFICANCE STATEMENT This study provides mechanistic insight into how the complement system, a key component of innate immunity, regulates the

  12. Metallopeptidase inhibition potentiates bradykinin-induced hyperalgesia

    PubMed Central

    Gomez, Ruben; Por, Elaine D.; Berg, Kelly A.; Clarke, William P.; Glucksman, Marc J.; Jeske, Nathaniel A.

    2011-01-01

    The neuropeptide bradykinin (BK) sensitizes nociceptor activation following its release in response to inflammatory injury. Thereafter, the bioactivity of bradykinin is controlled by the enzymatic activities of circulating peptidases. One such enzyme, the metalloendopeptidase EC3.4.24.15 (EP24.15), is co-expressed with bradykinin receptors in primary afferent neurons. In this study, utilizing approaches encompassing pharmacology, biochemistry, cell biology and behavioral animal models, we discover a crucial role for EP24.15 and the closely-related EP24.16 in modulating bradykinin-mediated hyperalgesia. Pharmacological analyses indicate that EP24.15 and EP24.16 inhibition significantly enhances bradykinin type-2 receptor activation by bradykinin in primary trigeminal ganglia cultures. In addition, bradykinin-induced sensitization of TRPV1 activation is increased in the presence of the EP24.15/16 inhibitor JA-2. Furthermore, behavioral analyses illustrate a significant dose-response relationship between JA-2 and bradykinin-mediated thermal hyperalgesia. These results indicate an important physiological role for the metallopeptidases EP24.15 and EP24.16 in regulating bradykinin-mediated sensitization of primary afferent nociceptors. PMID:21458920

  13. Persistent Nociception Triggered by Nerve Growth Factor (NGF) Is Mediated by TRPV1 and Oxidative Mechanisms.

    PubMed

    Eskander, Michael A; Ruparel, Shivani; Green, Dustin P; Chen, Paul B; Por, Elaine D; Jeske, Nathaniel A; Gao, Xiaoli; Flores, Eric R; Hargreaves, Kenneth M

    2015-06-03

    Nerve growth factor (NGF) is elevated in certain chronic pain conditions and is a sufficient stimulus to cause lasting pain in humans, but the actual mechanisms underlying the persistent effects of NGF remain incompletely understood. We developed a rat model of NGF-induced persistent thermal hyperalgesia and mechanical allodynia to determine the role of transient receptor potential vanilloid 1 (TRPV1) and oxidative mechanisms in the persistent effects of NGF. Persistent thermal hypersensitivity and mechanical allodynia require de novo protein translation and are mediated by TRPV1 and oxidative mechanisms. By comparing effects after systemic (subcutaneous), spinal (intrathecal) or hindpaw (intraplantar) injections of test compounds, we determined that TRPV1 and oxidation mediate persistent thermal hypersensitivity via peripheral and spinal sites of action and mechanical allodynia via only a spinal site of action. Therefore, NGF-evoked thermal and mechanical allodynia are mediated by spatially distinct mechanisms. NGF treatment evoked sustained increases in peripheral and central TRPV1 activity, as demonstrated by increased capsaicin-evoked nocifensive responses, increased calcitonin gene-related peptide release from hindpaw skin biopsies, and increased capsaicin-evoked inward current and membrane expression of TRPV1 protein in dorsal root ganglia neurons. Finally, we showed that NGF treatment increased concentrations of linoleic and arachidonic-acid-derived oxidized TRPV1 agonists in spinal cord and skin biopsies. Furthermore, increases in oxidized TRPV1-active lipids were reduced by peripheral and spinal injections of compounds that completely blocked persistent nociception. Collectively, these data indicate that NGF evokes a persistent nociceptive state mediated by increased TRPV1 activity and oxidative mechanisms, including increased production of oxidized lipid TRPV1 agonists.

  14. Fear of pain potentiates nocebo hyperalgesia.

    PubMed

    Aslaksen, Per M; Lyby, Peter S

    2015-01-01

    Nocebo hyperalgesia has received sparse experimental attention compared to placebo analgesia. The aim of the present study was to investigate if personality traits and fear of pain could predict experimental nocebo hyperalgesia. One hundred and eleven healthy volunteers (76 females) participated in an experimental study in which personality traits and fear of pain were measured prior to induction of thermal heat pain. Personality traits were measured by the Big-Five Inventory-10. Fear of pain was measured by the Fear of Pain Questionnaire III. Heat pain was induced by a PC-controlled thermode. Pain was measured by a computerized visual analog scale. Stress levels during the experiment were measured by numerical rating scales. The participants were randomized to a Nocebo group or to a no-treatment Natural History group. The results revealed that pain and stress levels were significantly higher in the Nocebo group after nocebo treatment. Mediation analysis showed that higher levels of the Fear of Pain Questionnaire III factor "fear of medical pain" significantly increased stress levels after nocebo treatment and that higher stress levels were associated with increased nocebo hyperalgesic responses. There were no significant associations between any of the personality factors and the nocebo hyperalgesic effect. The results from the present study suggest that dispositional fear of pain might be a useful predictor for nocebo hyperalgesia and emotional states concomitant with expectations of increased pain. Furthermore, measurement of traits that are specific to pain experience is probably better suited for prediction of nocebo hyperalgesic responses compared to broad measures of personality.

  15. TDAG8 involved in initiating inflammatory hyperalgesia and establishing hyperalgesic priming in mice

    PubMed Central

    Dai, Shih-Ping; Huang, Ya-Han; Chang, Chung-Jen; Huang, Yu-Fen; Hsieh, Wei-Shan; Tabata, Yasuhiko; Ishii, Satoshii; Sun, Wei-Hsin

    2017-01-01

    Chronic pain, resulting from injury, arthritis, and cancer, is often accompanied by inflammation. High concentrations of protons found in inflamed tissues results in tissue acidosis, a major cause of pain and hyperalgesia. Acidosis signals may mediate a transition from acute to chronic hyperalgesia (hyperalgesic priming) via proton-sensing G-protein-coupled receptors (GPCRs). The expression of T-cell death-associated gene 8 (TDAG8), a proton-sensing GPCR, is increased during inflammatory hyperalgesia. Attenuating TDAG8 expression in the spinal cord inhibits bone cancer pain, but whether TDAG8 is involved in inflammatory hyperalgesia or hyperalgesic priming remains unclear. In this study, we used TDAG8-knockout or -knockdown to explore the role of TDAG8 in pain. Suppressed TDAG8 expression delayed the onset of inflammatory hyperalgesia and shortened hyperalgesic time in mice. In a dual acid-injection model (acid [pH 5.0] injected twice, 5 days apart), shRNA inhibition of TDAG8 shortened the duration of the second hyperalgesia. Similar results were found in TDAG8-deficient mice. The dual administration of TDAG8 agonist also confirmed that TDAG8 is involved in hyperalgsic priming. Accordingly, TDAG8 may mediate acidosis signals to initiate inflammatory hyperalgesia and establish hyperalgesic priming. PMID:28145512

  16. Characterizing pinprick-evoked brain potentials before and after experimentally induced secondary hyperalgesia

    PubMed Central

    Mouraux, André; Groneberg, Antonia H.; Pfau, Doreen B.; Treede, Rolf-Detlef; Klein, Thomas

    2015-01-01

    Secondary hyperalgesia is believed to be a key feature of “central sensitization” and is characterized by enhanced pain to mechanical nociceptive stimuli. The aim of the present study was to characterize, using EEG, the effects of pinprick stimulation intensity on the magnitude of pinprick-elicited brain potentials [event-related potentials (ERPs)] before and after secondary hyperalgesia induced by intradermal capsaicin in humans. Pinprick-elicited ERPs and pinprick-evoked pain ratings were recorded in 19 healthy volunteers, with mechanical pinprick stimuli of varying intensities (0.25-mm probe applied with a force extending between 16 and 512 mN). The recordings were performed before (T0) and 30 min after (T1) intradermal capsaicin injection. The contralateral noninjected arm served as control. ERPs elicited by stimulation of untreated skin were characterized by 1) an early-latency negative-positive complex peaking between 120 and 250 ms after stimulus onset (N120-P240) and maximal at the vertex and 2) a long-lasting positive wave peaking 400–600 ms after stimulus onset and maximal more posterior (P500), which was correlated to perceived pinprick pain. After capsaicin injection, pinprick stimuli were perceived as more intense in the area of secondary hyperalgesia and this effect was stronger for lower compared with higher stimulus intensities. In addition, there was an enhancement of the P500 elicited by stimuli of intermediate intensity, which was significant for 64 mN. The other components of the ERPs were unaffected by capsaicin. Our results suggest that the increase in P500 magnitude after capsaicin is mediated by facilitated mechanical nociceptive pathways. PMID:26334010

  17. Descending facilitatory pathways from the RVM initiate and maintain bilateral hyperalgesia after muscle insult

    PubMed Central

    Tillu, DV; Gebhart, GF; Sluka, KA

    2008-01-01

    The rostral ventromedial medulla (RVM) is involved in facilitation of spinal nociceptive processing and generation of hyperalgesia in inflammatory and neuropathic pain models. We hypothesized that the bilateral hyperalgesia that develops after repeated intramuscular injections of acidic saline is initiated and maintained by activation of descending facilitatory pathways from the RVM. Male Sprague-Dawley rats were implanted with intracerebral guide cannulae into the nucleus raphe magnus (NRM) or the nucleus gigantocellularis (Gi). Two injections of acidic saline into one gastrocnemius muscle 5 days apart leads to robust hyperalgesia after the second injection. Either ropivacaine (local anesthetic) or vehicle (control) was microinjected into the RVM prior to the first intramuscular acid injection, prior to the second injection, or 24h after the second injection. Mechanical withdrawal thresholds of the paw (von Frey filaments) and the muscle (tweezer) were measured before and 24h after induction of hyperalgesia. The withdrawal thresholds for both the paw (cutaneous secondary hyperalgesia) and muscle (primary hyperalgesia) were decreased 24h after the second intramuscular acid injection in the vehicle control groups. Administration of ropivacaine prior to the first intramuscular acid injection had no effect on development of either cutaneous or muscle hyperalgesia that develops after the second injection. However, neither cutaneous nor muscle hyperalgesia developed in the group treated with ropivacaine prior to the second intramuscular injection. Ropivacaine also significantly reversed the hyperalgesia in the group treated 24h after the second intramuscular acid injection. Thus, the RVM is critical for both the development and maintenance of hyperalgesia after muscle insult. PMID:17764841

  18. ASIC3 is required for development of fatigue-induced hyperalgesia

    PubMed Central

    Gregory, Nicholas S.; Brito, Renan G.; Oliveira Fusaro, Maria Cláudia G; Sluka, Kathleen A.

    2015-01-01

    An acute bout of exercise can exacerbate pain, hindering participation in regular exercise and daily activities. The mechanisms underlying pain in response to acute exercise are poorly understood. We hypothesized that proton accumulation during muscle fatigue activates ASIC3 on muscle nociceptors to produce hyperalgesia. We investigated the role of ASIC3 using genetic and pharmacological approaches in a model of fatigue-enhanced hyperalgesia. This model uses two injections of pH 5.0 saline into muscle in combination with an electrically-induced fatigue of the same muscle just prior to the second injection of acid to induce mechanical hyperalgesia. We show a significant decrease in muscle force and decrease in muscle pH after 6 minutes of electrical stimulation. Genetic deletion of ASIC3 using knockout mice and pharmacological blockade of ASIC3 with APETx2 in muscle prevents the fatigue-enhanced hyperalgesia. However, ASIC3−/− mice and APETx2 have no effect on the fatigue response. Genetic deletion of ASIC3 in primary afferents innervating muscle using an HSV-1 expressing miRNA to ASIC3 surprisingly had no effect on the development of the hyperalgesia. Muscle fatigue increased the number of macrophages in muscle, and removal of macrophages from muscle with clodronate liposomes prevented the development of fatigue-enhanced hyperalgesia. Thus, these data suggest that fatigue reduces pH in muscle that subsequently activates ASIC3 on macrophages to enhance hyperalgesia to muscle insult. PMID:25577172

  19. Contributions of spinal D-amino acid oxidase to chronic morphine-induced hyperalgesia.

    PubMed

    Ma, Shuai; Li, Xin-Yan; Gong, Nian; Wang, Yong-Xiang

    2015-12-10

    Spinal D-amino acid oxidase (DAAO) is an FAD-dependent peroxisomal flavoenzyme which mediates the conversion of neutral and polar D-amino acids (including D-serine) to the corresponding α-keto acids, and simultaneously produces hydrogen peroxide and ammonia. This study has aimed to explore the potential contributions of spinal DAAO and its mediated hydrogen peroxide/D-serine metabolism to the development of morphine-induced hyperalgesia. Bi-daily subcutaneous injections of morphine to mice over 7 days induced thermal hyperalgesia as measured by both the hot-plate and tail-immersion tests, and spinal astroglial activation with increased spinal gene expression of DAAO, glial fibrillary acidic protein (GFAP) and pro-inflammatory cytokines (interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α)). Subcutaneous injections of the potent DAAO inhibitor CBIO (5-chloro-benzo[D]isoxazol-3-ol) prevented and reversed the chronic morphine-induced hyperalgesia. CBIO also inhibited both astrocyte activation and the expression of pro-inflammatory cytokines. Intrathecal injection of the hydrogen peroxide scavenger PBN (phenyl-N-tert-butylnitrone) and of catalase completely reversed established morphine hyperalgesia, whereas subcutaneous injections of exogenous D-serine failed to alter chronic morphine-induced hyperalgesia. These results provided evidence that spinal DAAO and its subsequent production of hydrogen peroxide rather than the D-serine metabolism contributed to the development of morphine-induced hyperalgesia.

  20. Opioid induced hyperalgesia in anesthetic settings

    PubMed Central

    Lee, Hyeon Jeong

    2014-01-01

    Pain is difficult to investigate and difficult to treat, in part, because of problems in quantification and assessment. The use of opioids, combined with classic anesthetics to maintain hemodynamic stability by controlling responses to intraoperative painful events has gained significant popularity in the anesthetic field. However, several side effects profiles concerning perioperative use of opioid have been published. Over the past two decades, many concerns have arisen with respect to opioid-induced hyperalgesia (OIH), which is the paradoxical effect wherein opioid usage may decrease pain thresholds and increase atypical pain unrelated to the original, preexisting pain. This brief review focuses on the evidence, mechanisms, and modulatory and pharmacologic management of OIH in order to elaborate on the clinical implication of OIH. PMID:25473457

  1. Mast Cell-Mediated Mechanisms of Nociception.

    PubMed

    Aich, Anupam; Afrin, Lawrence B; Gupta, Kalpna

    2015-12-04

    Mast cells are tissue-resident immune cells that release immuno-modulators, chemo-attractants, vasoactive compounds, neuropeptides and growth factors in response to allergens and pathogens constituting a first line of host defense. The neuroimmune interface of immune cells modulating synaptic responses has been of increasing interest, and mast cells have been proposed as key players in orchestrating inflammation-associated pain pathobiology due to their proximity to both vasculature and nerve fibers. Molecular underpinnings of mast cell-mediated pain can be disease-specific. Understanding such mechanisms is critical for developing disease-specific targeted therapeutics to improve analgesic outcomes. We review molecular mechanisms that may contribute to nociception in a disease-specific manner.

  2. Mast Cell-Mediated Mechanisms of Nociception

    PubMed Central

    Aich, Anupam; Afrin, Lawrence B.; Gupta, Kalpna

    2015-01-01

    Mast cells are tissue-resident immune cells that release immuno-modulators, chemo-attractants, vasoactive compounds, neuropeptides and growth factors in response to allergens and pathogens constituting a first line of host defense. The neuroimmune interface of immune cells modulating synaptic responses has been of increasing interest, and mast cells have been proposed as key players in orchestrating inflammation-associated pain pathobiology due to their proximity to both vasculature and nerve fibers. Molecular underpinnings of mast cell-mediated pain can be disease-specific. Understanding such mechanisms is critical for developing disease-specific targeted therapeutics to improve analgesic outcomes. We review molecular mechanisms that may contribute to nociception in a disease-specific manner. PMID:26690128

  3. Inflammation mobilizes local resources to control hyperalgesia: the role of endogenous opioid peptides.

    PubMed

    Alves, Daniela P; da Motta, Patrícia G; Lima, Patrícia P; Queiroz-Junior, Celso M; Caliari, Marcelo V; Pacheco, Daniela F; Pacheco, Cinthia F; Francischi, Janetti N; Duarte, Igor D G

    2012-01-01

    The aim of the present study was to investigate the mechanisms underlying the endogenous control of nociception at a peripheral level during inflammation. Using a pharmacological approach and the rat paw pressure test, we assessed the effect of an intraplantar injection of naloxone, an opioid receptor antagonist, and bestatin, an aminopeptidase inhibitor, on hyperalgesia induced by carrageenan, which mimics an inflammatory process, or prostaglandin E(2) (PGE(2)), which directly sensitizes nociceptors. Naloxone induced a significant and dose-dependent (25, 50 or 100 μg) increase in carrageenan-induced hyperalgesia, but not PGE(2)-induced hyperalgesia. Bestatin (400 μg/paw) significantly counteracted carrageenan-induced hyperalgesia, inducing an increase in the nociceptive threshold compared to control, but it did not modify hyperalgesia induced by PGE(2) injection into the rat paw. Positive β-endorphin immunoreactivity was increased in paw inflammation induced by carrageenan in comparison with the control group. However, PGE(2) did not significantly alter the immunostained area. These results provide evidence for activation of the endogenous opioidergic system during inflammation and indicate that this system regulates hyperalgesia through a negative feedback mechanism, modulating it at a peripheral level.

  4. Supraspinal Gβγ-dependent stimulation of PLCβ3 originating from G inhibitory protein-μ opioid receptor-coupling is necessary for morphine induced acute hyperalgesia

    PubMed Central

    Bianchi, Enrica; Norcini, Monica; Smrcka, Alan; Ghelardini, Carla

    2009-01-01

    Although alterations in μ-opioid receptor signaling mediate excitatory effects of opiates in opioid tolerance, the molecular mechanism for the excitatory effect of acute low dose morphine, as it relates to μ-opioid receptor coupling, is presently unknown. A pronounced coupling of μ-opioid receptor to the α subunit of G inhibitory protein emerged in periaqueductal gray from mice systemically administered with morphine at a dose producing acute thermal hyperalgesia. This coupling was abolished in presence of the selective μ-opioid receptor receptor antagonist CTOP administered at the periaqueductal gray site, showing that the low dose morphine effect is triggered by μ-opioid receptor activated G inhibitory protein at supraspinal level. When Gβγ downstream signalling was blocked by intra-periaqueductal gray co-administration of M119, a compound that inhibits Gβγ dimer-dependent signaling, a complete prevention of low dose morphine induced acute thermal hyperalgesia was obtained. Phospholipase C β3, an enzyme necessary to morphine hyperalgesia, was revealed to be associated with Gβγ in periaqueductal gray. Although opioid administration induces a shift in μ-opioid receptor-G protein coupling from Gi to Gs after chronic administration, our data support that this condition is not realized in acute treatment providing evidence that a separate molecular mechanism underlies morphine induced acute excitatory effect. PMID:19656263

  5. Traumatic Stress Promotes Hyperalgesia via Corticotropin-Releasing Factor-1 Receptor (CRFR1) Signaling in Central Amygdala.

    PubMed

    Itoga, Christy A; Roltsch Hellard, Emily A; Whitaker, Annie M; Lu, Yi-Ling; Schreiber, Allyson L; Baynes, Brittni B; Baiamonte, Brandon A; Richardson, Heather N; Gilpin, Nicholas W

    2016-09-01

    Hyperalgesia is an exaggerated response to noxious stimuli produced by peripheral or central plasticity. Stress modifies nociception, and humans with post-traumatic stress disorder (PTSD) exhibit co-morbid chronic pain and amygdala dysregulation. Predator odor stress produces hyperalgesia in rodents. Systemic blockade of corticotropin-releasing factor (CRF) type 1 receptors (CRFR1s) reduces stress-induced thermal hyperalgesia. We hypothesized that CRF-CRFR1 signaling in central amygdala (CeA) mediates stress-induced hyperalgesia in rats with high stress reactivity. Adult male Wistar rats were exposed to predator odor stress in a conditioned place avoidance paradigm and indexed for high (Avoiders) and low (Non-Avoiders) avoidance of predator odor-paired context, or were unstressed Controls. Rats were tested for the latency to withdraw hindpaws from thermal stimuli (Hargreaves test). We used pharmacological, molecular, and immunohistochemical techniques to assess the role of CRF-CRFR1 signaling in CeA in stress-induced hyperalgesia. Avoiders exhibited higher CRF peptide levels in CeA that did not appear to be locally synthesized. Intra-CeA CRF infusion mimicked stress-induced hyperalgesia. Avoiders exhibited thermal hyperalgesia that was reversed by systemic or intra-CeA injection of a CRFR1 antagonist. Finally, intra-CeA infusion of tetrodotoxin produced thermal hyperalgesia in unstressed rats and blocked the anti-hyperalgesic effect of systemic CRFR1 antagonist in stressed rats. These data suggest that rats with high stress reactivity exhibit hyperalgesia that is mediated by CRF-CRFR1 signaling in CeA.

  6. Molecular mechanisms regulating CD13-mediated adhesion

    PubMed Central

    Ghosh, Mallika; Gerber, Claire; Rahman, M Mamunur; Vernier, Kaitlyn M; Pereira, Flavia E; Subramani, Jaganathan; Caromile, Leslie A; Shapiro, Linda H

    2014-01-01

    CD13/Aminopeptidase N is a transmembrane metalloproteinase that is expressed in many tissues where it regulates various cellular functions. In inflammation, CD13 is expressed on myeloid cells, is up-regulated on endothelial cells at sites of inflammation and mediates monocyte/endothelial adhesion by homotypic interactions. In animal models the lack of CD13 alters the profiles of infiltrating inflammatory cells at sites of ischaemic injury. Here, we found that CD13 expression is enriched specifically on the pro-inflammatory subset of monocytes, suggesting that CD13 may regulate trafficking and function of specific subsets of immune cells. To further dissect the mechanisms regulating CD13-dependent trafficking we used the murine model of thioglycollate-induced sterile peritonitis. Peritoneal monocytes, macrophages and dendritic cells were significantly decreased in inflammatory exudates from global CD13KO animals when compared with wild-type controls. Furthermore, adoptive transfer of wild-type and CD13KO primary myeloid cells, or wild-type myeloid cells pre-treated with CD13-blocking antibodies into thioglycollate-challenged wild-type recipients demonstrated fewer CD13KO or treated cells in the lavage, suggesting that CD13 expression confers a competitive advantage in trafficking. Similarly, both wild-type and CD13KO cells were reduced in infiltrates in CD13KO recipients, confirming that both monocytic and endothelial CD13 contribute to trafficking. Finally, murine monocyte cell lines expressing mouse/human chimeric CD13 molecules demonstrated that the C-terminal domain of the protein mediates CD13 adhesion. Therefore, this work verifies that the altered inflammatory trafficking in CD13KO mice is the result of aberrant myeloid cell subset trafficking and further defines the molecular mechanisms underlying this regulation. PMID:24627994

  7. Histamine-induced itch converts into pain in neuropathic hyperalgesia.

    PubMed

    Baron, R; Schwarz, K; Kleinert, A; Schattschneider, J; Wasner, G

    2001-11-16

    Physiologically, itch and pain are transmitted in separate specific peripheral C-units and central afferent pathways. Some neuropathic pain patients with intact but sensitized (irritable) primary C-nociceptors have spontaneous pain, heat hyperalgesia, static and dynamic mechanical hyperalgesia. The question was whether cutaneous histamine application induces pain in these patients. For comparison histamine was applied into normal skin experimentally sensitized by capsaicin. Histamine application in the capsaicin-induced primary or secondary hyperalgesic skin did not change the intensity and quality of capsaicin pain. Itch was profoundly inhibited. Conversely, histamine application in neuropathic skin induced severe increase in spontaneous burning pain but no itch. In neuropathies irritable nociceptors may express histamine receptors or induce central sensitization to histaminergic stimuli so that itch converts into pain.

  8. Neuroprotective Mechanisms Mediated by CDK5 Inhibition

    PubMed Central

    Mushtaq, Gohar; Greig, Nigel H.; Anwar, Firoz; Al-Abbasi, Fahad A.; Zamzami, Mazin A.; Al-Talhi, Hasan A.; Kamal, Mohammad A.

    2016-01-01

    Cyclin-dependent kinase 5 (CDK5) is a proline-directed serine/threonine kinase belonging to the family of cyclin-dependent kinases. In addition to maintaining the neuronal architecture, CDK5 plays an important role in the regulation of synaptic plasticity, neurotransmitter release, neuron migration and neurite outgrowth. Although various reports have shown links between neurodegeneration and deregulation of cyclin-dependent kinases, the specific role of CDK5 inhibition in causing neuroprotection in cases of neuronal insult or in neurodegenerative diseases is not well-understood. This article discusses current evidence for the involvement of CDK5 deregulation in neurodegenerative disorders and neurodegeneration associated with stroke through various mechanisms. These include upregulation of cyclin D1 and overactivation of CDK5 mediated neuronal cell death pathways, aberrant hyperphosphorylation of human tau proteins and/or neurofilament proteins, formation of neurofibrillary lesions, excitotoxicity, cytoskeletal disruption, motor neuron death (due to abnormally high levels of CDK5/p25) and colchicine-induced apoptosis in cerebellar granule neurons. A better understanding of the role of CDK5 inhibition in neuroprotective mechanisms will help scientists and researchers to develop selective, safe and efficacious pharmacological inhibitors of CDK5 for therapeutic use against human neurodegenerative disorders, such as Alzheimer’s disease, amyotrophic lateral sclerosis and neuronal loss associated with stroke. PMID:26601962

  9. Celecoxib alleviates oxaliplatin-induced hyperalgesia through inhibition of spinal ERK1/2 signaling

    PubMed Central

    Chen, Hongping; Wang, Qinghua; Shi, Danni; Yao, Dongbo; Zhang, Lei; Xiong, Junping; Xu, Baohua

    2016-01-01

    Numerous pieces of evidence have revealed that oxaliplatin (OXA) evokes mechanical and cold hypersensitivity. However, the mechanism underlying these bothersome side effects needs to be further investigated. It is well known that cyclooxygenase-2 (COX-2) and extracellular signal-regulated kinases (ERK1/2) signaling play crucial roles in several pain states. Our previous data showed that Akt2 in the dorsal root ganglion (DRG) participated in the regulation of OXA-induced neuropathic pain. But it is still unclear whether spinal ERK1/2 signaling is involved in the regulation of OXA-induced hyperalgesia, and the linkage between COX-2 and ERK1/2 signaling in mediating OXA-induced hyperalgesia also remains unclear. In this research, we investigated the possible mechanism of celecoxib, a COX-2 inhibitor, in OXA-induced neuropathic pain. Our results show that single dose of OXA (12 mg/kg) significantly attenuated both the tail withdrawal latency (TWL) and mechanical withdrawal threshold (MWT) at days 4 after the OXA treatment. Administration of celecoxib (30 mg/kg/day) for 4 and 6 days inhibited the decrease in TWL and MWT, and each was significantly higher than that of the OXA+vehicle group and was equivalent to that of the vehicles group. OXA increased the expression of cyclooxygenase-2 (COX-2) mRNA and phosphorylated extracellular signal-regulated kinase1/2 (pERK1/2) protein in the lumbar 4-5 (L4-5) spinal cord dorsal horn neurons. Administration of celecoxib for 7 days suppressed the increase in expression of COX-2 and pERK1/2 induced by OXA. Our findings suggested that COX-2 and ERK1/2 signaling in spinal cord contributed to the OXA-induced neuropathic pain. PMID:27821910

  10. Mechanisms of PDGF siRNA-mediated inhibition of bone cancer pain in the spinal cord

    PubMed Central

    Xu, Yang; Liu, Jia; He, Mu; Liu, Ran; Belegu, Visar; Dai, Ping; Liu, Wei; Wang, Wei; Xia, Qing-Jie; Shang, Fei-Fei; Luo, Chao-Zhi; Zhou, Xue; Liu, Su; McDonald, JohnW.; Liu, Jin; Zuo, Yun-Xia; Liu, Fei; Wang, Ting-Hua

    2016-01-01

    Patients with tumors that metastasize to bone frequently suffer from debilitating pain, and effective therapies for treating bone cancer are lacking. This study employed a novel strategy in which herpes simplex virus (HSV) carrying a small interfering RNA (siRNA) targeting platelet-derived growth factor (PDGF) was used to alleviate bone cancer pain. HSV carrying PDGF siRNA was established and intrathecally injected into the cavum subarachnoidale of animals suffering from bone cancer pain and animals in the negative group. Sensory function was assessed by measuring thermal and mechanical hyperalgesia. The mechanism by which PDGF regulates pain was also investigated by comparing the differential expression of pPDGFRα/β and phosphorylated ERK and AKT. Thermal and mechanical hyperalgesia developed in the rats with bone cancer pain, and these effects were accompanied by bone destruction in the tibia. Intrathecal injection of PDGF siRNA and morphine reversed thermal and mechanical hyperalgesia in rats with bone cancer pain. In addition, we observed attenuated astrocyte hypertrophy, down-regulated pPDGFRα/β levels, reduced levels of the neurochemical SP, a reduction in CGRP fibers and changes in pERK/ERK and pAKT/AKT ratios. These results demonstrate that PDGF siRNA can effectively treat pain induced by bone cancer by blocking the AKT-ERK signaling pathway. PMID:27282805

  11. Suckling and sucrose ingestion suppress persistent hyperalgesia and spinal Fos expression after forepaw inflammation in infant rats.

    PubMed

    Ren, K; Blass, E M; Zhou, Q; Dubner, R

    1997-02-18

    Sweet taste and nonnutritive suckling produce analgesia to transient noxious stimuli in infant rats and humans. The present study evaluated the pain-modulating effects of sucrose and suckling in a rat model of persistent pain and hyperalgesia that mimics the response to tissue injury in humans. Fore- and hindpaw withdrawal latencies from a 30 degrees or 48 degrees C brass stylus were determined in 10-day-old rats following paw inflammation induced by complete Freund's adjuvant (CFA; 1:1 injected s.c. in a 0.01 ml volume). CFA markedly decreased escape latencies to both 48 degrees and 30 degrees C stimulation, thereby demonstrating thermal hyperalgesia and mechanical allodynia. The combination of nonnutritive suckling and sucrose (7.5%, 0.01-0.06 ml/min) infusion markedly increased escape latencies to forepaw stimulation in both CFA-treated and control rats. In contrast, intraoral sucrose and suckling did not increase hindpaw withdrawal latencies in either control or CFA-inflamed rats. The effect was specific to sweet taste because neither water nor isotonic saline infusion affected forepaw escape latencies. Parallel findings were obtained for CFA-induced Fos-like immunoreactivity (Fos-LI), a marker of neuronal activation. Fos-LI was selectively induced in cervical and lumbar regions ipsilateral to forepaw and hindpaw inflammation, respectively. Suckling-sucrose treatment significantly reduced Fos-LI at the cervical but not at the lumbar regions. These findings demonstrate: (i) the development of persistent pain and hyperalgesia in 10-day-old rats that can be attenuated by endogenous pain-modulating systems activated by taste and nonnutritive suckling; (ii) the mediation of the sucrose-suckling analgesia and antihyperalgesia at the spinal level; and (iii) a differential rostrocaudal maturation of descending pain-modulating systems to the spinal cord of 10-day-old rats. These findings may provide new clinical approaches for engaging endogenous analgesic mechanisms in

  12. Carbamazepine Withdrawal-induced Hyperalgesia in Chronic Neuropathic Pain.

    PubMed

    Ren, Zhenyu; Yang, Bing; Yang, Bin; Shi, Le; Sun, Qing-Li; Sun, A-Ping; Lu, Lin; Liu, Xiaoguang; Zhao, Rongsheng; Zhai, Suodi

    2015-11-01

    Combined pharmacological treatments are the most used approach for neuropathic pain. Carbamazepine, an antiepileptic agent, is generally used as a third-line treatment for neuropathic pain and can be considered an option only when patients have not responded to the first- and second-line medications. In the case presented herein, a patient with neuropathic pain was treated using a combined pharmacological regimen. The patient's pain deteriorated, despite increasing the doses of opioids, when carbamazepine was discontinued, potentially because carbamazepine withdrawal disrupted the balance that was achieved by the multifaceted pharmacological regimen, thus inducing hyperalgesia. Interestingly, when carbamazepine was prescribed again, the patient's pain was successfully managed. Animal research has reported that carbamazepine can potentiate the analgesic effectiveness of morphine in rodent models of neuropathic pain and postoperative pain. This clinical case demonstrates that carbamazepine may have a synergistic effect on the analgesic effectiveness of morphine and may inhibit or postpone opioid-induced hyperalgesia. We postulate that a probable mechanism of action of carbamazepine may involve -aminobutyric acid-ergic potentiation and the interruption of glutamatergic function via N-methyl-D-aspartate receptors. Further research is warranted to clarify the analgesic action of carbamazepine and its potential use for the prevention of opioid-induced hyperalgesia in chronic neuropathic pain patients.

  13. Spinal administration of a delta opioid receptor agonist attenuates hyperalgesia and allodynia in a rat model of neuropathic pain.

    PubMed

    Holdridge, Sarah V; Cahill, Catherine M

    2007-08-01

    Neuropathic (NP) pain is a debilitating chronic pain disorder considered by some to be inherently resistant to therapy with traditional analgesics. Indeed, micro opioid receptor (OR) agonists show reduced therapeutic benefit and their long term use is hindered by the high incidence of adverse effects. However, pharmacological and physiological evidence increasingly suggests a role for deltaOR agonists in modulating NP pain symptoms. In this study, we examined the antihyperalgesic and antiallodynic effects of the spinally administered deltaOR agonist, d-[Ala(2), Glu(4)]deltorphin II (deltorphin II), as well as the changes in deltaOR expression, in rats following chronic constriction injury (CCI) of the sciatic nerve. Rats with CCI exhibited cold hyperalgesia and mechanical allodynia over a 14-day testing period. Intrathecal administration of deltorphin II reversed cold hyperalgesia on day 14 and dose-dependently attenuated mechanical allodynia. The effects of deltorphin II were mediated via activation of the deltaOR as the effect was antagonized by co-treatment with the delta-selective antagonist, naltrindole. Western blotting experiments revealed no changes in deltaOR protein in the dorsal spinal cord following CCI. Taken together, these data demonstrate the antihyperalgesic and antiallodynic effectiveness of a spinally administered deltaOR agonist following peripheral nerve injury and support further investigation of deltaORs as potential therapeutic targets in the treatment of NP pain.

  14. Effects of intravenous Injections Paederiae and Stauntonia on spontaneous pain, hyperalgesia and inflammation induced by cutaneous chemical tissue injury in the rat.

    PubMed

    Peng, Xiao-Li; Gao, Xi-Ling; Chen, Jun; Huang, Xi; Chen, Hui-Sheng

    2003-10-25

    To study whether commercial traditional Chinese medicinal preparations Injection Paederiae (IP) or Injection Stauntonia (IS) has anti-nociceptive and/or anti-inflammatory effects, we used two persistent pain models (bee venom and formalin test) to evaluate the systemic effects of IP or IS on the chemical tissue injury-induced persistent spontaneous pain-related responses (PSPR), primary thermal/mechanical hyperalgesia and inflammation in conscious rats. Injection of bee venom (BV, 0.1 mg, 50 microl) into the plantar surface of one hind paw resulted in not only a 1-h monophasic PSPR such as flinching reflex in the injected paw and a subsequent period of 3-4 days primary heat and mechanical hyperalgesia, but also a marked sign of inflammation, including redness and swelling of the plantar surface in the injected paw. Intraplantar injection of formalin produced two phases of PSPR as reported previously. Systemic pre-treatment with three doses of IP (0.32, 1.6 and 9.0 ml/kg, 500%) or IS (0.32, 1.6 and 9.0 ml/kg, 250%) produced a dose-dependent suppression of the BV- or formalin-induced flinching reflex of 1 h time course as compared with the saline control group. Post-treatment with IP or IS 5 min after BV injection also produced a significant suppression of the flinching reflex in both BV test and formalin test respectively, as compared with the control group. However, neither pre- nor post-treatment with IP or IS produced any significantly suppressive effect on the BV-induced primary heat and mechanical hyperalgesia and inflammation. The analgesia produced by IP or IS was not mediated by the endogenous opioid receptors since naloxone, a non-selective opioid receptor antagonist, had no reversal effect on the IP and IS-produced analgesia in the BV-induced PSPR. Our present results suggest that IP or IS might prevent and relieve clinical persistent spontaneous pain, but without any anti-nociceptive and anti-inflammatory effects on the primary heat hyperalgesia

  15. Role of Corticotropin-releasing Factor Signaling in Stress-related Alterations of Colonic Motility and Hyperalgesia

    PubMed Central

    Taché, Yvette; Million, Mulugeta

    2015-01-01

    The corticotropin-releasing factor (CRF) signaling systems encompass CRF and the structurally related peptide urocortin (Ucn) 1, 2, and 3 along with 2 G-protein coupled receptors, CRF1 and CRF2. CRF binds with high and moderate affinity to CRF1 and CRF2 receptors, respectively while Ucn1 is a high-affinity agonist at both receptors, and Ucn2 and Ucn3 are selective CRF2 agonists. The CRF systems are expressed in both the brain and the colon at the gene and protein levels. Experimental studies established that the activation of CRF1 pathway in the brain or the colon recaptures cardinal features of diarrhea predominant irritable bowel syndrome (IBS) (stimulation of colonic motility, activation of mast cells and serotonin, defecation/watery diarrhea, and visceral hyperalgesia). Conversely, selective CRF1 antagonists or CRF1/CRF2 antagonists, abolished or reduced exogenous CRF and stress-induced stimulation of colonic motility, defecation, diarrhea and colonic mast cell activation and visceral hyperalgesia to colorectal distention. By contrast, the CRF2 signaling in the colon dampened the CRF1 mediated stimulation of colonic motor function and visceral hyperalgesia. These data provide a conceptual framework that sustained activation of the CRF1 system at central and/or peripheral sites may be one of the underlying basis of IBS-diarrhea symptoms. While targeting these mechanisms by CRF1 antagonists provided a relevant novel therapeutic venue, so far these promising preclinical data have not translated into therapeutic use of CRF1 antagonists. Whether the existing or newly developed CRF1 antagonists will progress to therapeutic benefits for stress-sensitive diseases including IBS for a subset of patients is still a work in progress. PMID:25611064

  16. Central activation of TRPV1 and TRPA1 by novel endogenous agonists contributes to mechanical allodynia and thermal hyperalgesia after burn injury.

    PubMed

    Green, Dustin; Ruparel, Shivani; Gao, Xiaoli; Ruparel, Nikita; Patil, Mayur; Akopian, Armen; Hargreaves, Kenneth

    2016-01-01

    The primary complaint of burn victims is an intense, often devastating spontaneous pain, with persistence of mechanical and thermal allodynia. The transient receptor potential channels, TRPV1 and TRPA1, are expressed by a subset of nociceptive sensory neurons and contribute to inflammatory hypersensitivity. Although their function in the periphery is well known, a role for these TRP channels in central pain mechanisms is less well defined. Lipid agonists of TRPV1 are released from peripheral tissues via enzymatic oxidation after burn injury; however, it is not known if burn injury triggers the release of oxidized lipids in the spinal cord. Accordingly, we evaluated whether burn injury evoked the central release of oxidized lipids . Analysis of lipid extracts of spinal cord tissue with HPLC-MS revealed a significant increase in levels of the epoxide and diol metabolites of linoleic acid: 9,10-DiHOME, 12,13-DiHOME, 9(10)-EpOME, and 12(13)-EpOME, that was reduced after intrathecal (i.t.) injection of the oxidative enzyme inhibitor ketoconazole. Moreover, we found that these four lipid metabolites were capable of specifically activating both TRPV1 and TRPA1. Intrathecal injection of specific antagonists to TRPV1 (AMG-517) or TRPA1 (HC-030031) significantly reduced post-burn mechanical and thermal allodynia. Finally, i.t. injection of ketoconazole significantly reversed post-burn mechanical and thermal allodynia. Our data indicate that spinal cord TRPV1 and TRPA1 contributes to pain after burn and identifies a novel class of oxidized lipids elevated in the spinal cord after burn injury. Since the management of burn pain is problematic, these findings point to a novel approach for treating post-burn pain.

  17. Elite swimmers with and without unilateral shoulder pain: mechanical hyperalgesia and active/latent muscle trigger points in neck-shoulder muscles.

    PubMed

    Hidalgo-Lozano, A; Fernández-de-las-Peñas, C; Calderón-Soto, C; Domingo-Camara, A; Madeleine, P; Arroyo-Morales, M

    2013-02-01

    Our aim was to investigate the presence of mechanical hypersensitivity and active trigger points (TrPs) in the neck-shoulder muscles in elite swimmers with/without unilateral shoulder pain. Seventeen elite swimmers with shoulder pain; 18 swimmers without shoulder pain; and 15 elite athletes matched controls were recruited. Pressure pain thresholds (PPT) were assessed over the levator scapulae, sternocleidomastoid, upper trapezius, infraspinatus, scalene, subscapularis and tibialis anterior muscles. TrPs in the levator scapulae, upper trapezius, infraspinatus, scalene, sternocleidomastoid and subscapularis muscles were also explored. Swimmers with shoulder pain showed significant lower PPT in all muscles compared with controls (P<0.01). No differences in PPT were found between swimmers with and without shoulder pain, underlining widespread mechanical hypersensitivity. The mean number of TrPs for elite swimmer with and without shoulder pain was, respectively, 4.7 ± 1 (2.1 ± 1.5 active; 2.6 ± 1.4 latent) and 4.7 ± 1.3 (1.3 ± 1.3 active; 3.4 ± 1.5 latent), whereas healthy athletes only showed latent TrPs (2.4 ± 1.2). Elite swimmers with shoulder pain showed higher number of active TrPs than swimmers without pain, whereas it was the opposite for the number of latent muscle TrP (P<0.05). The reported mechanical hypersensitivity suggests that active TrPs play a role in the development of shoulder pain in elite swimmers.

  18. TRPA1 contributes to capsaicin-induced facial cold hyperalgesia in rats.

    PubMed

    Honda, Kuniya; Shinoda, Masamichi; Furukawa, Akihiko; Kita, Kozue; Noma, Noboru; Iwata, Koichi

    2014-12-01

    Orofacial cold hyperalgesia is known to cause severe persistent pain in the face following trigeminal nerve injury or inflammation, and transient receptor potential (TRP) vanilloid 1 (TRPV1) and TRP ankylin 1 (TRPA1) are thought to be involved in cold hyperalgesia. However, how these two receptors are involved in cold hyperalgesia is not fully understood. To clarify the mechanisms underlying facial cold hyperalgesia, nocifensive behaviors to cold stimulation, the expression of TRPV1 and TRPA1 in trigeminal ganglion (TG) neurons, and TG neuronal excitability to cold stimulation following facial capsaicin injection were examined in rats. The head-withdrawal reflex threshold (HWRT) to cold stimulation of the lateral facial skin was significantly decreased following facial capsaicin injection. This reduction of HWRT was significantly recovered following local injection of TRPV1 antagonist as well as TRPA1 antagonist. Approximately 30% of TG neurons innervating the lateral facial skin expressed both TRPV1 and TRPA1, and about 64% of TRPA1-positive neurons also expressed TRPV1. The TG neuronal excitability to noxious cold stimulation was significantly increased following facial capsaicin injection and this increase was recovered by pretreatment with TRPA1 antagonist. These findings suggest that TRPA1 sensitization via TRPV1 signaling in TG neurons is involved in cold hyperalgesia following facial skin capsaicin injection.

  19. Rifaximin Alters Intestinal Bacteria and Prevents Stress-Induced Gut Inflammation and Visceral Hyperalgesia in Rats

    PubMed Central

    Xu, Dabo; Gao, Jun; Gillilland, Merritt; Wu, Xiaoyin; Song, Il; Kao, John Y.; Owyang, Chung

    2014-01-01

    Background & Aims Rifaximin is used to treat patients with functional gastrointestinal disorders, but little is known about its therapeutic mechanism. We propose that rifaximin modulates the ileal bacterial community, reduces subclinical inflammation of the intestinal mucosa, and improves gut barrier function to reduce visceral hypersensitivity. Methods We induced visceral hyperalgesia in rats, via chronic water avoidance or repeat restraint stressors, and investigated whether rifaximin altered the gut microbiota, prevented intestinal inflammation, and improved gut barrier function. Quantitative polymerase chain reaction and 454 pyrosequencing were used to analyze bacterial 16S rRNA in ileal contents from the rats. Reverse transcription, immunoblot, and histologic analyses were used to evaluate levels of cytokines, the tight junction protein occludin, and mucosal inflammation, respectively. Intestinal permeability and rectal sensitivity were measured. Results Water avoidance and repeat restraint stress each led to visceral hyperalgesia, accompanied by mucosal inflammation and impaired mucosal barrier function. Oral rifaximin altered the composition of bacterial communities in the ileum (Lactobacillus species became the most abundant) and prevented mucosal inflammation, impairment to intestinal barrier function, and visceral hyperalgesia in response to chronic stress. Neomycin also changed the composition of the ileal bacterial community (Proteobacteria became the most abundant species). Neomycin did not prevent intestinal inflammation or induction of visceral hyperalgesia induced by water avoidance stress. Conclusions Rifaximin alters the bacterial population in the ileum of rats, leading to a relative abundance of Lactobacillus. These changes prevent intestinal abnormalities and visceral hyperalgesia in response to chronic psychological stress. PMID:24161699

  20. Nerve growth factor induced hyperalgesia in the rat hind paw is dependent on circulating neutrophils.

    PubMed

    Bennett, G; al-Rashed, S; Hoult, J R; Brain, S D

    1998-09-01

    The mechanisms by which nerve growth factor (NGF) induces thermal hyperalgesia and neutrophil accumulation have been investigated in the rat. Thermal nociceptive thresholds in rat hind paw were measured as the time taken for paw withdrawal from a heat source and neutrophil accumulation was measured in hind paw and dorsal skin samples using a myeloperoxidase assay. NGF (23-80 pmol intraplantar (i.pl.) injection) induced a significant (P < 0.05, n = 6-16) thermal hyperalgesia at 5 h after injection and significant neutrophil accumulation (P < 0.05, n = 6) was observed with NGF (40 pmol). In dorsal skin, where multiple samples can be assessed, intradermal (i.d.) NGF was 10-30 times less potent than interleukin-1beta in inducing neutrophil accumulation. The 5-lipoxygenase inhibitor ZM230487 (10 nmol co-injected with NGF) significantly attenuated neutrophil accumulation and hyperalgesia induced by NGF; unlike the histamine and 5-hydroxytryptamine antagonists (mepyramine and methysergide) which were without effect at the times measured. Furthermore, depletion of circulating neutrophils (using a rabbit anti-rat neutrophil antibody) abolished NGF induced hyperalgesia. These results indicate that neutrophils, which accumulate in response to a 5-lipoxygenase product, play a crucial role in NGF-induced hyperalgesia.

  1. Secondary Hyperalgesia Phenotypes Exhibit Differences in Brain Activation during Noxious Stimulation

    PubMed Central

    Werner, Mads Utke; Mårtensson, Johan; Larsson, Henrik B. W.; Dahl, Jørgen Berg

    2015-01-01

    Noxious stimulation of the skin with either chemical, electrical or heat stimuli leads to the development of primary hyperalgesia at the site of injury, and to secondary hyperalgesia in normal skin surrounding the injury. Secondary hyperalgesia is inducible in most individuals and is attributed to central neuronal sensitization. Some individuals develop large areas of secondary hyperalgesia (high-sensitization responders), while others develop small areas (low-sensitization responders). The magnitude of each area is reproducible within individuals, and can be regarded as a phenotypic characteristic. To study differences in the propensity to develop central sensitization we examined differences in brain activity and anatomy according to individual phenotypical expression of secondary hyperalgesia by magnetic resonance imaging. Forty healthy volunteers received a first-degree burn-injury (47°C, 7 min, 9 cm2) on the non-dominant lower-leg. Areas of secondary hyperalgesia were assessed 100 min after the injury. We measured neuronal activation by recording blood-oxygen-level-dependent-signals (BOLD-signals) during mechanical noxious stimulation before burn injury and in both primary and secondary hyperalgesia areas after burn-injury. In addition, T1-weighted images were used to measure differences in gray-matter density in cortical and subcortical regions of the brain. We found significant differences in neuronal activity between high- and low-sensitization responders at baseline (before application of the burn-injury) (p < 0.05). After the burn-injury, we found significant differences between responders during noxious stimulation of both primary (p < 0.01) and secondary hyperalgesia (p ≤ 0.04) skin areas. A decreased volume of the right (p = 0.001) and left caudate nucleus (p = 0.01) was detected in high-sensitization responders in comparison to low-sensitization responders. These findings suggest that brain-structure and neuronal activation to noxious stimulation

  2. Secondary hyperalgesia phenotypes exhibit differences in brain activation during noxious stimulation.

    PubMed

    Asghar, Mohammad Sohail; Pereira, Manuel Pedro; Werner, Mads Utke; Mårtensson, Johan; Larsson, Henrik B W; Dahl, Jørgen Berg

    2015-01-01

    Noxious stimulation of the skin with either chemical, electrical or heat stimuli leads to the development of primary hyperalgesia at the site of injury, and to secondary hyperalgesia in normal skin surrounding the injury. Secondary hyperalgesia is inducible in most individuals and is attributed to central neuronal sensitization. Some individuals develop large areas of secondary hyperalgesia (high-sensitization responders), while others develop small areas (low-sensitization responders). The magnitude of each area is reproducible within individuals, and can be regarded as a phenotypic characteristic. To study differences in the propensity to develop central sensitization we examined differences in brain activity and anatomy according to individual phenotypical expression of secondary hyperalgesia by magnetic resonance imaging. Forty healthy volunteers received a first-degree burn-injury (47 °C, 7 min, 9 cm(2)) on the non-dominant lower-leg. Areas of secondary hyperalgesia were assessed 100 min after the injury. We measured neuronal activation by recording blood-oxygen-level-dependent-signals (BOLD-signals) during mechanical noxious stimulation before burn injury and in both primary and secondary hyperalgesia areas after burn-injury. In addition, T1-weighted images were used to measure differences in gray-matter density in cortical and subcortical regions of the brain. We found significant differences in neuronal activity between high- and low-sensitization responders at baseline (before application of the burn-injury) (p < 0.05). After the burn-injury, we found significant differences between responders during noxious stimulation of both primary (p < 0.01) and secondary hyperalgesia (p ≤ 0.04) skin areas. A decreased volume of the right (p = 0.001) and left caudate nucleus (p = 0.01) was detected in high-sensitization responders in comparison to low-sensitization responders. These findings suggest that brain-structure and neuronal activation to noxious

  3. Hyperalgesia and functional sensory loss in restless legs syndrome.

    PubMed

    Stiasny-Kolster, Karin; Pfau, Doreen B; Oertel, Wolfgang H; Treede, Rolf-Detlef; Magerl, Walter

    2013-08-01

    Pain and other sensory signs in patients with restless legs syndrome (RLS) are still poorly understood, as most investigations focus on motor system dysfunctions. This study aimed to investigate somatosensory changes in patients with primary RLS and the restoration of somatosensory function by guideline-based treatment. Forty previously untreated RLS patients were investigated unilaterally over hand and foot using quantitative sensory testing (QST) and were compared with 40 age- and gender-matched healthy subjects. The predominant finding in RLS patients was 3- to 4-fold increase of sensitivity to pinprick stimuli in both extremities (hand: P<.05; foot: P<.001), a sensory pathway involved in withdrawal reflexes. Pinprick hyperalgesia was not paralleled by dynamic mechanical allodynia. Additional significant sensory changes were tactile hypoesthesia in both extremities (hand: P<.05; foot P<.01) and dysesthesia to non-noxious cold stimuli (paradoxical heat sensation), which was present in the foot in an unusually high proportion (14 of 40 patients; P<.01). In 8 patients, follow-up QST 2 to 20 months after treatment with l-DOPA (L-3,4-dihydroxyphenylalanine) revealed a significant reduction of pinprick hyperalgesia (-60%, P<.001), improved tactile detection (+50%, P<.05), and disappearance of paradoxical heat sensation in half of the patients. QST suggested a type of spinal or supraspinal central sensitization differing from neuropathic pain or human experimental models of central sensitization by the absence of dynamic mechanical allodynia. Reversal of pinprick hyperalgesia by l-DOPA may be explained by impaired descending inhibitory dopaminergic control on spinal nociceptive neurons. Restoration of tactile sensitivity and paradoxical heat sensations suggest that they were functional disturbances resulting from central disinhibition.

  4. Molecular Mechanism of Cyclodextrin Mediated Cholesterol Extraction

    PubMed Central

    López, Cesar A.; de Vries, Alex H.; Marrink, Siewert J.

    2011-01-01

    The depletion of cholesterol from membranes, mediated by β-cyclodextrin (β-CD) is well known and documented, but the molecular details of this process are largely unknown. Using molecular dynamics simulations, we have been able to study the CD mediated extraction of cholesterol from model membranes, in particular from a pure cholesterol monolayer, at atomic resolution. Our results show that efficient cholesterol extraction depends on the structural distribution of the CDs on the surface of the monolayer. With a suitably oriented dimer, cholesterol is extracted spontaneously on a nanosecond time scale. Additional free energy calculations reveal that the CDs have a strong affinity to bind to the membrane surface, and, by doing so, destabilize the local packing of cholesterol molecules making their extraction favorable. Our results have implications for the interpretation of experimental measurements, and may help in the rational design of efficient CD based nano-carriers. PMID:21455285

  5. Pioglitazone inhibits the development of hyperalgesia and sensitization of spinal nociresponsive neurons in type 2 diabetes

    PubMed Central

    Griggs, Ryan B.; Donahue, Renee R.; Adkins, Braxton G.; Anderson, Katie L.; Thibault, Olivier; Taylor, Bradley K.

    2016-01-01

    Thiazolidinedione drugs (TZDs) such as pioglitazone are FDA-approved for the treatment of insulin resistance in type 2 diabetes. However, whether TZDs reduce painful diabetic neuropathy (PDN) remains unknown. Therefore we tested the hypothesis that chronic administration of pioglitazone would reduce PDN in Zucker Diabetic Fatty (ZDFfa/fa) rats. Compared to Zucker Lean (ZLfa/+) controls, ZDF developed: (1) elevated blood glucose, HbA1c, methylglyoxal and insulin; (2) mechanical and thermal hyperalgesia at the hindpaw; (3) increased avoidance of noxious mechanical probes in a mechanical conflict avoidance behavioral assay, the first report of a measure of affective-motivational pain-like behavior in ZDF; and (4) exaggerated lumbar dorsal horn immunohistochemical expression of pressure-evoked phosphorylated extracellular signal-regulated kinase (pERK). Seven weeks of pioglitazone (30 mg · kg−1 · d−1 in food) reduced blood glucose, HbA1c, hyperalgesia, and pERK expression in ZDF. This is the first report to reveal hyperalgesia and spinal sensitization in the same ZDF animals, both evoked by a noxious mechanical stimulus that reflects pressure pain frequently associated with clinical PDN. As pioglitazone provides the combined benefit of reducing hyperglycemia, hyperalgesia, and central sensitization, we suggest that TZDs represent an attractive pharmacotherapy in patients with type 2 diabetes-associated pain. PMID:26687453

  6. Role for Peroxynitrite in Sphingosine-1-Phosphate Induced Hyperalgesia in Rats

    PubMed Central

    Doyle, Tim; Finley, Amanda; Chen, Zhoumou; Salvemini, Daniela

    2011-01-01

    Sphingosine-1-phosphate (S1P) is an important mediator of inflammation recently shown in in vitro studies to increase the excitability of small diameter sensory neurons at least in part via activation of the S1P1 receptor subtype. Activation of S1PR1 has been reported to increase the formation of NADPH oxidase-derived superoxide (O2•−) and nitric oxide synthase (NOS)-derived nitric oxide (NO). This process favors the formation of peroxynitrite (ONOO−, PN), a potent mediator of hyperalgesia associated with peripheral and central sensitization. The aims of our study were to determine whether S1P causes peripheral sensitization and thermal hyperalgesia via S1PR1 activation and PN formation. Intraplantar injection of S1P in rats led to a time-dependent development of thermal hyperalgesia that was blocked by the S1PR1 antagonist, W146 but not its inactive enantiomer, W140. The hyperalgesic effects of S1P were mimicked by intraplantar injection of the well characterized S1PR1 agonist, SEW2871. The development of S1P-induced hyperalgesia was blocked by apocynin, a NADPH oxidase inhibitor, L-NAME, a non-selective NOS inhibitor and by the potent PN decomposition catalysts (FeTM-4-PyP5+ and MnTE-2-PyP5+). Our findings provide mechanistic insight into the signaling pathways engaged by S1P in the development of hyperalgesia and highlight the contribution of the S1P1 receptor-to-PN signaling in this process. PMID:21239112

  7. Effect of subcutaneous administration of calcium channel blockers on nerve injury-induced hyperalgesia.

    PubMed

    White, D M; Cousins, M J

    1998-08-10

    Recent studies suggest that calcium contributes to peripheral neural mechanisms of hyperalgesia associated with nerve damage. In this animal behavioural study, we examined further the contribution of calcium in neuropathic pain by testing whether subcutaneous administration of either a calcium chelating agent or voltage-dependent calcium channel blockers attenuate nerve injury-induced hyperalgesia to mechanical stimulation. Studies were carried out in animals with partially ligated sciatic nerves, an established animal model of neuropathic pain. The nociceptive flexion reflex was quantified using an Ugo Basile Analgesymeter. Partial nerve injury induced a significant decrease in mechanical threshold compared to the sham operated controls. Daily subcutaneous injections of the calcium chelating agent, Quin 2 (20 microgram/2.5 microliter), significantly attenuated the nerve injury-induced hyperalgesia. Similarly, SNX-111, a N-type channel blocker, also significantly attenuated the nerve injury-induced hyperalgesia. SNX-230, a P and/or Q-type channel blocker, and nifedipine, a L-type channel blocker, had no effect on the hyperalgesia to mechanical stimulation. In control experiments, SNX-111 had no effect on mechanical thresholds when administered subcutaneously in either the hindpaw of normal animals or the back of the neck in nerve injury animals. This study shows that neuropathic pain involves a local calcium-dependent mechanism in the receptive field of intact neurons of an injured nerve, since it can be alleviated by subcutaneous injections of either a calcium chelating agent or SNX-111, a N-type calcium channel blocker. These agents may be effective, peripherally acting therapeutic agents for neuropathic pain.

  8. Transient receptor potential ankyrin-1 has a major role in mediating visceral pain in mice.

    PubMed

    Cattaruzza, Fiore; Spreadbury, Ian; Miranda-Morales, Marcela; Grady, Eileen F; Vanner, Stephen; Bunnett, Nigel W

    2010-01-01

    The excitatory ion channel transient receptor potential ankyrin-1 (TRPA1) is prominently expressed by primary afferent neurons and is a mediator of inflammatory pain. Inflammatory agents can directly activate [e.g., hydroxynonenal (HNE), prostaglandin metabolites] or indirectly sensitize [e.g., agonists of protease-activated receptor (PAR(2))] TRPA1 to induce somatic pain and hyperalgesia. However, the contribution of TRPA1 to visceral pain is unknown. We investigated the role of TRPA1 in visceral hyperalgesia by measuring abdominal visceromotor responses (VMR) to colorectal distention (CRD) after intracolonic administration of TRPA1 agonists [mustard oil (MO), HNE], sensitizing agents [PAR(2) activating peptide (PAR(2)-AP)], and the inflammatory agent trinitrobenzene sulfonic acid (TNBS) in trpa1(+/+) and trpa1(-/-) mice. Sensory neurons innervating the colon, identified by retrograde tracing, coexpressed immunoreactive TRPA1, calcitonin gene-related peptide, and substance P, expressed TRPA1 mRNA and responded to MO with depolarizing currents. Intracolonic MO and HNE increased VMR to CRD and induced immunoreactive c-fos in spinal neurons in trpa1+/+ but not in trpa1(-/-) mice. Intracolonic PAR(2)-AP induced mechanical hyperalgesia in trpa1+/+ but not in trpa1(-/-) mice. TNBS-induced colitis increased in VMR to CRD and induced c-fos in spinal neurons in trpa1(+/+) but not in trpa1(-/-) mice. Thus TRPA1 is expressed by colonic primary afferent neurons. Direct activation of TRPA1 causes visceral hyperalgesia, and TRPA1 mediates PAR(2)-induced hyperalgesia. TRPA1 deletion markedly reduces colitis-induced mechanical hyperalgesia in the colon. Our results suggest that TRPA1 has a major role in visceral nociception and may be a therapeutic target for colonic inflammatory pain.

  9. Lateral Hypothalamic Stimulation Reduces Hyperalgesia Through Spinally Descending Orexin-A Neurons in Neuropathic Pain.

    PubMed

    Wardach, Jacob; Wagner, Monica; Jeong, Younhee; Holden, Janean E

    2016-03-01

    No evidence to date shows that lateral hypothalamic (LH) stimulation produces orexin-A-mediated antinociception in the spinal cord dorsal horn (SCDH) in a model of neuropathic pain. We conducted experiments to examine the effect of orexin-A-mediated LH stimulation in female rats with chronic constriction injury (CCI) on thermal hyperalgesia. Rats receiving carbachol into the LH demonstrated antinociception on both the left CCI and right nonligated paws (p < .05). Rats were given carbachol in the LH followed by intrathecal injection of the orexin-1 (OX1) receptor antagonist SB-334867, which blocked LH-induced antinociception compared with control groups (p < .05) in the left paw, but not in the right paw. These findings support the hypothesis that LH stimulation produces antinociception in rats with thermal hyperalgesia from neuropathic pain via an orexin-A connection between the LH and the SCDH. Identification of this pathway may lead to studies using orexins to manage clinical pain.

  10. Mechanisms of cytoskeleton-mediated mechanical signal transmission in cells

    PubMed Central

    Hwang, Yongyun; Gouget, Cecile L.M.; Barakat, Abdul I.

    2012-01-01

    Recent experiments have demonstrated very rapid long-distance transmission of mechanical forces within cells. Because the speed of this transmission greatly exceeds that of reaction-diffusion signaling, it has been conjectured that it occurs via the propagation of elastic waves through the actin stress fiber network. To explore the plausibility of this conjecture, we recently developed a model of small amplitude stress fiber deformations in prestressed viscoelastic stress fibers subjected to external forces. The model results demonstrated that rapid mechanical signal transmission is only possible when the external force is applied orthogonal to the stress fiber axis and that the dynamics of this transmission are governed by a balance between the prestress in the stress fiber and the stress fiber's material viscosity. The present study, which is a follow-up on our previous model, uses dimensional analysis to: (1) further evaluate the plausibility of the elastic wave conjecture and (2) obtain insight into mechanical signal transmission dynamics in simple stress fiber networks. We show that the elastic wave scenario is likely not the mechanism of rapid mechanical signal transmission in actin stress fibers due to the highly viscoelastic character of these fibers. Our analysis also demonstrates that the time constant characterizing mechanical stimulus transmission is strongly dependent on the topology of the stress fiber network, implying that network organization plays an important role in determining the dynamics of cellular responsiveness to mechanical stimulation. PMID:23336020

  11. Mechanisms of cytoskeleton-mediated mechanical signal transmission in cells.

    PubMed

    Hwang, Yongyun; Gouget, Cecile L M; Barakat, Abdul I

    2012-11-01

    Recent experiments have demonstrated very rapid long-distance transmission of mechanical forces within cells. Because the speed of this transmission greatly exceeds that of reaction-diffusion signaling, it has been conjectured that it occurs via the propagation of elastic waves through the actin stress fiber network. To explore the plausibility of this conjecture, we recently developed a model of small amplitude stress fiber deformations in prestressed viscoelastic stress fibers subjected to external forces. The model results demonstrated that rapid mechanical signal transmission is only possible when the external force is applied orthogonal to the stress fiber axis and that the dynamics of this transmission are governed by a balance between the prestress in the stress fiber and the stress fiber's material viscosity. The present study, which is a follow-up on our previous model, uses dimensional analysis to: (1) further evaluate the plausibility of the elastic wave conjecture and (2) obtain insight into mechanical signal transmission dynamics in simple stress fiber networks. We show that the elastic wave scenario is likely not the mechanism of rapid mechanical signal transmission in actin stress fibers due to the highly viscoelastic character of these fibers. Our analysis also demonstrates that the time constant characterizing mechanical stimulus transmission is strongly dependent on the topology of the stress fiber network, implying that network organization plays an important role in determining the dynamics of cellular responsiveness to mechanical stimulation.

  12. Mechanisms involved in antibody- and complement-mediated allograft rejection

    PubMed Central

    2010-01-01

    Antibody-mediated rejection has become critical clinically because this form of rejection is usually unresponsive to conventional anti-rejection therapy, and therefore, it has been recognized as a major cause of allograft loss. Our group developed experimental animal models of vascularized organ transplantation to study pathogenesis of antibody- and complement-mediated endothelial cell injury leading to graft rejection. In this review, we discuss mechanisms of antibody-mediated graft rejection resulting from activation of complement by C1q- and MBL (mannose-binding lectin)-dependent pathways and interactions with a variety of effector cells, including macrophages and monocytes through Fcγ receptors and complement receptors. PMID:20135240

  13. Phospholipase C and protein kinase A mediate bradykinin sensitization of TRPA1: a molecular mechanism of inflammatory pain.

    PubMed

    Wang, Shenglan; Dai, Yi; Fukuoka, Tetsuo; Yamanaka, Hiroki; Kobayashi, Kimiko; Obata, Koichi; Cui, Xiuyu; Tominaga, Makoto; Noguchi, Koichi

    2008-05-01

    Bradykinin is an inflammatory mediator that plays a pivotal role in pain and hyperalgesia in inflamed tissues by exciting and/or sensitizing nociceptors. TRPA1 is an important component of the transduction machinery through which environmental irritants and endogenous proalgesic agents depolarize nociceptors to elicit inflammatory pain. Here, using electrophysiological, immunocytochemical and behavioural analyses, we showed a functional interaction of these two inflammation-related molecules in both heterologous expressing systems and primary sensory neurons. We found that bradykinin increased the TRPA1 currents evoked by allyl isothiocyanate (AITC) or cinnamaldehyde in HEK293 cells expressing TRPA1 and bradykinin receptor 2 (B2R). This potentiation was inhibited by phospholipase C (PLC) inhibitor or protein kinase A (PKA) inhibitor, and mimicked by PLC or PKA activator. The functional interaction between B2R and TRPA1, as well as the modulation mechanism, was also observed in rat dorsal root ganglia neurons. In an occlusion experiment, the PLC activator could enhance AITC-induced TRPA1 current further even in saturated PKA-mediated potentiation, indicating the additive potentiating effects of the PLC and PKA pathways. These data for the first time indicate that a cAMP-PKA signalling is involved in the downstream from B2R in dorsal root ganglia neurons in addition to PLC. Finally, subcutaneous pre-injection of a sub-inflammatory dose of bradykinin into rat hind paw enhanced AITC-induced pain behaviours, which was consistent with the observations in vitro. Collectively, these results represent a novel mechanism through which bradykinin released in response to tissue inflammation might trigger the sensation of pain by TRPA1 activation.

  14. Immunological Mechanisms Mediating Hantavirus Persistence in Rodent Reservoirs

    PubMed Central

    Easterbrook, Judith D.; Klein, Sabra L.

    2008-01-01

    Hantaviruses, similar to several emerging zoonotic viruses, persistently infect their natural reservoir hosts, without causing overt signs of disease. Spillover to incidental human hosts results in morbidity and mortality mediated by excessive proinflammatory and cellular immune responses. The mechanisms mediating the persistence of hantaviruses and the absence of clinical symptoms in rodent reservoirs are only starting to be uncovered. Recent studies indicate that during hantavirus infection, proinflammatory and antiviral responses are reduced and regulatory responses are elevated at sites of increased virus replication in rodents. The recent discovery of structural and non-structural proteins that suppress type I interferon responses in humans suggests that immune responses in rodent hosts could be mediated directly by the virus. Alternatively, several host factors, including sex steroids, glucocorticoids, and genetic factors, are reported to alter host susceptibility and may contribute to persistence of hantaviruses in rodents. Humans and reservoir hosts differ in infection outcomes and in immune responses to hantavirus infection; thus, understanding the mechanisms mediating viral persistence and the absence of disease in rodents may provide insight into the prevention and treatment of disease in humans. Consideration of the coevolutionary mechanisms mediating hantaviral persistence and rodent host survival is providing insight into the mechanisms by which zoonotic viruses have remained in the environment for millions of years and continue to be transmitted to humans. PMID:19043585

  15. EVALUATION OF HYPERALGESIA AND HISTOLOGICAL CHANGES OF DORSAL ROOT GANGLION INDUCED BY NUCLEUS PULPOSUS

    PubMed Central

    Grava, André Luiz de Souza; Ferrari, Luiz Fernando; Parada, Carlos Amílcar; Defino, Helton Luiz Aparecido

    2015-01-01

    To evaluate the hyperalgesia and histological abnormalities induced by contact between the dorsal root ganglion and the nucleus pulposus. Methods: Twenty Wistar rats were used, divided into two experimental groups. In one of the groups, a fragment of autologous nucleus pulposus was removed from the sacrococcygeal region and deposited on the L5 dorsal root ganglia. In the other group (control), a fragment of adipose tissue was deposited on the L5 dorsal root ganglia. Mechanical and thermal hyperalgesia was evaluated on the third day and the first, third, fifth and seventh weeks after the operation. A L5 dorsal root ganglion was removed in the first, third, fifth and seventh weeks after the operation for histological study using HE staining and histochemical study using specific labeling for iNOS. Results: Higher intensity of mechanical and thermal hyperalgesia was observed in the group of animals in which the nucleus pulposus was placed in contact with the dorsal root ganglion. In this group, the histological study showed abnormalities of the dorsal root ganglion tissue, characterized by an inflammatory process and axonal degeneration. The histopathological abnormalities of the dorsal root ganglion tissue presented increasing intensity with increasing length of observation, and there was a correlation with maintenance of the hyperalgesia observed in the behavioral assessment. Immunohistochemistry using specific labeling for iNOS in the group of animals in which the nucleus pulposus was placed in contact with the dorsal root ganglion showed higher expression of this enzyme in the nuclei of the inflammatory cells (glial cells) surrounding the neurons. Conclusion: Contact between the nucleus pulposus and the dorsal root ganglion induced mechanical and thermal hyperalgesia and caused histological abnormalities in the dorsal root ganglion components. These abnormalities were characterized by an inflammatory and degenerative process in the structures of the dorsal root

  16. Protective effects of dexmedetomidine combined with flurbiprofen axetil on remifentanil-induced hyperalgesia: A randomized controlled trial

    PubMed Central

    Yu, Zenggui; Wu, Weilan; Wu, Xiaodan; Lei, Hongyi; Gong, Cansheng; Xu, Shiyuan

    2016-01-01

    High dosages of intra-operative remifentanil are associated with opioid-induced hyperalgesia (OIH). The aim of the present study was to investigate the effect of combined dexmedetomidine and flurbiprofen axetil treatment on remifentanil-induced hyperalgesia. Patients with an American Society of Anesthesiologists physical status of I–II who were diagnosed with hysteromyoma and scheduled for laparoscopic assisted vaginal hysterectomy (LAVH) were randomly divided into three groups. Group hyperalgesia (Group H, n=29) received intra-operative remifentanil, Group hyperalgesia and dexmedetomidine (Group HD, n=28) received remifentanil and a continuous infusion of dexmedetomidine, and Group hyperalgesia, dexmedetomidine and flurbiprofen axetil (Group HDF, n=29) received remifentanil, flurbiprofen axetil and dexmedetomidine. Mechanical pain thresholds were measured during the preoperative visit and postoperatively at 1, 6 and 24-h time points. Visual analog scale (VAS) scores, time to analgesic requirement, total sufentanil consumption and side effects were assessed postoperatively. Mechanical pain threshold at the incision site was significantly lower in Group H compared with Groups HD and HDF (both P<0.05), and significantly higher in Group HDF than in Group HD (P<0.05). The area of secondary hyperalgesia at the incision site was greater in Group H than in the other two groups (both P<0.05), and significantly smaller in Group HDF compared with Group HD (P<0.05). VAS scores and total sufentanil consumption were significantly higher in Group H compared with the other two groups (both P<0.05), and were significantly lower in Group HDF compared with Group HD (P<0.05). Dexmedetomidine combined with flurbiprofen axetil exhibits synergetic effects in the prevention of remifentanil-induced hyperalgesia in patients undergoing LAVH. PMID:27698764

  17. Distinct Neural Mechanisms Mediate Olfactory Memory Formation at Different Timescales

    ERIC Educational Resources Information Center

    McNamara, Ann Marie; Magidson, Phillip D.; Linster, Christiane; Wilson, Donald A.; Cleland, Thomas A.

    2008-01-01

    Habituation is one of the oldest forms of learning, broadly expressed across sensory systems and taxa. Here, we demonstrate that olfactory habituation induced at different timescales (comprising different odor exposure and intertrial interval durations) is mediated by different neural mechanisms. First, the persistence of habituation memory is…

  18. Metformin attenuates hyperalgesia and allodynia in rats with painful diabetic neuropathy induced by streptozotocin.

    PubMed

    Ma, Junxiong; Yu, Hailong; Liu, Jun; Chen, Yu; Wang, Qi; Xiang, Liangbi

    2015-10-05

    Painful diabetic neuropathy is a common complication of diabetes mellitus, which often makes the patients suffer from severe hyperalgesia and allodynia. Thus far, the treatment of painful diabetic neuropathy remains unsatisfactory. Metformin, which is the first-line drug for type-2 diabetes, has been proved to attenuate hyperexcitability in sensory neurons linked to chemotherapy-induced neuropathic pain, highlighting its potential in alleviating pain related with painful diabetic neuropathy. The present study was designed to investigate the potential beneficial effect of metformin on hyperalgesia and allodynia in diabetic rats. The mechanical sensitivity, heat nociception, and cold allodynia were examined. The levels of malondialdehyde, superoxide dismutase, and advanced glycation end-products in the blood were measured. The expression of adenosine monophosphate-activated protein kinase (AMPK) phosphorylation and AMPK target genes were examined in the sciatic nerves of the animals. It was found that metformin was capable of attenuating diabetes-induced mechanical hyperalgesia, heat hyperalgesia and cold allodynia. In addition, metformin was capable of decreasing malondialdehyde and glycation end-products levels in blood, as well as increasing superoxide dismutas activity, indicating the inhibitory effect of metformin against diabetes-induced oxidative stress. Further studies showed that metformin could activate AMPK and increase the AMPK target genes in sciatic nerves in diabetic rats. In conclusion, metformin is able to attenuate diabetes-induced hyperalgesia and allodynia, which might be associated its anti-oxidative effect through AMPK pathway. Metformin might be used as an effective drug, especially with fewer side effects, for abnormal sensation in painful diabetic neuropathy.

  19. Methylglyoxal modification of Nav1.8 facilitates nociceptive neuron firing and causes hyperalgesia in diabetic neuropathy.

    PubMed

    Bierhaus, Angelika; Fleming, Thomas; Stoyanov, Stoyan; Leffler, Andreas; Babes, Alexandru; Neacsu, Cristian; Sauer, Susanne K; Eberhardt, Mirjam; Schnölzer, Martina; Lasitschka, Felix; Lasischka, Felix; Neuhuber, Winfried L; Kichko, Tatjana I; Konrade, Ilze; Elvert, Ralf; Mier, Walter; Pirags, Valdis; Lukic, Ivan K; Morcos, Michael; Dehmer, Thomas; Rabbani, Naila; Thornalley, Paul J; Edelstein, Diane; Nau, Carla; Forbes, Josephine; Humpert, Per M; Schwaninger, Markus; Ziegler, Dan; Stern, David M; Cooper, Mark E; Haberkorn, Uwe; Brownlee, Michael; Reeh, Peter W; Nawroth, Peter P

    2012-06-01

    This study establishes a mechanism for metabolic hyperalgesia based on the glycolytic metabolite methylglyoxal. We found that concentrations of plasma methylglyoxal above 600 nM discriminate between diabetes-affected individuals with pain and those without pain. Methylglyoxal depolarizes sensory neurons and induces post-translational modifications of the voltage-gated sodium channel Na(v)1.8, which are associated with increased electrical excitability and facilitated firing of nociceptive neurons, whereas it promotes the slow inactivation of Na(v)1.7. In mice, treatment with methylglyoxal reduces nerve conduction velocity, facilitates neurosecretion of calcitonin gene-related peptide, increases cyclooxygenase-2 (COX-2) expression and evokes thermal and mechanical hyperalgesia. This hyperalgesia is reflected by increased blood flow in brain regions that are involved in pain processing. We also found similar changes in streptozotocin-induced and genetic mouse models of diabetes but not in Na(v)1.8 knockout (Scn10(-/-)) mice. Several strategies that include a methylglyoxal scavenger are effective in reducing methylglyoxal- and diabetes-induced hyperalgesia. This previously undescribed concept of metabolically driven hyperalgesia provides a new basis for the design of therapeutic interventions for painful diabetic neuropathy.

  20. Homocysteine-induced attenuation of vascular endothelium-dependent hyperalgesia in the rat

    PubMed Central

    Joseph, Elizabeth K.; Green, Paul G.; Ferrari, Luiz F.; Levine, Jon D.

    2014-01-01

    We have recently demonstrated a role of the vascular endothelium in peripheral pain mechanism by disrupting endothelial cell function using intravascular administration of octoxynol-9, a non-selective membrane active agent. As an independent test of the role of endothelial cells in pain mechanisms, we evaluated the effect of homocysteine, an agent that damages endothelial cell function. Mechanical stimulus-induced enhancement of endothelin-1 hyperalgesia in the gastrocnemius muscle of the rat was first prevented then enhanced by intravenous administration of homocysteine, but was only inhibited by its precursor, methionine. Both homocysteine and methionine significantly attenuated mechanical hyperalgesia in two models of ergonomic muscle pain, induced by exposure to vibration, and by eccentric exercise, and cutaneous mechanical hyperalgesia in an ischemia-reperfusion injury model of Complex Regional Pain Syndrome type I, all previously shown responsive to octoxynol-9. This study provides independent support for a role of the endothelial cell in pain syndromes thought to have a vascular basis, and suggests that substances that are endothelial cell toxins can enhance vascular pain. PMID:25451284

  1. Chronic CRF1 receptor blockade reduces heroin intake escalation and dependence-induced hyperalgesia.

    PubMed

    Park, Paula E; Schlosburg, Joel E; Vendruscolo, Leandro F; Schulteis, Gery; Edwards, Scott; Koob, George F

    2015-03-01

    Opioids represent effective drugs for the relief of pain, yet chronic opioid use often leads to a state of increased sensitivity to pain that is exacerbated during withdrawal. A sensitization of pain-related negative affect has been hypothesized to closely interact with addiction mechanisms. Neuro-adaptive changes occur as a consequence of excessive opioid exposure, including a recruitment of corticotropin-releasing factor (CRF) and norepinephrine (NE) brain stress systems. To better understand the mechanisms underlying the transition to dependence, we determined the effects of functional antagonism within these two systems on hyperalgesia-like behavior during heroin withdrawal utilizing models of both acute and chronic dependence. We found that passive or self-administered heroin produced a significant mechanical hypersensitivity. During acute opioid dependence, systemic administration of the CRF1 receptor antagonist MPZP (20 mg/kg) alleviated withdrawal-induced mechanical hypersensitivity. In contrast, several functional adrenergic system antagonists (clonidine, prazosin, propranolol) failed to alter mechanical hypersensitivity in this state. We then determined the effects of chronic MPZP or clonidine treatment on extended access heroin self-administration and found that MPZP, but not clonidine, attenuated escalation of heroin intake, whereas both drugs alleviated chronic dependence-associated hyperalgesia. These findings suggest that an early potentiation of CRF signaling occurs following opioid exposure that begins to drive both opioid-induced hyperalgesia and eventually intake escalation.

  2. Polymeric mechanical amplifiers of immune cytokine-mediated apoptosis

    NASA Astrophysics Data System (ADS)

    Mitchell, Michael J.; Webster, Jamie; Chung, Amanda; Guimarães, Pedro P. G.; Khan, Omar F.; Langer, Robert

    2017-03-01

    Physical forces affect tumour growth, progression and metastasis. Here, we develop polymeric mechanical amplifiers that exploit in vitro and in vivo physical forces to increase immune cytokine-mediated tumour cell apoptosis. Mechanical amplifiers, consisting of biodegradable polymeric particles tethered to the tumour cell surface via polyethylene glycol linkers, increase the apoptotic effect of an immune cytokine on tumour cells under fluid shear exposure by as much as 50% compared with treatment under static conditions. We show that targeted polymeric particles delivered to tumour cells in vivo amplify the apoptotic effect of a subsequent treatment of immune cytokine, reduce circulating tumour cells in blood and overall tumour cell burden by over 90% and reduce solid tumour growth in combination with the antioxidant resveratrol. The work introduces a potentially new application for a broad range of micro- and nanoparticles to maximize receptor-mediated signalling and function in the presence of physical forces.

  3. Polymeric mechanical amplifiers of immune cytokine-mediated apoptosis

    PubMed Central

    Mitchell, Michael J.; Webster, Jamie; Chung, Amanda; Guimarães, Pedro P. G.; Khan, Omar F.; Langer, Robert

    2017-01-01

    Physical forces affect tumour growth, progression and metastasis. Here, we develop polymeric mechanical amplifiers that exploit in vitro and in vivo physical forces to increase immune cytokine-mediated tumour cell apoptosis. Mechanical amplifiers, consisting of biodegradable polymeric particles tethered to the tumour cell surface via polyethylene glycol linkers, increase the apoptotic effect of an immune cytokine on tumour cells under fluid shear exposure by as much as 50% compared with treatment under static conditions. We show that targeted polymeric particles delivered to tumour cells in vivo amplify the apoptotic effect of a subsequent treatment of immune cytokine, reduce circulating tumour cells in blood and overall tumour cell burden by over 90% and reduce solid tumour growth in combination with the antioxidant resveratrol. The work introduces a potentially new application for a broad range of micro- and nanoparticles to maximize receptor-mediated signalling and function in the presence of physical forces. PMID:28317839

  4. Novel Lipid Mediators and Resolution Mechanisms in Acute Inflammation

    PubMed Central

    Serhan, Charles N.

    2010-01-01

    Because inflammation is appreciated as a unifying basis of many widely occurring diseases, the mechanisms involved in its natural resolution are of considerable interest. Using contained, self-limited inflammatory exudates and a systems approach, novel lipid-derived mediators and pathways were uncovered in the resolution of inflammatory exudates. These new families of local mediators control both the duration and magnitude of acute inflammation as well as the return of the site to homeostasis in the process of catabasis. This new genus of specialized proresolving mediators (SPM) includes essential fatty acid–derived lipoxins, resolvins, protectins, and, most recently, maresins. These families were named based on their unique structures and potent stereoselective actions. The temporally initiated biosynthesis of SPM and their direct impact on leukocyte trafficking and macrophage-directed clearance mechanisms provide clear evidence that resolution is an active, programmed response at the tissue level. Moreover, SPM that possess anti-inflammatory (ie, limiting PMN infiltration) and proresolving (enhance macrophage uptake and clearance of apoptotic PMN and microbial particles) actions as well as stimulating mucosal antimicrobial responses demonstrate that anti-inflammation and proresolution are different responses of the host and novel defining properties of these molecules. The mapping of new resolution circuits has opened the possibility for understanding mechanisms that lead from acute to chronic inflammation, or to the resolution thereof, as well as to potential, resolution-based immunopharmacological therapies. PMID:20813960

  5. Attenuation of thermal nociception and hyperalgesia by VR1 blockers

    PubMed Central

    García-Martínez, Carolina; Humet, Marc; Planells-Cases, Rosa; Gomis, Ana; Caprini, Marco; Viana, Felix; De la Peña, Elvira; Sanchez-Baeza, Francisco; Carbonell, Teresa; De Felipe, Carmen; Pérez-Payá, Enrique; Belmonte, Carlos; Messeguer, Angel; Ferrer-Montiel, Antonio

    2002-01-01

    Vanilloid receptor subunit 1 (VR1) appears to play a critical role in the transduction of noxious chemical and thermal stimuli by sensory nerve endings in peripheral tissues. Thus, VR1 antagonists are useful compounds to unravel the contribution of this receptor to pain perception, as well as to induce analgesia. We have used a combinatorial approach to identify new, nonpeptidic channel blockers of VR1. Screening of a library of trimers of N-alkylglycines resulted in the identification of two molecules referred to as DD161515 {N-[2-(2-(N-methylpyrrolidinyl)ethyl]glycyl]-[N-[2,4-dichlorophenethyl]glycyl]-N-(2,4-dichlorophenethyl)glycinamide} and DD191515 {[N-[3-(N,N-diethylamino)propyl]glycyl]-[N-[2,4-dichlorophenethyl]glycyl]-N-(2,4-dichlorophenethyl)glycinamide} that selectively block VR1 channel activity with micromolar efficacy, rivaling that characteristic of vanilloid-related inhibitors. These compounds appear to be noncompetitive VR1 antagonists that recognize a receptor site distinct from that of capsaicin. Intraperitoneal administration of both trialkylglycines into mice significantly attenuated thermal nociception as measured in the hot plate test. It is noteworthy that these compounds eliminated pain and neurogenic inflammation evoked by intradermal injection of capsaicin into the animal hindpaw, as well as the thermal hyperalgesia induced by tissue irritation with nitrogen mustard. In contrast, responses to mechanical stimuli were not modified by either compound. Modulation of sensory nerve fibers excitability appears to underlie the peptoid analgesic activity. Collectively, these results indicate that blockade of VR1 activity attenuates chemical and thermal nociception and hyperalgesia, supporting the tenet that this ionotropic receptor contributes to chemical and thermal sensitivity and pain perception in vivo. These trialkylglycine-based, noncompetitive VR1 antagonists may likely be developed into analgesics to treat inflammatory pain. PMID:11854530

  6. Quinolone resistance in bacteria: emphasis on plasmid-mediated mechanisms.

    PubMed

    Li, Xian-Zhi

    2005-06-01

    Bacterial resistance to quinolones/fluoroquinolones has emerged rapidly and such resistance has traditionally been attributed to the chromosomally mediated mechanisms that alter the quinolone targets (i.e. DNA gyrase and topoisomerase IV) and/or overproduce multidrug resistance efflux pumps. However, the discovery of the plasmid-borne quinolone resistance determinant, named qnr, has substantially broadened our horizon on the molecular mechanisms of quinolone resistance. Several recent reports of Qnr or its homologues encoded by transferable plasmids in Gram-negative bacteria isolated worldwide highlight the significance of the emerging plasmid-mediated mechanism(s). This also alerts us to the potential rapid dissemination of quinolone resistance determinants. Qnr belongs to the pentapeptide repeat family and protects DNA gyrase from the action of quinolone agents including the newer fluoroquinolones. This protection interplays with chromosomal mechanisms to raise significantly the resistance levels. The qnr-bearing strains generate quinolone-resistant mutants at a much higher frequency than those qnr-free strains. Furthermore, the qnr-plasmids are integron-associated and carry multiple resistance determinants providing resistance to several classes of antimicrobials including beta-lactams and aminoglycosides. The high quinolone resistance rates in Escherichia coli are used to address issues of quinolone resistance, and possible strategies for minimising quinolone resistance are discussed.

  7. Mitotic wavefronts mediated by mechanical signaling in early Drosophila embryos

    NASA Astrophysics Data System (ADS)

    Kang, Louis; Idema, Timon; Liu, Andrea; Lubensky, Tom

    2013-03-01

    Mitosis in the early Drosophila embryo demonstrates spatial and temporal correlations in the form of wavefronts that travel across the embryo in each cell cycle. This coordinated phenomenon requires a signaling mechanism, which we suggest is mechanical in origin. We have constructed a theoretical model that supports nonlinear wavefront propagation in a mechanically-excitable medium. Previously, we have shown that this model captures quantitatively the wavefront speed as it varies with cell cycle number, for reasonable values of the elastic moduli and damping coefficient of the medium. Now we show that our model also captures the displacements of cell nuclei in the embryo in response to the traveling wavefront. This new result further supports that mechanical signaling may play an important role in mediating mitotic wavefronts.

  8. Activation of JNK pathway in spinal astrocytes contributes to acute ultra-low-dose morphine thermal hyperalgesia.

    PubMed

    Sanna, Maria Domenica; Ghelardini, Carla; Galeotti, Nicoletta

    2015-07-01

    Accumulating evidence suggests that opioid analgesics can lead to paradoxical sensitization to pain when delivered in different administration patterns. Although opioid tolerance-induced hyperalgesia is largely studied, little is known about the mechanisms underlying acute ultra-low-dose morphine hyperalgesia. Activation of spinal glial cells is reported to regulate pain hypersensitivity. To elucidate the mechanism involved in acute ultra-low-dose morphine hyperalgesia, we tested whether an opioid agonist promoted the activation of spinal astrocytes and microglia and investigated the cellular pathways involved. Ultra-low-dose morphine activated spinal astrocytes with no effect on microglia. The astrocyte activation was selectively prevented by the opioid antagonist naloxone, the μ-opioid receptor (MOR) silencing and the JNK inhibitor SP600125. Morphine elevated spinal JNK1, JNK2, and c-Jun phosphorylation. Conversely, phosphorylation of cAMP response element-binding protein (CREB) and signal transducer and activator of transcription-1 (STAT-1) was not elevated, and nuclear factor kappa B (NF-κB) levels remained unmodified. Administration of SP600125 and the N-methyl-D-aspartate (NMDA) antagonist MK801 prevented morphine hyperalgesia. Ultra-low-dose morphine increased protein kinase C (PKC) γ phosphorylation. Pretreatment with a PKC inhibitor prevented morphine hyperalgesia and JNK and c-Jun overphosphorylation, indicating PKC is a JNK upstream modulator and illustrating the presence of a pathway involving PKC, NMDA, and JNK activated by morphine. Immunofluorescence experiments indicated the neuronal localization of spinal MOR. However, JNK was not detected in MOR-expressing cells, showing the presence of a neuron-astrocyte signaling pathway. These results illustrate the selective activation of an astrocyte JNK pathway after the stimulation of neuronal MOR, which contributes to ultra-low-dose morphine hyperalgesia.

  9. Pathogenesis and mechanisms of antibody-mediated hemolysis

    PubMed Central

    Flegel, Willy A

    2015-01-01

    Background The clinical consequences of antibodies to red blood cells (RBC) have been studied for a century. Most clinically relevant antibodies can be detected by sensitive in vitro assays. Several mechanisms of antibody-mediated hemolysis are well understood. Such hemolysis following transfusion is reliably avoided in a donor/recipient pair, if one individual is negative for the cognate antigen to which the other has the antibody. Study design and results Mechanisms of antibody-mediated hemolysis were reviewed based on a presentation at the Strategies to Address Hemolytic Complications of Immune Globulin Infusions Workshop addressing intravenous immunoglobulin (IVIG) and ABO antibodies. The presented topics included the rates of intravascular and extravascular hemolysis; IgM and IgG isoagglutinins; auto- and alloantibodies; antibody specificity; A, B, A,B and A1 antigens; A1 versus A2 phenotypes; monocytes/macrophages, other immune cells and complement; monocyte monolayer assay (MMA); antibody-dependent cell-mediated cytotoxicity (ADCC); and transfusion reactions due to ABO and other antibodies. Conclusion Several clinically relevant questions remained unresolved, and diagnostic tools were lacking to routinely and reliably predict the clinical consequences of RBC antibodies. Most hemolytic transfusion reactions associated with IVIG were due to ABO antibodies. Reducing the titers of such antibodies in IVIG may lower the frequency of this kind of adverse event. The only way to stop these events is to have no anti-A or anti-B antibodies in the IVIG products. PMID:26174897

  10. Mechanisms of strain-mediated mesenchymal stem cell apoptosis.

    PubMed

    Kearney, E M; Prendergast, P J; Campbell, V A

    2008-12-01

    Mechanical conditioning of mesenchymal stem cells (MSCs) has been adopted widely as a biophysical signal to aid tissue engineering applications. The replication of in vivo mechanical signaling has been used in in vitro environments to regulate cell differentiation, and extracellular matrix synthesis, so that both the chemical and mechanical properties of the tissue-engineered construct are compatible with the implant site. While research in these areas contributes to tissue engineering, the effects of mechanical strain on MSC apoptosis remain poorly defined. To evaluate the effects of uniaxial cyclic tensile strain on MSC apoptosis and to investigate mechanotransduction associated with strain-mediated cell death, MSCs seeded on a 2D silicone membrane were stimulated by a range of strain magnitudes for 3 days. Mechanotransduction was investigated using the stretch-activated cation channel blocker gadolinium chloride, the L-type voltage-activated calcium channel blocker nicardipine, the c-jun NH(2)-terminal kinase (JNK) blocker D-JNK inhibitor 1, and the calpain inhibitor MDL 28170. Apoptosis was assessed through DNA fragmentation using the terminal deoxynucleotidyl transferase mediated-UTP-end nick labeling method. Results demonstrated that tensile strains of 7.5% or greater induce apoptosis in MSCs. L-type voltage-activated calcium channels coupled mechanical stress to activation of calpain and JNK, which lead to apoptosis through DNA fragmentation. The definition of the in vitro boundary conditions for tensile strain and MSCs along with a proposed mechanism for apoptosis induced by mechanical events positively contributes to the development of MSC biology, bioreactor design for tissue engineering, and development of computational methods for mechanobiology.

  11. Oral administration of stavudine induces hyperalgesia without affecting activity in rats.

    PubMed

    Weber, Juliane; Mitchell, Duncan; Kamerman, Peter R

    2007-12-05

    We have investigated whether long-term oral administration of the nucleoside reverse transcriptase inhibitor (NRTI) stavudine affects nociception in Sprague-Dawley rats, and whether any changes of nociception are accompanied by deterioration in activity and appetite. Stavudine (50 mg kg(-1)) was administered to rats orally once daily for six weeks in gelatine cubes. Mechanical hyperalgesia of the tail was assessed using a bar algometer, and thermal hyperalgesia by tail immersion in 49 degrees C water. Withdrawal latencies were compared to those of rats receiving placebo gelatine cubes. Withdrawal latencies to the noxious thermal challenge were not affected by stavudine, but those to the mechanical challenge were significantly decreased in rats receiving stavudine, compared to rats receiving placebo, from week three to week six of drug administration (P<0.05, ANCOVA with Newman Keuls post-hoc comparisons). The overall condition of the rats was assessed by recording daily voluntary wheel running distance and maximum running speed, food intake and body mass. Daily stavudine administration did not adversely affect voluntary running activity, appetite or growth. We have shown that long-term daily oral administration of the NRTI stavudine results in mechanical hyperalgesia in rats within three weeks without affecting appetite, growth and physical activity.

  12. Movement-evoked hyperalgesia induced by lipopolysaccharides is not suppressed by glucocorticoids

    PubMed Central

    Kovács, Katalin J.; Papic, Jonathan C.; Larson, Alice A.

    2008-01-01

    Systemic exposure to lipopolysaccharides (LPS) produces a variety of effects, including movement-evoked hyperalgesia that can be measured using the grip force assay in mice. Because both lethality and enhanced sensitivity to cutaneous pain following exposure to endotoxins have each been attributed to inflammatory mediators, we explored the possibility that LPS-induced movement-evoked hyperalgesia is also sensitive to manipulations of glucocorticoids that regulate these other LPS responses. We found that the hyperalgesic effect of LPS (5 mg/kg s.c.) in mice that were adrenalectomized did not differ from that in control mice that were sham-operated, even though mortality after LPS was potentiated by adrenalectomy. The development of tolerance to the movement-evoked hyperalgesic effect of LPS also did not differ between adrenalectomized and sham-operated control mice. In addition, mifepristone (25 mg/kg s.c.), a glucocorticoid antagonist, did not attenuate the hyperalgesic effect of LPS (2 mg/kg s.c.), yet this dose of mifepristone was sufficient to enhance the incidence of lethality induced by LPS. Enhancement of glucocorticoid activity by two injections of dexamethasone (1 mg/kg s.c.) had no effect on the degree of hyperalgesia in mice injected with LPS (5 mg/kg s.c.), yet this dose of dexamethasone was sufficient to attenuate the incidence of mortality induced by LPS in adrenalectomized mice. Finally, morphine (10 mg/kg i.p.) reversed the decrease in grip force caused by LPS (5 mg/kg i.p.), supporting the interpretation that decreases in grip force produced by LPS reflect muscle hyperalgesia that is not sensitive to glucocorticoids. PMID:17686584

  13. Chronic infection by Leishmania amazonensis mediated through MAPK ERK mechanisms.

    PubMed

    Martinez, Pedro A; Petersen, Christine A

    2014-08-01

    Leishmania amazonensis is an intracellular protozoan parasite responsible for chronic cutaneous leishmaniasis (CL). CL is a neglected tropical disease responsible for infecting millions of people worldwide. L. amazonensis promotes alteration of various signaling pathways that are essential for host cell survival. Specifically, through parasite-mediated phosphorylation of extracellular signal regulated kinase (ERK), L. amazonensis inhibits cell-mediated parasite killing and promotes its own survival by co-opting multiple host cell functions. In this review, we highlight Leishmania-host cell signaling alterations focusing on those specific to (1) motor proteins, (2) prevention of NADPH subunit phosphorylation impairing reactive oxygen species production, and (3) localized endosomal signaling to up-regulate ERK phosphorylation. This review will focus upon mechanisms and possible explanations as to how Leishmania spp. evades the various layers of defense employed by the host immune response.

  14. Intact subepidermal nerve fibers mediate mechanical hypersensitivity via the activation of protein kinase C gamma in spared nerve injury

    PubMed Central

    Ko, Miau-Hwa; Yang, Ming-Ling; Youn, Su-Chung; Tseng, To-Jung

    2016-01-01

    Background Spared nerve injury is an important neuropathic pain model for investigating the role of intact primary afferents in the skin on pain hypersensitivity. However, potential cellular mechanisms remain poorly understood. In phosphoinositide-3 kinase pathway, pyruvate dehydrogenase kinase 1 (PDK1) participates in the regulation of neuronal plasticity for central sensitization. The downstream cascades of PDK1 include: (1) protein kinase C gamma (PKCγ) controls the trafficking and phosphorylation of ionotropic glutamate receptor; (2) protein kinase B (Akt)/the mammalian target of rapamycin (mTOR) signaling is responsible for local protein synthesis. Under these statements, we therefore hypothesized that an increase of PKCγ activation and mTOR-dependent PKCγ synthesis in intact primary afferents after SNI might contribute to pain hypersensitivity. Results The variants of spared nerve injury were performed in Sprague-Dawley rats by transecting any two of the three branches of the sciatic nerve, leaving only one branch intact. Following SNIt (spared tibial branch), mechanical hyperalgesia and mechanical allodynia, but not thermal hyperalgesia, were significantly induced. In the first footpad, normal epidermal innervations were verified by the protein gene product 9.5 (PGP9.5)- and growth-associated protein 43 (GAP43)-immunoreactive (IR) intraepidermal nerve fibers (IENFs) densities. Furthermore, the rapid increases of phospho-PKCγ- and phospho-mTOR-IR subepidermal nerve fibers (SENFs) areas were distinct gathered from the results of PGP9.5-, GAP43-, and neurofilament 200 (NF200)-IR SENFs areas. The efficacy of PKC inhibitor (GF 109203X) or mTOR complex 1 inhibitor (rapamycin) for attenuating mechanical hyperalgesia and mechanical allodynia by intraplantar injection was dose-dependent. Conclusions From results obtained in this study, we strongly recommend that the intact SENFs persistently increase PKCγ activation and mTOR-dependent PKCγ synthesis participate

  15. Opioid-induced hyperalgesia in chronic pain patients and the mitigating effects of gabapentin

    PubMed Central

    Stoicea, Nicoleta; Russell, Daric; Weidner, Greg; Durda, Michael; Joseph, Nicholas C.; Yu, Jeffrey; Bergese, Sergio D.

    2015-01-01

    Chronic pain patients receiving opioid drugs are at risk for opioid-induced hyperalgesia (OIH), wherein opioid pain medication leads to a paradoxical pain state. OIH involves central sensitization of primary and secondary afferent neurons in the dorsal horn and dorsal root ganglion, similar to neuropathic pain. Gabapentin, a gamma-aminobutyric acid (GABA) analog anticonvulsant used to treat neuropathic pain, has been shown in animal models to reduce fentanyl hyperalgesia without compromising analgesic effect. Chronic pain patients have also exhibited lower opioid consumption and improved pain response when given gabapentin. However, few human studies investigating gabapentin use in OIH have been performed in recent years. In this review, we discuss the potential mechanisms that underlie OIH and provide a critical overview of interventional therapeutic strategies, especially the clinically-successful drug gabapentin, which may reduce OIH. PMID:26074817

  16. Tyrosine phosphorylation of the N-Methyl-D-Aspartate receptor 2B subunit in spinal cord contributes to remifentanil-induced postoperative hyperalgesia: the preventive effect of ketamine

    PubMed Central

    2009-01-01

    Background Experimental and clinical studies showed that intraoperative infusionof remifentanil has been associated with postoperative hyperalgesia. Previous reports suggested that spinal N-methyl-D-aspartate (NMDA) receptors may contribute to the development and maintenance of opioid-induced hyperalgesia. In the present study, we used a rat model of postoperative pain to investigate the role of tyrosine phosphorylation of NMDA receptor 2B (NR2B) subunit in spinal cord in the postoperative hyperalgesia induced by remifentanil and the intervention of pretreatment with ketamine. Results Intraoperative infusion of remifentanil (0.04 mg/kg, subcutaneous) significantly enhanced mechanical allodynia and thermal hyperalgesia induced by the plantar incision during the postoperative period (each lasting between 2 h and 48 h), which was attenuated by pretreatment with ketamine (10 mg/kg, subcutaneous). Correlated with the pain behavior changes, immunocytochemical and western blotting experiments in our study revealed that there was a marked increase in NR2B phosphorylation at Tyr1472 in the superficial dorsal horn after intraoperative infusion of remifentanil, which was attenuated by pretreatment with ketamine. Conclusions This study provides direct evidence that tyrosine phosphorylation of the NR2B at Tyr1472 in spinal dosal horn contributes to postoperative hyperalgesia induced by remifentanil and supports the potential therapeutic value of ketamine for improving postoperative hyperalgesia induced by remifentanil. PMID:20042082

  17. Intraneural dexamethasone applied simultaneously to rat sciatic nerve constriction delays the development of hyperalgesia and allodynia.

    PubMed

    Bastos, Leandro F S; Medeiros, Daniel C; Vieira, Rafael P; Watkins, Linda R; Coelho, Márcio M; Moraes, Márcio F D

    2012-02-21

    Although neuroimmune interactions associated with the development of pain sensitization in models of neuropathic pain have been widely studied, there are some aspects that require further investigation. Thus, we aimed to evaluate whether the local intraneural or perineural injections of dexamethasone, an efficacious anti-inflammatory and immunosuppressant drug, delays the development of both thermal hyperalgesia and mechanical allodynia in an experimental model of neuropathic pain in rats. Hargreaves and electronic von Frey tests were applied. The chronic constriction injury (CCI) of right sciatic nerve was performed. Single intraneural dexamethasone administration at the moment of constriction delayed the development of sensitization for thermal hyperalgesia and mechanical allodynia. However, perineural administration of dexamethasone, at the highest dose, did not delay experimental pain development. These results show that inflammation/immune response at the site of nerve lesion is an essential trigger for the pathological changes that lead to both hyperalgesia and allodynia. In conclusion, this approach opens new opportunities to study cellular and molecular neuroimmune interactions associated with the development of pain derived from peripheral neuropathies.

  18. Mechanisms of PDL1-mediated regulation of autoimmune diabetes.

    PubMed

    Guleria, Indira; Gubbels Bupp, Melanie; Dada, Shirine; Fife, Brian; Tang, Qizhi; Ansari, Mohammed Javeed; Trikudanathan, Subbulaxmi; Vadivel, Nidyanandh; Fiorina, Paolo; Yagita, Hideo; Azuma, Miyuki; Atkinson, Mark; Bluestone, Jeffrey A; Sayegh, Mohamed H

    2007-10-01

    The PD-1-PDL1 pathway plays a critical role in regulating autoimmune diabetes as blockade or deficiency of PD-1 or PDL1 results in accelerated disease in NOD mice. We explored the cellular mechanisms involved in the regulation of these autoimmune responses by investigations involving various gene-deficient mice on the NOD background. Administration of blocking anti-PDL1 antibody to CD4+ T cell-deficient, CD8+ T cell-deficient and B cell-deficient mice demonstrated that PDL1-mediated regulation of autoreactive CD4+ and CD8+ T cells is critical for diabetes development. This concept was confirmed by adoptive transfer studies utilizing lymphocytes from BDC2.5 and 4.1 (CD4+) TCR transgenic mice and 8.3 (CD8+) TCR transgenic mice; efforts showing increased proliferation of both CD4+ and CD8+ T cells following PDL1 blockade in vivo. Furthermore, we observed that anti-PDL1-mediated acceleration is dependent upon events occurring in the pancreatic lymph nodes during early disease stages, but becomes independent of the pancreatic lymph nodes during later disease stages. These data provide strong evidence that PDL1 regulates autoimmune diabetes by limiting the expansion of CD4+ and CD8+ autoreactive T cells, and define the timing and locale of PDL1-mediated regulation of type 1 diabetes.

  19. Probing the mechanisms of silicon-mediated pathogen resistance.

    PubMed

    Cai, Kunzheng; Gao, Dan; Chen, Jining; Luo, Shiming

    2009-01-01

    Silicon is the second most abundant mineral element in soil, it has important role in alleviating various environmental stresses and enhancing plant resistance against pathogen, but the exact mechanism by which Si mediates pathogen resistance remains unclear. One of the resistance mechanisms is related to silicon deposition in leaf that acts as a physical barrier to hinder pathogen penetration. But more evidence show that silicon can induce defense responses that are functionally similar to systemic acquired resistance, Si-treated plants can significantly increase antioxidant enzyme activities and the production of antifungal compounds such as phenolic metabolism product, phytoalexins and pathogenesis-related proteins etc. Molecular and biochemical detections show that Si can activate the expression of defense-related genes and may play important role in the transduction of plant stress signal such as salicylic acid, jasmonic acid and ethylene.

  20. Receptor- and reactive intermediate-mediated mechanisms of teratogenesis.

    PubMed

    Wells, Peter G; Lee, Crystal J J; McCallum, Gordon P; Perstin, Julia; Harper, Patricia A

    2010-01-01

    Drugs and environmental chemicals can adversely alter the development of the fetus at critical periods during pregnancy, resulting in death, or in structural and functional birth defects in the surviving offspring. This process of teratogenesis may not be evident until a decade or more after birth. Postnatal functional abnormalities include deficits in brain function, a variety of metabolic diseases, and cancer. Due to the high degree of fetal cellular division and differentiation, and to differences from the adult in many biochemical pathways, the fetus is highly susceptible to teratogens, typically at low exposure levels that do not harm the mother. Insights into the mechanisms of teratogenesis come primarily from animal models and in vitro systems, and involve either receptor-mediated or reactive intermediate-mediated processes. Receptor-mediated mechanisms involving the reversible binding of xenobiotic substrates to a specific receptor are exemplified herein by the interaction of the environmental chemical 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or "dioxin") with the cytosolic aryl hydrocarbon receptor (AHR), which translocates to the nucleus and, in association with other proteins, binds to AH-responsive elements (AHREs) in numerous genes, initiating changes in gene transcription that can perturb development. Alternatively, many xenobiotics are bioactivated by fetal enzymes like the cytochromes P450 (CYPs) and prostaglandin H synthases (PHSs) to highly unstable electrophilic or free radical reactive intermediates. Electrophilic reactive intermediates can covalently (irreversibly) bind to and alter the function of essential cellular macromolecules (proteins, DNA), causing developmental anomalies. Free radical reactive intermediates can enhance the formation of reactive oxygen species (ROS), resulting in oxidative damage to cellular macromolecules and/or altered signal transduction. The teratogenicity of reactive intermediates is determined to a large extent

  1. Mechanisms mediating parallel action monitoring in fronto-striatal circuits.

    PubMed

    Beste, Christian; Ness, Vanessa; Lukas, Carsten; Hoffmann, Rainer; Stüwe, Sven; Falkenstein, Michael; Saft, Carsten

    2012-08-01

    Flexible response adaptation and the control of conflicting information play a pivotal role in daily life. Yet, little is known about the neuronal mechanisms mediating parallel control of these processes. We examined these mechanisms using a multi-methodological approach that integrated data from event-related potentials (ERPs) with structural MRI data and source localisation using sLORETA. Moreover, we calculated evoked wavelet oscillations. We applied this multi-methodological approach in healthy subjects and patients in a prodromal phase of a major basal ganglia disorder (i.e., Huntington's disease), to directly focus on fronto-striatal networks. Behavioural data indicated, especially the parallel execution of conflict monitoring and flexible response adaptation was modulated across the examined cohorts. When both processes do not co-incide a high integrity of fronto-striatal loops seems to be dispensable. The neurophysiological data suggests that conflict monitoring (reflected by the N2 ERP) and working memory processes (reflected by the P3 ERP) differentially contribute to this pattern of results. Flexible response adaptation under the constraint of high conflict processing affected the N2 and P3 ERP, as well as their delta frequency band oscillations. Yet, modulatory effects were strongest for the N2 ERP and evoked wavelet oscillations in this time range. The N2 ERPs were localized in the anterior cingulate cortex (BA32, BA24). Modulations of the P3 ERP were localized in parietal areas (BA7). In addition, MRI-determined caudate head volume predicted modulations in conflict monitoring, but not working memory processes. The results show how parallel conflict monitoring and flexible adaptation of action is mediated via fronto-striatal networks. While both, response monitoring and working memory processes seem to play a role, especially response selection processes and ACC-basal ganglia networks seem to be the driving force in mediating parallel conflict

  2. Atenolol Reduces Leishmania major-Induced Hyperalgesia and TNF-α Without Affecting IL-1β or Keratinocyte Derived Chemokines (KC)

    PubMed Central

    Karam, Marc C.; Merckbawi, Rana; Salman, Sara; Mobasheri, Ali

    2016-01-01

    Infection with a high dose of the intracellular parasitic protozoan Leishmania major induces a sustained hyperalgesia in susceptible BALB/c mice accompanied by up-regulation of the pro-inflammatory cytokines IL-1β and IL-6. Interleukin-13 (IL-13) has been shown to reduce this hyperalgesia (despite increased levels of IL-6) and the levels of IL-1β during and after the treatment period. These findings favor the cytokine cascade leading to the production of sympathetic amines (involving TNF-α and KC) over prostaglandins (involving IL-lβ and IL-6) as the final mediators of hyperalgesia. The aim of this study was to investigate the effect of daily treatment with the β-blockers atenolol on L. major-induced inflammation in mice with respect to hyperalgesia as well as the levels of TNF-α and KC (the analog of IL-8 in mice). Our data demonstrates that atenolol is able to reduce the L. major induced sustained peripheral hyperalgesia, which does not seem to involve a direct role for neither IL-lβ nor KC. Moreover, our results show that TNF-α may play a pivotal and direct role in sensitizing the peripheral nerve endings (nociceptors) since its level was reduced during the period of atenolol treatment, which correlates well with the reduction of the observed peripheral, but not central, hyperalgesia. These findings contribute to a better understanding of the cytokine cascade leading to hyperalgesia and may lead to the development of new and more efficient medications for many types of pain. PMID:26913003

  3. Down-regulation of PPARα in the spinal cord contributes to augmented peripheral inflammation and inflammatory hyperalgesia in diet-induced obese rats.

    PubMed

    Wang, J; Zhang, Q; Zhao, L; Li, D; Fu, Z; Liang, L

    2014-10-10

    Obesity is associated with augmented peripheral inflammation and pain sensitivity in response to inflammatory stimulation, but the underlying mechanisms remain unclear. Emerging evidence has shown that activation of peroxisome proliferator-activated receptor-α (PPARα) in the central nervous system controls peripheral inflammation and pain. We hypothesized that obesity might down-regulate PPARα in the spinal cord, leading to enhanced peripheral inflammation and inflammatory hyperalgesia. Sprague-Dawley rats fed a high-fat diet (HF) for 12weeks developed metabolic disorder and displayed significantly decreased spinal PPARα expression and activity. Interestingly, intracerebroventricular (ICV) infusion of the PPARα activator palmitoylethanolamide (PEA) in HF-fed rats for 2weeks normalized spinal PPARα expression and activity without altering metabolic parameters. HF-fed rats were more sensitive to stimulation of the inflamed paw, and exhibited more severe paw edema following carrageenan injection, whereas HF-fed rats receiving ICV PEA had similar pain sensitivity and paw edema to LF-fed rats. No difference in the expression of inflammatory mediators or nuclear factor (NF)-κB activity was observed at baseline among groups. Carrageenan induced decreased PPARα expression and activity, increased spinal cord inflammatory mediator expression and NF-κB activity in both LF-and HF-fed rats. However, the increase was more pronounced in HF-fed rats and corrected by PEA. Intrathecal injection of small interfering RNA (siRNA) against PPARα in HF-fed rats completely abolished PEA effects on peripheral pain sensitivity and paw edema. These findings suggest that diet-induced obesity causes down-regulation of spinal PPARα, which facilitates the susceptibility to peripheral inflammatory challenge by increasing inflammatory response in the spinal cord, contributing to augmented peripheral inflammation and inflammatory hyperalgesia in obesity.

  4. Sildenafil Treatment Eliminates Pruritogenesis and Thermal Hyperalgesia in Rats with Portacaval Shunts.

    PubMed

    Belghiti, Majedeline; Agusti, Ana; Hernandez-Rabaza, Vicente; Cabrera-Pastor, Andrea; Llansola, Marta; Felipo, Vicente

    2017-03-01

    Pruritus is a common symptom in chronic liver diseases, which may also alter thermal sensitivity. The underlying mechanisms remain unclear and treatments are not satisfactory. Portal-systemic shunting has been proposed to alter thermal sensitivity in cirrhotics. Inflammation-induced enhanced activity of the Transient Receptor Potential Vanilloid 1 (TRPV1) may contribute to pruritus and thermal hyperalgesia. Sildenafil reduces neuroinflammation in portacaval shunt (PCS) rats. The aims were to assess whether: (1) PCS rats show enhanced scratching or thermal sensitivity; (2) TRPV1 activity is enhanced in PCS rats; (3) treatment with sildenafil reduces TRPV1 activation, scratching and thermal hyperalgesia. Rats were treated with sildenafil beginning 3 weeks after surgery. The number of scratches performed were counted. Thermal hyperalgesia was analyzed using the Hargreaves' Plantar Test. TRPV1 activation by measuring the increase in Ca(2+) induced by capsaicin in dorsal root ganglia neurons. PCS rats show enhanced scratching behavior, reaching 66 ± 5 scratches/h (p < 0.01) at 21 days after surgery, while controls show 37 ± 2 scratches/h. PCS rats show thermal hyperalgesia. Paw withdrawal latency was reduced (p < 0.05) to 10 ± 1 s compared to controls (21 ± 2 s). Capsaicin-induced calcium increase was higher in dorsal root ganglia cultures from PCS rats, indicating TRPV1functional increase. PCS rats show enhanced scratching behavior and thermal sensitivity and are a good model to study these alterations in chronic liver diseases. Enhanced sensitivity and activity of TRPV1 channel underlies these alterations. Treatment with sildenafil reduces TRPV1 channel sensitivity and activity and normalizes scratching behavior and thermal sensitivity.

  5. The effect of the sex of a model on nocebo hyperalgesia induced by social observational learning.

    PubMed

    Swider, Karolina; Bąbel, Przemysław

    2013-08-01

    Research shows that placebo analgesia can be induced through social observational learning. Our aim was to replicate and extend this result by studying the effect of the sex of both the model and the subject on the magnitude of placebo analgesia induced by social observational learning. Four experimental (1 through 4) and 2 control (5 and 6) groups were observed: groups 1, 3, and 5 were female; groups 2, 4, and 6 were male. All subjects received pain stimuli of the same intensity preceded by green and red lights. Before receiving pain stimuli, groups 1 and 4 observed a female model and groups 2 and 3 a male model; both models simulated responses to pain stimuli preceded by green lights as less painful than those preceded by red lights. Groups 1 through 4 also rated pain stimuli preceded by green lights as less painful. Further investigation revealed that in fact subjects in experimental groups rated red-associated stimuli as more painful than subjects from control groups who did not observe a model before receiving the same pain stimuli, indicating that nocebo hyperalgesia rather than placebo analgesia was induced. Empathy traits predicted the magnitude of nocebo hyperalgesia. Regardless of the sex of the subject, nocebo hyperalgesia was greater after the male model was observed. The results show that social observational learning is a mechanism that produces placebo effects. They also indicate that the sex of the model plays an important role in this process.

  6. Experimental hypoglycemia is a human model of stress-induced hyperalgesia.

    PubMed

    Gibbons, Christopher H; Adler, Gail K; Bonyhay, Istvan; Freeman, Roy

    2012-11-01

    Hypoglycemia is a physiological stress that leads to the release of stress hormones, such as catecholamines and glucocorticoids, and proinflammatory cytokines. These factors, in euglycemic animal models, are associated with stress-induced hyperalgesia. The primary aim of this study was to determine whether experimental hypoglycemia in humans would lead to a hyperalgesic state. In 2 separate 3-day admissions separated by 1 to 3 months, healthy study participants were exposed to two 2-hour euglycemic hyperinsulinemic clamps or two 2-hour hypoglycemic hyperinsulinemic clamps. Thermal quantitative sensory testing and thermal pain assessments were measured the day before and the day after euglycemia or hypoglycemia. In contrast to prior euglycemia exposure, prior hypoglycemia exposure resulted in enhanced pain sensitivity to hot and cold stimuli as well as enhanced temporal summation to repeated heat-pain stimuli. These findings suggest that prior exposure to hypoglycemia causes a state of enhanced pain sensitivity that is consistent with stress-induced hyperalgesia. This human model may provide a framework for hypothesis testing and targeted, mechanism-based pharmacological interventions to delineate the molecular basis of hyperalgesia and pain susceptibility.

  7. [Tc1-mediated contact sensitivity reaction, its mechanism and regulation].

    PubMed

    Zemelka-Wiącek, Magdalena; Szczepanik, Marian

    2014-07-04

    The contact hypersensitivity reaction (CHS) to haptens is a classic example of cell-mediated immune response. In the effector phase, two stages can be distinguished: an early component, that appears only 2 hours after subsequent contact with the hapten, and the late component that develops approximately 24 hours later which is mediated by TCRαβ+ cells. The effector lymphocytes may be CD4+ T helper 1 (Th1) cells or CD8+ T cytotoxic 1 (Tc1) cells, which depends on the employed hapten and/or mice strain. NKT lymphocytes play the crucial role in the CHS initiation, by supporting B1 cells in the antigen-specific IgM antibodies production. The development of an early component is essential for the recruitment of T effector (Teff) cells to the side of hapten deposition and for the complete expansion of inflammatory reaction. The CHS reaction is under T regulatory (Treg) cells control, both in the induction phase as well as in the effector phase. A new view of a negative regulation of the Tc1 mediated CHS response is based on the suppression induced by epicutaneous (EC) application of protein antigen. The DNP-BSA skin application, on a gauze patch, leads to a state of immunosuppression. This maneuver results in rising the population of Treg cells with TCRαβ+CD4+CD25+Foxp3+ phenotype. The mechanism of suppression requires direct contact between Treg cells and Teff cells and the participation of CTLA-4 molecule is also necessary. The described method of evoking immune tolerance via EC immunization may contribute to elaborate a new method of allergic contact dermatitis therapy. This is because of its effectiveness, ease of induction and non-invasive protein antigen application.

  8. Regionally selective activation of ERK and JNK in morphine paradoxical hyperalgesia: a step toward improving opioid pain therapy.

    PubMed

    Sanna, Maria Domenica; Ghelardini, Carla; Galeotti, Nicoletta

    2014-11-01

    In addition to analgesia, opioid agonists may increase pain sensitivity under different conditions varying dose and administration pattern. While opioid hyperalgesia induced by tolerance and withdrawal is largely studied, little is known on the mechanisms underlying ultra-low dose morphine hyperalgesia. This pronociceptive response appears to play an opposing role in morphine analgesia and might have clinical relevance. Ultra-low dose morphine elicited thermal hyperalgesia through activation of μ opioid receptors. To elucidate the intracellular mechanism of morphine nociceptive behaviour, we investigated the mitogen-activated protein kinase (MAPK), crucial pathways in pain hypersensitivity. The catalytic activity of extracellular signal-regulated kinase (ERK), p38, c-Jun-N-terminal kinase (JNK), upstream modulators and transcription factors was investigated in the mouse periaqueductal grey matter (PAG), thalamus and prefrontal cortex by western blotting. Ultra-low dose morphine intensively increased pERK1 contents in the PAG and cortex and, to a lesser extent, increased cortical ERK2 and JNK phosphorylation. No involvement of p38 was detected. Morphine exposure also increased phosphorylation of cortical c-Jun whereas levels of phosphorylated cAMP response element-binding protein (CREB) remained unmodified. Blockade of protein kinase C (PKC) prevented increases in phosphorylation showing a PKC-dependent mechanism of activation. Pharmacological inhibitors of PKC, ERK, and JNK activity prevented morphine hyperalgesia. No modulation of MAPK and transcription factors' activity was detected in the thalamus. These results support the concept that selective activation of ERK and JNK on descending pathways plays an important role in ultra-low dose morphine hyperalgesia. The modulation of these signalling processes might improve pain management with opiate analgesics.

  9. A possible mechanism in DHEA-mediated protection against osteoarthritis.

    PubMed

    Li, Wei-Jun; Tang, Lu-Ping; Xiong, Yan; Chen, Wei-Ping; Zhou, Xin-Die; Ding, Qian-Hai; Wu, Li-Dong

    2014-11-01

    Dehydroepiandrosterone (DHEA) and its ester form, DHEA-S, are the most abundant steroids in human plasma. Our previous studies showed that DHEA protects against osteoarthritis (OA). The aim of this paper was to explore the possible mechanisms that underlie DHEA-mediated protection against OA. We tested the expression of β-catenin, it was increased significantly in OA. Rabbit cartilage was treated with various concentrations of DHEA in both IL-1β-induced rabbit chondrocytes and in rabbit cartilage from the anterior cruciate ligament transaction-induced OA model. We found DHEA decreased the expression of β-catenin. Then we further activated Wnt/β-catenin signaling by β-catenin transfection and inactivated it by the inhibitor Dickkopf1 in chondrocytes to reveal its role in the pathogenesis of OA. It turns out the protective effect of DHEA was significantly decreased when Wnt/β-catenin signaling was activated, while inactivating Wnt/β-catenin signaling enhanced the effects of DHEA. Therefore, we hypothesize that DHEA probably exerted its chondroprotective effect by regulating Wnt/β-catenin signaling. Our findings demonstrate the critical role of Wnt/β-catenin signaling in DHEA-mediated protection against OA.

  10. IL-1beta in the trigeminal subnucleus caudalis contributes to extra-territorial allodynia/hyperalgesia following a trigeminal nerve injury.

    PubMed

    Takahashi, Kouji; Watanabe, Mineo; Suekawa, Yohei; Ito, Goshi; Inubushi, Toshihiro; Hirose, Naoto; Murasaki, Kyoko; Hiyama, Shinji; Uchida, Takashi; Tanne, Kazuo

    2011-05-01

    It has been reported that the whisker pad (WP) area, which is innervated by the second branch of the trigeminal nerve, shows allodynia/hyperalgesia following transection of the mental nerve (MN: the third branch of the trigeminal nerve). However, the mechanisms of this extra-territorial pain induction still remain unclear. Glia and cytokines are known to facilitate perception of noxious input, raising a possibility that these non-neuronal elements are involved in the induction and spread of allodynia/hyperalgesia at non-injured skin territory. One day after MN transection, tactile allodynia/hyperalgesia developed on the ipsilateral WP area, which is in the non-injured skin territory. The tactile allodynia/hyperalgesia lasted for more than 56 days. In response to MN transection, astrocytes and microglia appeared to be in an activated state, and interleukin (IL)-1beta was up-regulated in astrocytes in the trigeminal subnucleus caudalis (Vc). Allodynia/hyperalgesia at WP area induced by MN transection was attenuated dose-dependently by IL-1 receptor antagonist IL-1ra (i.t., 0.05, 0.5, and 5 pg/rat). Fos-like immunoreactive (Fos-Li) neurons were observed in the Vc after non-noxious mechanical stimulation of the WP area in the rats with MN transection. Administration of IL-1ra also attenuated the number of Fos-Li neurons dose-dependently. Administration of a noncompetitive antagonist of NMDA receptors MK-801 (i.t., 5 μg/rat) reversed allodynia/hyperalgesia. IL-1 receptor type I (IL-1RI) was localized in Fos- and phospho NR1-immunoreactive neurons. These results suggest that IL-1beta in the Vc plays an important role in the development of extra-territorial tactile allodynia/hyperalgesia after MN transection.

  11. The degree of acute descending control of spinal nociception in an area of primary hyperalgesia is dependent on the peripheral domain of afferent input.

    PubMed

    Drake, Robert A R; Hulse, Richard P; Lumb, Bridget M; Donaldson, Lucy F

    2014-08-15

    Descending controls of spinal nociceptive processing play a critical role in the development of inflammatory hyperalgesia. Acute peripheral nociceptor sensitization drives spinal sensitization and activates spino-supraspinal-spinal loops leading to descending inhibitory and facilitatory controls of spinal neuronal activity that further modify the extent and degree of the pain state. The afferent inputs from hairy and glabrous skin are distinct with respect to both the profile of primary afferent classes and the degree of their peripheral sensitization. It is not known whether these differences in afferent input differentially engage descending control systems to different extents or in different ways. Injection of complete Freund's adjuvant resulted in inflammation and swelling of hairy hind foot skin in rats, a transient thermal hyperalgesia lasting <2 h, and longlasting primary mechanical hyperalgesia (≥7 days). Much longer lasting thermal hyperalgesia was apparent in glabrous skin (1 h to >72 h). In hairy skin, transient hyperalgesia was associated with sensitization of withdrawal reflexes to thermal activation of either A- or C-nociceptors. The transience of the hyperalgesia was attributable to a rapidly engaged descending inhibitory noradrenergic mechanism, which affected withdrawal responses to both A- and C-nociceptor activation and this could be reversed by intrathecal administration of yohimbine (α-2-adrenoceptor antagonist). In glabrous skin, yohimbine had no effect on an equivalent thermal inflammatory hyperalgesia. We conclude that acute inflammation and peripheral nociceptor sensitization in hind foot hairy skin, but not glabrous skin, rapidly activates a descending inhibitory noradrenergic system. This may result from differences in the engagement of descending control systems following sensitization of different primary afferent classes that innervate glabrous and hairy skin.

  12. Mechanisms of peroxynitrite-mediated nitration of tyrosine.

    PubMed

    Gunaydin, Hakan; Houk, K N

    2009-05-01

    The mechanisms of tyrosine nitration by peroxynitrous acid or nitrosoperoxycarbonate were investigated with the CBS-QB3 method. Either the protonation of peroxynitrite or a reaction with carbon dioxide gives a reactive peroxide intermediate. Peroxynitrous acid-mediated nitration of phenol occurs via unimolecular decomposition to give nitrogen dioxide and hydroxyl radicals. Nitrosoperoxycarbonate also undergoes unimolecular decomposition to give carbonate and nitrogen dioxide radicals. The reactions of tyrosine with the hydroxyl or carbonate radicals give a phenoxy radical intermediate. The reaction of the nitrogen dioxide with this radical intermediate followed by tautomerization gives nitrated tyrosine in both cases. According to CBS-QB3 calculations, the rate-limiting step for the nitration of phenol is the decomposition of peroxynitrous acid or nitrosoperoxycarbonate.

  13. Quantification of Grain Boundary Mediated Plasticity Mechanisms in Nanocrystalline Metals

    NASA Astrophysics Data System (ADS)

    Panzarino, Jason F.

    Nanocrystalline metals have been a topic of great discussion over recent years due to their exceptional strengths and novel grain boundary-mediated deformation mechanisms. Their microstructures are known to evolve through dynamic processes such as grain boundary migration and grain rotation, but how the collective interaction of these mechanisms alter the microstructure on a larger scale is not completely understood. In this thesis, we present coupled atomistic modeling and experimental tasks that aim to understand how the grain structure, grain boundaries, and associated grain boundary network change during nanocrystalline plasticity. Due to the complex three-dimensional nature of these mechanisms and the limited spatial and temporal resolution of current in-situ experimental techniques, we turn to atomistic modeling to help understand the dynamics by which these mechanisms unfold. In order to provide a quantitative analysis of this behavior, we develop a tool which fully characterizes nanocrystalline microstructures in atomistic models and subsequently tracks their evolution during molecular dynamics simulations. We then use this algorithm to quantitatively track grain structure and boundary network evolution in plastically deformed nanocrystalline Al, finding that higher testing temperature and smaller average grain size results in increased evolution of grain structure with evidence of larger scale changes to the grain boundary network also taking place. This prompts us to extend our analysis technique to include full characterization of grain boundary networks and rigorous topographical feature identification. We then employ this tool on simulations of Al subject to monotonic tension, cycling loading, and simple annealing, and find that each case results in different evolution of the grain boundary network. Finally, our computational work is complemented synergistically by experimental analyses which track surface microstructure evolution during sliding wear

  14. Continuing to illuminate the mechanisms underlying UV-mediated melanomagenesis.

    PubMed

    Dellinger, Ryan W; Liu-Smith, Feng; Meyskens, Frank L

    2014-09-05

    The incidence of melanoma is one of the fastest growing of all tumor types in the United States and the number of cases worldwide has doubled in the past 30 years. Melanoma, which arises from melanocytes, is an extremely aggressive tumor that invades the vascular and lymphatic systems to establish tumors elsewhere in the body. Melanoma is a particularly resilient cancer and systemic therapy approaches have achieved minimal success against metastatic melanoma resulting in only a few FDA-approved treatments with limited benefit. Leading treatments offer minimal efficacy with response rates generally under 15% in the long term with no clear effect on melanoma-related mortality. Even the recent success of the specific BRAF mutant inhibitor vemurafenib has been tempered somewhat since acquired resistance is rapidly observed. Thus, understanding the mechanism(s) of melanoma carcinogenesis is paramount to combating this deadly disease. Not only for the treatment of melanoma but, ultimately, for prevention. In this report, we will summarize our work to date regarding the characterization of ultraviolet radiation (UVR)-mediated melanomagenesis and highlight several promising avenues of ongoing research.

  15. 5-hydroxyindolacetic acid (5-HIAA), a main metabolite of serotonin, is responsible for complete Freund's adjuvant-induced thermal hyperalgesia in mice

    PubMed Central

    2011-01-01

    Background The role of serotonin (5-hydroxytrptamine, 5-HT) in the modulation of pain has been widely studied. Previous work led to the hypothesis that 5-hydroxyindolacetic acid (5-HIAA), a main metabolite of serotonin, might by itself influence pain thresholds. Results In the present study, we investigated the role of 5-HIAA in inflammatory pain induced by intraplantar injection of complete Freund's adjuvant (CFA) into the hind paw of mice. Wild-type mice were compared to mice deficient of the 5-HT transporter (5-HTT-/- mice) using behavioral tests for hyperalgesia and high-performance liquid chromatography (HPLC) to determine tissue levels of 5-HIAA. Wild-type mice reproducibly developed thermal hyperalgesia and paw edema for 5 days after CFA injection. 5-HTT-/- mice treated with CFA had reduced thermal hyperalgesia on day 1 after CFA injection and normal responses to heat thereafter. The 5-HIAA levels in spinal cord and sciatic nerve as measured with HPLC were lower in 5-HTT-/- mice than in wild-type mice after CFA injection. Pretreatment of wild-type mice with intraperitoneal injection of para-chlorophenylalanine (p-CPA), a serotonin synthesis inhibitor, resulted in depletion of the 5-HIAA content in spinal cord and sciatic nerve and decrease in thermal hyperalgesia in CFA injected mice. The application of exogenous 5-HIAA resulted in potentiation of thermal hyperalgesia induced by CFA in 5-HTT-/- mice and in wild-type mice pretreated with p-CPA, but not in wild-type mice without p-CPA pretreatment. Further, methysergide, a broad-spectrum serotonin receptor antagonist, had no effect on 5-HIAA-induced potentiation of thermal hyperalgesia in CFA-treated wild-type mice. Conclusion Taken together, the present results suggest that 5-HIAA plays an important role in modulating peripheral thermal hyperalgesia in CFA induced inflammation, probably via a non-serotonin receptor mechanism. PMID:21447193

  16. Activation of cannabinoid CB2 receptors reduces hyperalgesia in an experimental autoimmune encephalomyelitis mouse model of multiple sclerosis.

    PubMed

    Fu, Weisi; Taylor, Bradley K

    2015-05-19

    Clinical trials investigating the analgesic efficacy of cannabinoids in multiple sclerosis have yielded mixed results, possibly due to psychotropic side effects mediated by cannabinoid CB1 receptors. We hypothesized that, a CB2-specific agonist (JWH-133) would decrease hyperalgesia in an experimental autoimmune encephalomyelitis mouse model of multiple sclerosis. Four weeks after induction of experimental autoimmune encephalomyelitis, we found that intrathecal administration of JWH-133 (10-100μg) dose-dependently reduced both mechanical and cold hypersensitivity without producing signs of sedation or ataxia. The anti-hyperalgesic effects of JWH-133 could be dose-dependently prevented by intrathecal co-administration of the CB2 antagonist, AM-630 (1-3μg). Our results suggest that JWH-133 acts at CB2 receptors, most likely within the dorsal horn of the spinal cord, to suppress the hypersensitivity associated with experimental autoimmune encephalomyelitis. These are the first pre-clinical studies to directly promote CB2 as a promising target for the treatment of central pain in an animal model of multiple sclerosis.

  17. ARID1B-mediated disorders: Mutations and possible mechanisms

    PubMed Central

    Sim, Joe C. H.; White, Susan M; Lockhart, Paul J.

    2015-01-01

    Summary Mutations in the gene encoding AT-rich interactive domain-containing protein 1B (ARID1B) were recently associated with multiple syndromes characterized by developmental delay and intellectual disability, in addition to nonsyndromic intellectual disability. While the majority of ARID1B mutations identified to date are predicted to result in haploinsufficiency, the underlying pathogenic mechanisms have yet to be fully understood. ARID1B is a DNA-binding subunit of the Brahma-associated factor chromatin remodelling complexes, which play a key role in the regulation of gene activity. The function of remodelling complexes can be regulated by their subunit composition, and there is some evidence that ARID1B is a component of the neuron-specific chromatin remodelling complex. This complex is involved in the regulation of stem/progenitor cells exiting the cell cycle and differentiating into postmitotic neurons. Recent research has indicated that alterations in the cell cycle contribute to the underlying pathogenesis of syndromes associated with ARID1B haploinsufficiency in fibroblasts derived from affected individuals. This review describes studies linking ARID1B to neurodevelopmental disorders and it summarizes the function of ARID1B to provide insights into the pathogenic mechanisms underlying ARID1B-mediated disorders. In conclusion, ARID1B is likely to play a key role in neurodevelopment and reduced levels of wild-type protein compromise normal brain development. Additional studies are required to determine the mechanisms by which impaired neural development contributes to the intellectual disability and speech impairment that are consistently observed in individuals with ARID1B haploinsufficiency. PMID:25674384

  18. Neuropeptide Y in the rostral ventromedial medulla reverses inflammatory and nerve injury hyperalgesia in rats via non-selective excitation of local neurons

    PubMed Central

    Cleary, Daniel R.; Roeder, Zachary; Elkhatib, Rania; Heinricher, Mary M.

    2014-01-01

    Chronic pain reflects not only sensitization of the ascending nociceptive pathways, but also changes in descending modulation. The rostral ventromedial medulla (RVM) is a key structure in a well-studied descending pathway, and contains two classes of modulatory neurons, the ON-cells and the OFF-cells. Disinhibition of OFF-cells depresses nociception; increased ON-cell activity facilitates nociception. Multiple lines of evidence show that sensitization of ON-cells contributes to chronic pain, and reversing or blocking this sensitization is of interest as a treatment of persistent pain. Neuropeptide Y (NPY) acting via the Y1 receptor has been shown to attenuate hypersensitivity in nerve-injured animals without affecting normal nociception when microinjected into the RVM, but the neural basis for this effect was unknown. We hypothesized that behavioral anti-hyperalgesia was due to selective inhibition of ON-cells by NPY at the Y1 receptor. To explore the possibility of Y1 selectivity on ON-cells, we stained for the NPY-Y1 receptor in the RVM, and found it broadly expressed on both serotonergic and non-serotonergic neurons. In subsequent behavioral experiments, NPY microinjected into the RVM in lightly anesthetized animals reversed signs of mechanical hyperalgesia following either nerve injury or chronic hindpaw inflammation. Unexpectedly, rather than decreasing ON-cell activity, NPY increased spontaneous activity of both ON- and OFF-cells without altering noxious-evoked changes in firing. Based on these results, we conclude that the anti-hyperalgesic effects of NPY in the RVM are not explained by selective inhibition of ON-cells, but rather by increased spontaneous activity of OFF-cells. Although ON-cells undoubtedly facilitate nociception and contribute to hypersensitivity, the present results highlight the importance of parallel OFF-cell mediated descending inhibition in limiting the expression of chronic pain. PMID:24792711

  19. Structural mechanisms of PriA-mediated DNA replication restart.

    PubMed

    Bhattacharyya, Basudeb; George, Nicholas P; Thurmes, Tiffany M; Zhou, Ruobo; Jani, Niketa; Wessel, Sarah R; Sandler, Steven J; Ha, Taekjip; Keck, James L

    2014-01-28

    Collisions between cellular DNA replication machinery (replisomes) and damaged DNA or immovable protein complexes can dissociate replisomes before the completion of replication. This potentially lethal problem is resolved by cellular "replication restart" reactions that recognize the structures of prematurely abandoned replication forks and mediate replisomal reloading. In bacteria, this essential activity is orchestrated by the PriA DNA helicase, which identifies replication forks via structure-specific DNA binding and interactions with fork-associated ssDNA-binding proteins (SSBs). However, the mechanisms by which PriA binds replication fork DNA and coordinates subsequent replication restart reactions have remained unclear due to the dearth of high-resolution structural information available for the protein. Here, we describe the crystal structures of full-length PriA and PriA bound to SSB. The structures reveal a modular arrangement for PriA in which several DNA-binding domains surround its helicase core in a manner that appears to be poised for binding to branched replication fork DNA structures while simultaneously allowing complex formation with SSB. PriA interaction with SSB is shown to modulate SSB/DNA complexes in a manner that exposes a potential replication initiation site. From these observations, a model emerges to explain how PriA links recognition of diverse replication forks to replication restart.

  20. Adipokines and the cardiovascular system: mechanisms mediating health and disease.

    PubMed

    Northcott, Josette M; Yeganeh, Azadeh; Taylor, Carla G; Zahradka, Peter; Wigle, Jeffrey T

    2012-08-01

    This review focuses on the role of adipokines in the maintenance of a healthy cardiovascular system, and the mechanisms by which these factors mediate the development of cardiovascular disease in obesity. Adipocytes are the major cell type comprising the adipose tissue. These cells secrete numerous factors, termed adipokines, into the blood, including adiponectin, leptin, resistin, chemerin, omentin, vaspin, and visfatin. Adipose tissue is a highly vascularised endocrine organ, and different adipose depots have distinct adipokine secretion profiles, which are altered with obesity. The ability of many adipokines to stimulate angiogenesis is crucial for adipose tissue expansion; however, excessive blood vessel growth is deleterious. As well, some adipokines induce inflammation, which promotes cardiovascular disease progression. We discuss how these 7 aforementioned adipokines act upon the various cardiovascular cell types (endothelial progenitor cells, endothelial cells, vascular smooth muscle cells, pericytes, cardiomyocytes, and cardiac fibroblasts), the direct effects of these actions, and their overall impact on the cardiovascular system. These were chosen, as these adipokines are secreted predominantly from adipocytes and have known effects on cardiovascular cells.

  1. Molecular mechanism of statin-mediated LOX-1 inhibition.

    PubMed

    Biocca, Silvia; Iacovelli, Federico; Matarazzo, Sara; Vindigni, Giulia; Oteri, Francesco; Desideri, Alessandro; Falconi, Mattia

    2015-01-01

    Statins are largely used in clinics in the treatment of patients with cardiovascular diseases for their effect on lowering circulating cholesterol. Lectin-like oxidized low-density lipoprotein (LOX-1), the primary receptor for ox-LDL, plays a central role in the pathogenesis of atherosclerosis and cardiovascular disorders. We have recently shown that chronic exposure of cells to lovastatin disrupts LOX-1 receptor cluster distribution in plasma membranes, leading to a marked loss of LOX-1 function. Here we investigated the molecular mechanism of statin-mediated LOX-1 inhibition and we demonstrate that all tested statins are able to displace the binding of fluorescent ox-LDL to LOX-1 by a direct interaction with LOX-1 receptors in a cell-based binding assay. Molecular docking simulations confirm the interaction and indicate that statins completely fill the hydrophobic tunnel that crosses the C-type lectin-like (CTLD) recognition domain of LOX-1. Classical molecular dynamics simulation technique applied to the LOX-1 CTLD, considered in the entire receptor structure with or without a statin ligand inside the tunnel, indicates that the presence of a ligand largely increases the dimer stability. Electrophoretic separation and western blot confirm that different statins binding stabilize the dimer assembly of LOX-1 receptors in vivo. The simulative and experimental results allow us to propose a CTLD clamp motion, which enables the receptor-substrate coupling. These findings reveal a novel and significant functional effect of statins.

  2. Statistical mechanics of DNA-mediated colloidal aggregation

    NASA Astrophysics Data System (ADS)

    Licata, Nicholas A.; Tkachenko, Alexei V.

    2006-10-01

    We present a statistical mechanical model of aggregation in colloidal systems with DNA-mediated interactions. We obtain a general result for the two-particle binding energy in terms of the hybridization free energy ΔG of DNA and two model-dependent properties: the average number of available DNA bridges ⟨N⟩ and the effective DNA concentration ceff . We calculate these parameters for a particular DNA bridging scheme. The fraction of all the n -mers, including the infinite aggregate, are shown to be universal functions of a single parameter directly related to the two-particle binding energy. We explicitly take into account the partial ergodicity of the problem resulting from the slow DNA binding-unbinding dynamics, and introduce the concept of angular localization of DNA linkers. In this way, we obtain a direct link between DNA thermodynamics and the global aggregation and melting properties in DNA-colloidal systems. The results of the theory are shown to be in quantitative agreement with two recent experiments with particles of micron and nanometer size.

  3. The effects of opioid receptor antagonists on electroacupuncture-produced anti-allodynia/hyperalgesia in rats with paclitaxel-evoked peripheral neuropathy

    PubMed Central

    Meng, Xianze; Zhang, Yu; Li, Aihui; Xin, Jiajia; Lao, Lixing; Ren, Ke; Berman, Brian M.; Tan, Ming; Zhang, Rui-Xin

    2011-01-01

    Research supports the effectiveness of acupuncture for conditions such as chronic low back and knee pain. In a five-patient pilot study the modality also improved the symptoms of chemotherapy-induced neuropathic pain. Using an established rat model of paclitaxel-induced peripheral neuropathy, we evaluated the effect of electroacupuncture (EA) on paclitaxel-induced hyperalgesia and allodynia that has not been studied in an animal model. We hypothesize that EA would relieve the paclitaxel-induced mechanical allodynia and hyperalgesia, which was assessed 30 minutes after EA using von Frey filaments. Beginning on day 13, the response frequency to von Frey filaments (4-15 g) was significantly increased in paclitaxel-injected rats compared to those injected with vehicle. EA at 10Hz significantly (p<0.05) decreased response frequency at 4-15 g compared to sham EA; EA at 100Hz only decreased response frequency at 15 g stimulation. Compared to sham EA plus vehicle, EA at 10Hz plus either a μ, δ, or κ opioid receptor antagonist did not significantly decrease mechanical response frequency, indicating that all three antagonists blocked EA inhibition of allodynia and hyperalgesia. Since we previously demonstrated that μ and δ but not κ opioid receptors affect EA anti-hyperalgesia in an inflammatory pain model, these data show that EA inhibits pain through different opioid receptors under varying conditions. Our data indicate that EA at 10Hz inhibits mechanical allodynia/hyperalgesia more potently than does EA at 100Hz. Thus, EA significantly inhibits paclitaxel-induced allodynia/hyperalgesia through spinal opioid receptors, and EA may be a useful complementary treatment for neuropathic pain patients. PMID:21872220

  4. Loss of neurons in rostral ventromedial medulla that express neurokinin-1 receptors decreases the development of hyperalgesia.

    PubMed

    Khasabov, S G; Simone, D A

    2013-10-10

    It is well known that neurons in the rostral ventromedial medulla (RVM) are involved in descending modulation of nociceptive transmission in the spinal cord. It has been shown that activation of neurokinin-1 receptors (NK-1Rs) in the RVM, which are presumably located on pain facilitating ON cells, produces hyperalgesia whereas blockade of NK-1Rs attenuates hyperalgesia. To obtain a better understanding of the functions of NK-1R expressing neurons in the RVM, we selectively ablated these neurons by injecting the stable analog of substance P (SP), Sar(9),Met(O2)(11)-Substance P, conjugated to the ribosomal toxin saporin (SSP-SAP) into the RVM. Rats received injections of SSP-SAP (1 μM) or an equal volume of 1 μM of saporin conjugated to artificial peptide (Blank-SAP). Stereological analysis of NK-1R- and NeuN-labeled neurons in the RVM was determined 21-24 days after treatment. Withdrawal responses to mechanical and heat stimuli applied to the plantar hindpaw were determined 5-28 days after treatment. Withdrawal responses were also determined before and after intraplantar injection of capsaicin (acute hyperalgesia) or complete Freund's adjuvant (CFA) (prolonged hyperalgesia). The proportion of NK-1R-labeled neurons in the RVM was 8.8 ± 1.3% in naïve rats and 8.1 ± 0.8% in rats treated with Blank-SAP. However, injection of SSP-SAP into the RVM resulted in a 90% decrease in NK-1R-labeled neurons. SSP-SAP did not alter withdrawal responses to mechanical or heat stimuli under normal conditions, and did not alter analgesia produced by morphine administered into the RVM. In contrast, the duration of nocifensive behaviors produced by capsaicin and mechanical and heat hyperalgesia produced by capsaicin and CFA were decreased in rats pretreated with SSP-SAP as compared to those that received Blank-SAP. These data support our earlier studies using NK-1R antagonists in the RVM and demonstrate that RVM neurons that possess the NK-1R do not play a significant role in

  5. Pain referral and regional deep tissue hyperalgesia in experimental human hip pain models.

    PubMed

    Izumi, Masashi; Petersen, Kristian Kjær; Arendt-Nielsen, Lars; Graven-Nielsen, Thomas

    2014-04-01

    Hip disorder patients typically present with extensive pain referral and hyperalgesia. To better understand underlying mechanisms, an experimental hip pain model was established in which pain referrals and hyperalgesia could be studied under standardized conditions. In 16 healthy subjects, pain was induced by hypertonic saline injection into the gluteus medius tendon (GMT), adductor longus tendon (ALT), or gluteus medius muscle (GMM). Isotonic saline was injected contralaterally as control. Pain intensity was assessed on a visual analogue scale (VAS), and subjects mapped the pain distribution. Before, during, and after injections, passive hip joint pain provocation tests were completed, together with quantitative sensory testing as follows: pressure pain thresholds (PPTs), cuff algometry pain thresholds (cuff PPTs), cutaneous pin-prick sensitivity, and thermal pain thresholds. Hypertonic saline injected into the GMT resulted in higher VAS scores than hypertonic injections into the ALT and GMM (P<.05). Referred pain areas spread to larger parts of the leg after GMT and GMM injections compared with more regionalized pain pattern after ALT injections (P<.05). PPTs at the injection site were decreased after hypertonic saline injections into GMT and GMM compared with baseline, ALT injections, and isotonic saline. Cuff PPTs from the thigh were decreased after hypertonic saline injections into the ALT compared with baseline, GMT injections, and isotonic saline (P<.05). More subjects had positive joint pain provocation tests after hypertonic compared with isotonic saline injections (P<.05), indicating that this provocation test also assessed hyperalgesia in extra-articular soft tissues. The experimental models may open for better understanding of pain mechanisms associated with painful hip disorders.

  6. A comprehensive review of opioid-induced hyperalgesia.

    PubMed

    Lee, Marion; Silverman, Sanford M; Hansen, Hans; Patel, Vikram B; Manchikanti, Laxmaiah

    2011-01-01

    Opioid-induced hyperalgesia (OIH) is defined as a state of nociceptive sensitization caused by exposure to opioids. The condition is characterized by a paradoxical response whereby a patient receiving opioids for the treatment of pain could actually become more sensitive to certain painful stimuli. The type of pain experienced might be the same as the underlying pain or might be different from the original underlying pain. OIH appears to be a distinct, definable, and characteristic phenomenon that could explain loss of opioid efficacy in some patients. Findings of the clinical prevalence of OIH are not available. However, several observational, cross-sectional, and prospective controlled trials have examined the expression and potential clinical significance of OIH in humans. Most studies have been conducted using several distinct cohorts and methodologies utilizing former opioid addicts on methadone maintenance therapy, perioperative exposure to opioids in patients undergoing surgery, and healthy human volunteers after acute opioid exposure using human experimental pain testing. The precise molecular mechanism of OIH, while not yet understood, varies substantially in the basic science literature, as well as clinical medicine. It is generally thought to result from neuroplastic changes in the peripheral and central nervous system (CNS) that lead to sensitization of pronociceptive pathways. While there are many proposed mechanisms for OIH, 5 mechanisms involving the central glutaminergic system, spinal dynorphins, descending facilitation, genetic mechanisms, and decreased reuptake and enhanced nociceptive response have been described as the important mechanisms. Of these, the central glutaminergic system is considered the most common possibility. Another is the hypothesis that N-methyl-D-aspartate (NMDA) receptors in OIH include activation, inhibition of the glutamate transporter system, facilitation of calcium regulated intracellular protein kinase C, and cross

  7. Visceral hyperalgesia caused by peptide YY deletion and Y2 receptor antagonism

    PubMed Central

    Hassan, Ahmed M.; Jain, Piyush; Mayerhofer, Raphaela; Fröhlich, Esther E.; Farzi, Aitak; Reichmann, Florian; Herzog, Herbert; Holzer, Peter

    2017-01-01

    Altered levels of colonic peptide YY (PYY) have been reported in patients suffering from functional and inflammatory bowel disorders. While the involvement of neuropeptide Y (NPY) and Y receptors in the regulation of nociception is well established, the physiological role of PYY in somatic and visceral pain is poorly understood. In this work, the role of PYY in pain sensitivity was evaluated using PYY knockout (PYY(−/−)) mice and Y2 receptor ligands. PYY(−/−) mice were more sensitive to somatic thermal pain compared to wild type (WT) mice. Visceral pain was assessed by evaluating pain-related behaviors, mouse grimace scale (MGS) and referred hyperalgesia after intrarectal administration of allyl isothiocyanate (AITC, 1 or 2%) or its vehicle, peanut oil. The pain-related behaviors induced by AITC were significantly exaggerated by PYY deletion, whereas the MGS readout and the referred hyperalgesia were not significantly affected. The Y2 receptor antagonist, BII0246, increased pain-related behaviors in response to intrarectal AITC compared to vehicle treatment while the Y2 receptor agonist, PYY(3–36), did not have a significant effect. These results indicate that endogenous PYY has a hypoalgesic effect on somatic thermal and visceral chemical pain. The effect on visceral pain seems to be mediated by peripheral Y2 receptors. PMID:28106168

  8. TRPV1 function is modulated by Cdk5-mediated phosphorylation: insights into the molecular mechanism of nociception.

    PubMed

    Jendryke, Thomas; Prochazkova, Michaela; Hall, Bradford E; Nordmann, Grégory C; Schladt, Moritz; Milenkovic, Vladimir M; Kulkarni, Ashok B; Wetzel, Christian H

    2016-02-23

    TRPV1 is a polymodally activated cation channel acting as key receptor in nociceptive neurons. Its function is strongly affected by kinase-mediated phosphorylation leading to hyperalgesia and allodynia. We present behavioral and molecular data indicating that TRPV1 is strongly modulated by Cdk5-mediated phosphorylation at position threonine-407(mouse)/T406(rat). Increasing or decreasing Cdk5 activity in genetically engineered mice has severe consequences on TRPV1-mediated pain perception leading to altered capsaicin consumption and sensitivity to heat. To understand the molecular and structural/functional consequences of TRPV1 phosphorylation, we generated various rTRPV1T406 receptor variants to mimic phosphorylated or dephosphorylated receptor protein. We performed detailed functional characterization by means of electrophysiological whole-cell and single-channel recordings as well as Ca(2+)-imaging and challenged recombinant rTRPV1 receptors with capsaicin, low pH, or heat. We found that position T406 is critical for the function of TRPV1 by modulating ligand-sensitivity, activation, and desensitization kinetics as well as voltage-dependence. Based on high resolution structures of TRPV1, we discuss T406 being involved in the molecular transition pathway, its phosphorylation leading to a conformational change and influencing the gating of the receptor. Cdk5-mediated phosphorylation of T406 can be regarded as an important molecular switch modulating TRPV1-related behavior and pain sensitivity.

  9. TRPV1 function is modulated by Cdk5-mediated phosphorylation: insights into the molecular mechanism of nociception

    PubMed Central

    Jendryke, Thomas; Prochazkova, Michaela; Hall, Bradford E.; Nordmann, Grégory C.; Schladt, Moritz; Milenkovic, Vladimir M.; Kulkarni, Ashok B.; Wetzel, Christian H.

    2016-01-01

    TRPV1 is a polymodally activated cation channel acting as key receptor in nociceptive neurons. Its function is strongly affected by kinase-mediated phosphorylation leading to hyperalgesia and allodynia. We present behavioral and molecular data indicating that TRPV1 is strongly modulated by Cdk5-mediated phosphorylation at position threonine-407(mouse)/T406(rat). Increasing or decreasing Cdk5 activity in genetically engineered mice has severe consequences on TRPV1-mediated pain perception leading to altered capsaicin consumption and sensitivity to heat. To understand the molecular and structural/functional consequences of TRPV1 phosphorylation, we generated various rTRPV1T406 receptor variants to mimic phosphorylated or dephosphorylated receptor protein. We performed detailed functional characterization by means of electrophysiological whole-cell and single-channel recordings as well as Ca2+-imaging and challenged recombinant rTRPV1 receptors with capsaicin, low pH, or heat. We found that position T406 is critical for the function of TRPV1 by modulating ligand-sensitivity, activation, and desensitization kinetics as well as voltage-dependence. Based on high resolution structures of TRPV1, we discuss T406 being involved in the molecular transition pathway, its phosphorylation leading to a conformational change and influencing the gating of the receptor. Cdk5-mediated phosphorylation of T406 can be regarded as an important molecular switch modulating TRPV1-related behavior and pain sensitivity. PMID:26902776

  10. Intrathecal PKA-selective siRNA treatment blocks sustained morphine-mediated pain sensitization and antinociceptive tolerance in rats.

    PubMed

    Tumati, S; Roeske, W R; Largent-Milnes, T M; Vanderah, T W; Varga, E V

    2011-07-15

    Sustained morphine treatment has been shown to produce paradoxical pain sensitization (opioid-induced hyperalgesia) and also causes increase in spinal pain neurotransmitter, such as calcitonin gene related peptide (CGRP), concentration in experimental animals. Studies have also shown that cyclic adenosine-monophosphate (cAMP)-dependent protein kinase (PKA) plays a major role in the regulation of presynaptic neurotransmitter (such as CGRP and substance P) synthesis and release. We have previously shown that in cultured primary sensory dorsal root ganglion (DRG) neurons sustained in vitro opioid agonist treatment upregulates cAMP levels (adenylyl cyclase (AC) superactivation) and augments basal and capsaicin evoked CGRP release in a PKA dependent manner. In the present study, we investigated the in vivo role of PKA in sustained morphine-mediated pain sensitization. Our data indicate that selective knock-down of spinal PKA activity by intrathecal (i.th.) pretreatment of rats with a PKA-selective small interference RNA (siRNA) mixture significantly attenuates sustained morphine-mediated augmentation of spinal CGRP immunoreactivity, thermal hyperalgesia, mechanical allodynia and antinociceptive tolerance. The present findings indicate that sustained morphine-mediated activation of spinal cAMP/PKA-dependent signaling may play an important role in opioid induced hyperalgesia.

  11. Metamizole (dipyrone) effects on sevoflurane requirements and postoperative hyperalgesia in rats.

    PubMed

    Ruiz-Pérez, Daniel; Benito, Javier; Largo, Carlota; Polo, Gonzalo; Canfrán, Susana; Gómez de Segura, Ignacio Alvarez

    2016-09-30

    Unlike non-steroidal anti-inflammatory drugs (NSAIDs), metamizole has poor anti-inflammatory effects; and is suitable for models where analgesia, but not anti-inflammatory effects, is desirable. Like opioids, these drugs produce perioperative analgesia while reducing anaesthetic requirements, but it remains unclear whether they may develop tolerance or hyperalgesia, and thus decrease in analgesic efficacy. The aim was to determine whether tolerance or hyperalgesia to metamizole occurred in rats, and whether the sevoflurane minimum alveolar concentration (MAC) was affected. In a randomized, prospective, controlled study, male Wistar rats (n = 8 per group) were administered metamizole (300 mg/kg, day 4). Previously, the following treatments were provided: daily metamizole for four days (0-3), morphine (10 mg/kg; positive control, day 0 only) or saline (negative control). The main outcome measures were mechanical (MNT) and warm thermal (WNT) nociceptive quantitative sensory thresholds. The baseline sevoflurane MAC and the reduction produced by the treatments were also determined. The mean (SD) baseline MAC [2.4(0.2)%vol] was decreased by morphine and metamizole by 45(11)% and 33(7)% (P = 0.000, both), respectively. Baseline MNT [35.4(4.5) g] and WNT [13.2(2.4) s] were decreased by morphine and metamizole: MNT reduction of 22(6)% (P = 0.000) and 22(7)% (P = 0.001), respectively and WNT reduction of 34(14)% (P = 0.000) and 24(13)% (P = 0.001). The baseline MAC on day 4 was neither modified by treatments nor the MAC reduction produced by metamizole (days 0 and 4; P > 0.05). In conclusion, repeated metamizole administration may produce hyperalgesia, although it may not modify its anaesthetic sparing effect. The clinical relevance of this effect in painful research models requiring prolonged analgesic therapy warrants further investigation.

  12. Bilateral hand/wrist heat and cold hyperalgesia, but not hypoesthesia, in unilateral carpal tunnel syndrome.

    PubMed

    de la Llave-Rincón, Ana Isabel; Fernández-de-las-Peñas, César; Fernández-Carnero, Josué; Padua, Luca; Arendt-Nielsen, Lars; Pareja, Juan A

    2009-10-01

    The aim of the current study was to evaluate bilaterally warm/cold detection and heat/cold pain thresholds over the hand/wrist in patients with carpal tunnel syndrome (CTS). A total of 25 women with strictly unilateral CTS (mean 42 +/- 10 years), and 20 healthy matched women (mean 41 +/- 8 years) were recruited. Warm/cold detection and heat/cold pain thresholds were assessed bilaterally over the carpal tunnel and the thenar eminence in a blinded design. Self-reported measures included both clinical pain history (intensity, location and area) and Boston Carpal Tunnel Questionnaire. No significant differences between groups for both warm and cold detection thresholds in either carpal tunnel or thenar eminence (P > 0.5) were found. Further, significant differences between groups, but not between sides, for both heat and cold pain thresholds in both the carpal tunnel and thenar eminence were found (all P < 0.001). Heat pain thresholds (P < 0.01) were negatively correlated, whereas cold pain thresholds (P < 0.001) were positively correlated with hand pain intensity and duration of symptoms. Our findings revealed bilateral thermal hyperalgesia (lower heat pain and reduced cold pain thresholds) but not hypoesthesia (normal warm/cold detection thresholds) in patients with strictly unilateral CTS when compared to controls. We suggest that bilateral heat and cold hyperalgesia may reflect impairments in central nociceptive processing in patients with unilateral CTS. The bilateral thermal hyperalgesia associated with pain intensity and duration of pain history supports a role of generalized sensitization mechanisms in the initiation, maintenance and spread of pain in CTS.

  13. Early transcutaneous electrical nerve stimulation reduces hyperalgesia and decreases activation of spinal glial cells in mice with neuropathic pain.

    PubMed

    Matsuo, Hideaki; Uchida, Kenzo; Nakajima, Hideaki; Guerrero, Alexander Rodriguez; Watanabe, Shuji; Takeura, Naoto; Sugita, Daisuke; Shimada, Seiichiro; Nakatsuka, Terumasa; Baba, Hisatoshi

    2014-09-01

    Although transcutaneous electrical nerve stimulation (TENS) is widely used for the treatment of neuropathic pain, its effectiveness and mechanism of action in reducing neuropathic pain remain uncertain. We investigated the effects of early TENS (starting from the day after surgery) in mice with neuropathic pain, on hyperalgesia, glial cell activation, pain transmission neuron sensitization, expression of proinflammatory cytokines, and opioid receptors in the spinal dorsal horn. Following nerve injury, TENS and behavioral tests were performed every day. Immunohistochemical, immunoblot, and flow cytometric analysis of the lumbar spinal cord were performed after 8 days. Early TENS reduced mechanical and thermal hyperalgesia and decreased the activation of microglia and astrocytes (P<0.05). In contrast, the application of TENS at 1 week (TENS-1w) or 2 weeks (TENS-2w) after injury was ineffective in reducing hyperalgesia (mechanical and thermal) or activation of microglia and astrocytes. Early TENS decreased p-p38 within microglia (P<0.05), the expression levels of protein kinase C (PKC-γ), and phosphorylated anti-phospho-cyclic AMP response element-binding protein (p-CREB) in the superficial spinal dorsal horn neurons (P<0.05), mitogen-activated protein (MAP) kinases, and proinflammatory cytokines, and increased the expression levels of opioid receptors (P<0.05). The results suggested that the application of early TENS relieved hyperalgesia in our mouse model of neuropathic pain by inhibiting glial activation, MAP kinase activation, PKC-γ, and p-CREB expression, and proinflammatory cytokines expression, as well as maintenance of spinal opioid receptors. The findings indicate that TENS treatment is more effective when applied as early after nerve injury as possible.

  14. Leishmania (L). amazonensis induces hyperalgesia in balb/c mice: Contribution of endogenous spinal cord TNFα and NFκB activation.

    PubMed

    Borghi, Sergio M; Fattori, Victor; Ruiz-Miyazawa, Kenji W; Miranda-Sapla, Milena M; Casagrande, Rúbia; Pinge-Filho, Phileno; Pavanelli, Wander R; Verri, Waldiceu A

    2017-02-17

    Cutaneous leishmaniasis (CL) is the most common form of the leishmaniasis in humans. Ulcerative painless skin lesions are predominant clinical features of CL. Wider data indicate pain accompanies human leishmaniasis, out with areas of painless ulcerative lesions per se. In rodents, Leishmania (L.) major infection induces nociceptive behaviors that correlate with peripheral cytokine levels. However, the role of the spinal cord in pain processing after Leishmania infection has not been investigated. Balb/c mice received intraplantar (i.pl.) injection of Leishmania (L). amazonensis and hyperalgesia, edema, parasitism, and spinal cord TNFα, TNFR1 and TNFR2 mRNA expression, and NFκB activation were evaluated. The effects of intrathecal (i.t.) injection of morphine, TNFα, TNFα inhibitors (etanercept and adalimumab) and NFκB inhibitor (PDTC) were investigated. The present study demonstrates that Leishmania (L.) amazonensis infection in balb/c mice induces chronic mechanical and thermal hyperalgesia in an opioid-sensitive manner. Spinal cord TNFα mRNA expression increased in a time-dependent manner, peaking between 30 and 40 days after infection. At the peak of TNFα mRNA expression (day 30), there was a concomitant increase in TNFR1 and TNFR2 mRNA expression. TNFα i.t. injection enhanced L. (L.) amazonensis-induced hyperalgesia. Corroborating a role for TNFα in L. (L.) amazonensis-induced hyperalgesia, i.t. treatment with the TNFα inhibitors, etanercept and adalimumab inhibited the hyperalgesia. L. (L.) amazonensis also induced spinal cord activation of NFκB, and PDTC (given i.t.), also inhibited L. (L.) amazonensis-induced hyperalgesia, and spinal cord TNFα, TNFR1 and TNFR2 mRNA expression. Moreover, L. (L.) amazonensis-induced spinal cord activation of NFκB was also inhibited by etanercept and adalimumab as well as PDTC i.t.

  15. Molecular adsorption at particle surfaces: a PM toxicity mediation mechanism.

    PubMed

    Kendall, Michaela; Brown, Leslie; Trought, Katherine

    2004-01-01

    surfactant was added at a fixed concentration lower than physiological lung lavage concentrations to ensure the lipid remained in suspension during experimentation ex situ. For dipalmitoylphosphatidylcholine (DPPC) combinations with particles, visible particle agglomeration occurred within 1 h. Marked changes in the size distribution of the immersed particles were observed, compared to a phosphate buffer control. Differences in particle agglomeration and particle settling were observed between M120, R330, and R400. Reduction of DPPC occurred in a surface- and size-dependent manner. This indicates that surface adsorption was responsible for the observed agglomeration and the gross reductions in phospholipid concentrations. Combination of particles with fibrinogen and albumin revealed little agglomeration/precipitation at the protein concentrations chosen. However, surfactant protein (SP-D) was completely eliminated from suspension upon combination with all three-particle types. This reaction between SP-D particles was therefore concluded to be independent of surface chemistry. Further investigation as to whether this is size- or surface-area-dependent is recommended. The biological implication is that molecular adsorption at nonbiological particulate matter (PM) surfaces in BALF may mediate the toxicity of PM via one or both of these mechanisms, as in the case of biological particles.

  16. Vibration-mediated Kondo transport in molecular junctions: conductance evolution during mechanical stretching

    PubMed Central

    Rakhmilevitch, David

    2015-01-01

    Summary The vibration-mediated Kondo effect attracted considerable theoretical interest during the last decade. However, due to lack of extensive experimental demonstrations, the fine details of the phenomenon were not addressed. Here, we analyze the evolution of vibration-mediated Kondo effect in molecular junctions during mechanical stretching. The described analysis reveals the different contributions of Kondo and inelastic transport. PMID:26734532

  17. Cellular Mechanisms of Calcium-Mediated Triggered Activity

    NASA Astrophysics Data System (ADS)

    Song, Zhen

    Life-threatening cardiac arrhythmias continue to pose a major health problem. Ventricular fibrillation, which is a complex form of electrical wave turbulence in the lower chambers of the heart, stops the heart from pumping and is the largest cause of natural death in the United States. Atrial fibrillation, a related form of wave turbulence in the upper heart chambers, is in turn the most common arrhythmia diagnosed in clinical practice. Despite extensive research to date, mechanisms of cardiac arrhythmias remain poorly understood. It is well established that both spatial disorder of the refractory period of heart cells and triggered activity (TA) jointly contribute to the initiation and maintenance of arrhythmias. TA broadly refers to the abnormal generation of a single or a sequence of abnormal excitation waves from a small submillimeter region of the heart in the interval of time between two normal waves generated by the heart's natural pacemaker (the sinoatrial node). TA has been widely investigated experimentally and occurs in several pathological conditions where the intracellular concentration of free Ca2+ ions in heart cells becomes elevated. Under such conditions, Ca2+ can be spontaneously released from intracellular stores, thereby driving an electrogenic current that exchanges 3Na+ ions for one Ca2+ ion across the cell membrane. This current in turn depolarizes the membrane of heart cells after a normal excitation. If this calcium-mediated "delayed after depolarization'' (DAD) is sufficiently large, it can generate an action potential. While the arrhythmogenic importance of spontaneous Ca2+ release and DADs is well appreciated, the conditions under which they occur in heart pathologies remain poorly understood. Calcium overload is only one factor among several other factors that can promote DADs, including sympathetic nerve stimulation, different expression levels of membrane ion channels and calcium handling proteins, and different mutations of those

  18. SHANK3 Deficiency Impairs Heat Hyperalgesia and TRPV1 Signaling in Primary Sensory Neurons.

    PubMed

    Han, Qingjian; Kim, Yong Ho; Wang, Xiaoming; Liu, Di; Zhang, Zhi-Jun; Bey, Alexandra L; Lay, Mark; Chang, Wonseok; Berta, Temugin; Zhang, Yan; Jiang, Yong-Hui; Ji, Ru-Rong

    2016-12-21

    Abnormal pain sensitivity is commonly associated with autism spectrum disorders (ASDs) and affects the life quality of ASD individuals. SHANK3 deficiency was implicated in ASD and pain dysregulation. Here, we report functional expression of SHANK3 in mouse dorsal root ganglion (DRG) sensory neurons and spinal cord presynaptic terminals. Homozygous and heterozygous Shank3 complete knockout (Δe4-22) results in impaired heat hyperalgesia in inflammatory and neuropathic pain. Specific deletion of Shank3 in Nav1.8-expressing sensory neurons also impairs heat hyperalgesia in homozygous and heterozygous mice. SHANK3 interacts with transient receptor potential subtype V1 (TRPV1) via Proline-rich region and regulates TRPV1 surface expression. Furthermore, capsaicin-induced spontaneous pain, inward currents in DRG neurons, and synaptic currents in spinal cord neurons are all reduced after Shank3 haploinsufficiency. Finally, partial knockdown of SHANK3 expression in human DRG neurons abrogates TRPV1 function. Our findings reveal a peripheral mechanism of SHANK3, which may underlie pain deficits in SHANK3-related ASDs.

  19. The C-terminus of murine S100A9 protein inhibits hyperalgesia induced by the agonist peptide of protease-activated receptor 2 (PAR2)

    PubMed Central

    Dale, C S; Cenac, N; Britto, L R G; Juliano, M A; Juliano, L; Vergnolle, N; Giorgi, R

    2006-01-01

    Background and purpose: S100A9 protein induces anti-nociception in rodents, in different experimental models of inflammatory pain. Herein, we investigated the effects of a fragment of the C-terminus of S100A9 (mS100A9p), on the hyperalgesia induced by serine proteases, through the activation of protease-activated receptor-2 (PAR2). Experimental approach: Mechanical and thermal hyperalgesia induced by PAR2 agonists (SLIGRL-NH2 and trypsin) was measured in rats submitted to the paw pressure or plantar tests, and Egr-1 expression was determined by immunohistochemistry in rat spinal cord dorsal horn. Calcium flux in human embryonic kidney cells (HEK), which naturally express PAR2, in Kirsten virus-transformed kidney cells, transfected (KNRK-PAR2) or not (KNRK) with PAR2, and in mouse dorsal root ganglia neurons (DRG) was measured by fluorimetric methods. Key results: mS100A9p inhibited mechanical hyperalgesia induced by trypsin, without modifying its enzymatic activity. Mechanical and thermal hyperalgesia induced by SLIGRL-NH2 were inhibited by mS100A9p. SLIGRL-NH2 enhanced Egr-1 expression, a marker of nociceptor activation, and this effect was inhibited by concomitant treatment with mS100A9p. mS100A9p inhibited calcium mobilization in DRG neurons in response to the PAR2 agonists trypsin and SLIGRL-NH2, but also in response to capsaicin and bradykinin, suggesting a direct effect of mS100A9 on sensory neurons. No effect on the calcium flux induced by trypsin or SLIGRL in HEK cells or KNRK-PAR2 cells was observed. Conclusions and implications: These data demonstrate that mS100A9p interferes with mechanisms involved in nociception and hyperalgesia and modulates, possibly directly on sensory neurons, the PAR2-induced nociceptive signal. PMID:16967049

  20. Effects of Different Electroacupuncture Scheduling Regimens on Murine Bone Tumor-Induced Hyperalgesia: Sex Differences and Role of Inflammation

    PubMed Central

    Smeester, Branden A.; Al-Gizawiy, Mona; Beitz, Alvin J.

    2012-01-01

    Previous studies have shown that electroacupuncture (EA) is able to reduce hyperalgesia in rodent models of persistent pain, but very little is known about the analgesic effects and potential sex differences of different EA treatment regimens. In the present study, we examined the effects of five different EA treatments on tumor-induced hyperalgesia in male and female mice. EA applied to the ST-36 acupoint either twice weekly (EA-2X/3) beginning on postimplantation day (PID) 3 or prophylactically three times prior to implantation produced the most robust and longest lasting antinociceptive effects. EA treatment given once per week beginning at PID 7 only produced an antinociceptive effect in female animals. The analgesic effect of EA-2X/3 began earlier in males, but lasted longer in females indicating sex differences in EA. We further demonstrate that EA-2X/3 elicits a marked decrease in tumor-associated inflammation as evidenced by a significant reduction in tumor-associated neutrophils at PID 7. Moreover, EA-2X/3 produced a significant reduction in tumor-associated PGE2 as measured in microperfusate samples. Collectively, these data provide evidence that EA-2X/3 treatment reduces tumor-induced hyperalgesia, which is associated with a decrease in tumor-associated inflammation and PGE2 concentration at the tumor site suggesting possible mechanisms by which EA reduces tumor nociception. PMID:23320035

  1. Exploration of a Novel Persistent Reversal of Pathological Pain: Mechanisms and Mediators

    DTIC Science & Technology

    2015-04-01

    Award Number: W81XWH-11-2-0070 TITLE: Exploration of a Novel Persistent Reversal of Pathological Pain : Mechanisms and Mediators PRINCIPAL...Jan 2015 4. TITLE AND SUBTITLE Exploration of a Novel Persistent Reversal of Pathological Pain : Mechanisms and Mediators 5a. CONTRACT NUMBER 5b...Approved for Public Release; Distribution Unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT Neuropathic pain , resulting from nerve injury or inflammation

  2. Pharmacokinetics of Lipophilic Agents Following Preexposure: Non-Cytochrome P-450 Mediated Mechanisms

    DTIC Science & Technology

    1990-05-30

    FOLLOWING PREEXPOSUR G - AFOSR-87- 185 U’) NON -CYTOCHROME P-450 MEDIATED MECHAN’ICS PE - 61102F (NJ . AUTHOKS) PR - 2312 N TA - A5 Dr Lawrence R Curtis, Dr...Z39- I*. PHARMACOKINETICS OF LIPOPHILIC AGENTS FOLLOWING PREEXPOSURE: NON -CYTOCHROME P-450 MEDIATED MECHANISMS Air Force Grant No. 87-0185 Hillary M...amounts located in non -hepatic tissues. These studies showed that, despite the differences in PDR, the systems involved respond in a dose related manner

  3. Vacancy Mediated Mechanism of Nitrogen Substitution in Carbon Nanotubes

    NASA Technical Reports Server (NTRS)

    Srivastava, Deepak; Menon, Madhu; Sadanadan, Bindu; Rao, Apparao M.

    2003-01-01

    Nitrogen substitution reaction in a graphene sheet and carbon nanotubes of different diameter are investigated using the generalized tight-binding molecular dynamics method. The formation of a vacancy in curved graphene sheet or a carbon nanotube is found to cause a curvature dependent local reconstruction of the surface. Our simulations and analysis show that vacancy mediated N substitution (rather than N chemisorption) is favored on the surface of nanotubes with diameter larger than 8 nm. This predicted value of the critical minimum diameter for N incorporation is confirmed by experimental results presented.

  4. Hydrogen-rich saline prevents remifentanil-induced hyperalgesia and inhibits MnSOD nitration via regulation of NR2B-containing NMDA receptor in rats.

    PubMed

    Zhang, L; Shu, R; Wang, H; Yu, Y; Wang, C; Yang, M; Wang, M; Wang, G

    2014-11-07

    Remifentanil administration may subsequently cause paradoxical hyperalgesia in animals and humans, but mechanisms remain unclear. Manganese superoxide dismutase (MnSOD) nitration and inactivation caused by generation of reactive oxygen species and activation of N-methyl-D-aspartate (NMDA) receptors are involved in the induction and maintenance of central neuropathic pain. Hydrogen which selectively removes superoxide has gained much attention in recent years. In this study, we investigated antinociceptive effects of hydrogen-rich saline (HRS) on remifentanil-induced postsurgical hyperalgesia in a rat model of incisional pain. HRS was injected intraperitoneally 10 min before remifentanil infusion (1 μg kg(-1) min(-1) for 60 min). A selective NR2B antagonist Ro25-6981 was used to investigate whether antihypernociception of HRS is associated with NMDA receptor (NMDAR). Nociception was evaluated by the paw withdrawal mechanical threshold and thermal latency respectively. Then we assessed MnSOD, NR2A and NR2B in spinal cord dorsal horn via Western blot and immunohistochemistry after nociceptive tests. Here, we found that the analgesic effect of remifentanil was followed by long-term hyperalgesia lasting at least postoperative 7 days, which was accompanied with increase in NR2B expression and trafficking from cytoplasm to surface and MnSOD nitration in dorsal horn. Pretreatment with HRS (10 ml/kg) significantly attenuated mechanical and thermal hyperalgesia, blocked NR2B trafficking and MnSOD nitration in dorsal horn after remifentanil infusion. Ro25-6981 not 5 μg but 10 and 50 μg dosage-dependently attenuated hyperalgesia, and inhibited MnSOD nitration. Hyperalgesia and MnSOD nitration were attenuated after the combination of HRS (2.5 ml/kg) and Ro25-6981 (5 μg). In conclusion, HRS (10 ml/kg) might reverse remifentanil-induced hyperalgesia, through regulating NR2B-containing NMDAR trafficking to control MnSOD nitration and enhance MnSOD activity.

  5. Mechanisms of uric acid crystal-mediated autoinflammation.

    PubMed

    Martinon, Fabio

    2010-01-01

    Gout is an arthritis characterized by elevated uric acid in the bloodstream. In this condition, crystals of uric acid are formed and accumulate in the synovial fluids. Crystal deposition leads to acute inflammation, which is associated with the spontaneous resolution of the disease. Recent studies have led to significant advances in the understanding of the basic biology of crystal-mediated inflammation. Uric acid has been identified as a danger signal that triggers a cytosolic sensor, the inflammasome. This signaling platform is required for the activation of interleukin-1, a cytokine that is critical to the initiation of acute inflammation in gout. Importantly, both molecular and pathological evidence support the notion that gout is a prototypical member of the growing family of autoinflammatory diseases. This review discusses the role of the inflammasome in gout and the emerging new therapeutic strategies aimed at controlling inflammation in crystal arthritis.

  6. A cascade of cytokines mediates mechanical inflammatory hypernociception in mice

    PubMed Central

    Cunha, T. M.; Verri, W. A.; Silva, J. S.; Poole, S.; Cunha, F. Q.; Ferreira, S. H.

    2005-01-01

    The hypernociceptive effects of cytokines [TNF-α, keratinocyte-derived chemokine (KC), and IL-1β] and their participation in carrageenan (Cg)-induced inflammatory hypernociception in mice were investigated. Nociceptor sensitization (hypernociception) was quantified with an electronic version of the von Frey filament test in WT and TNF receptor type 1 knockout mice (TNF-R1–/–). TNF-α-induced hypernociception was abolished in TNF-R1–/– mice, partially inhibited by pretreatment with IL-1 receptor antagonist (IL-1ra) or indomethacin and unaffected by Ab against KC (AbKC) or guanethidine. IL-1ra and indomethacin pretreatment strongly inhibited the hypernociception induced by IL-1β, which was not altered by AbKC or guanethidine or by knocking out TNF-R1. KC-induced hypernociception was abolished by AbKC, inhibited by pretreatment with indomethacin plus guanethidine, and partially inhibited by IL-1ra, indomethacin, or guanethidine. In contrast, KC-induced hypernociception was not altered by knocking out TNF-R1. Cg-induced hypernociception was abolished by administration of indomethacin plus guanethidine, diminished in TNF-R1–/– mice, and partially inhibited in WT mice pretreated with AbKC, IL-1ra, indomethacin, or guanethidine. TNF-α, KC, and IL-1β concentrations were elevated in the skin of Cg-injected paws. The TNF-α and KC concentrations rose concomitantly and peaked before that of IL-1β. In mice, the cytokine cascade begins with the release of TNF-α (acting on TNF-R1 receptor) and KC, which stimulate the release of IL-1β. As in rats, the final mediators of this cascade were prostaglandins released by IL-1β and sympathetic amines released by KC. These results extend to mice the concept that the release of primary mediators responsible for hypernociception is preceded by a cascade of cytokines. PMID:15665080

  7. Genetic Mechanisms in Apc-Mediated Mammary Tumorigenesis

    PubMed Central

    Kuraguchi, Mari; Ohene-Baah, Nana Yaw; Sonkin, Dmitriy; Bronson, Roderick Terry; Kucherlapati, Raju

    2009-01-01

    Many components of Wnt/β-catenin signaling pathway also play critical roles in mammary tumor development, yet the role of the tumor suppressor gene APC (adenomatous polyposis coli) in breast oncongenesis is unclear. To better understand the role of Apc in mammary tumorigenesis, we introduced conditional Apc mutations specifically into two different mammary epithelial populations using K14-cre and WAP-cre transgenic mice that express Cre-recombinase in mammary progenitor cells and lactating luminal cells, respectively. Only the K14-cre–mediated Apc heterozygosity developed mammary adenocarcinomas demonstrating histological heterogeneity, suggesting the multilineage progenitor cell origin of these tumors. These tumors harbored truncation mutation in a defined region in the remaining wild-type allele of Apc that would retain some down-regulating activity of β-catenin signaling. Activating mutations at codons 12 and 61 of either H-Ras or K-Ras were also found in a subset of these tumors. Expression profiles of acinar-type mammary tumors from K14-cre; ApcCKO/+ mice showed luminal epithelial gene expression pattern, and clustering analysis demonstrated more correlation to MMTV-neu model than to MMTV-Wnt1. In contrast, neither WAP-cre–induced Apc heterozygous nor homozygous mutations resulted in predisposition to mammary tumorigenesis, although WAP-cre–mediated Apc deficiency resulted in severe squamous metaplasia of mammary glands. Collectively, our results suggest that not only the epithelial origin but also a certain Apc mutations are selected to achieve a specific level of β-catenin signaling optimal for mammary tumor development and explain partially the colon- but not mammary-specific tumor development in patients that carry germline mutations in APC. PMID:19197353

  8. Kinin B(1) and B(2) receptors contribute to orofacial heat hyperalgesia induced by infraorbital nerve constriction injury in mice and rats.

    PubMed

    Luiz, Ana Paula; Schroeder, Samilla Driessen; Chichorro, Juliana Geremias; Calixto, João Batista; Zampronio, Aleksander Roberto; Rae, Giles Alexander

    2010-04-01

    Mechanisms coupled to kinin B(1) and B(2) receptors have been implicated in sensory changes associated to various models of neuropathy. The current study aimed to investigate if kinins also participate in orofacial thermal hyperalgesia induced by constriction of the infraorbital nerve (CION), a model of trigeminal neuropathic pain which displays persistent hypersensitivity to orofacial sensory stimulation, in rats and mice. Male Swiss mice (30-35g) or Wistar rats (200-250g; n=6-10 per group in both cases) underwent CION or sham surgery and were submitted repeatedly to application of heat ( approximately 50 degrees C) to the ipsilateral or contralateral snout, delivered by a heat source placed 1cm from the vibrissal pad. Decreases in latency to display head withdrawal or vigorous snout flicking were considered indicative of heat hyperalgesia. CION caused long-lasting heat hyperalgesia which started on Day 2 after surgery in both species and lasted up to Day 17 in mice and Day 10 in rats. Administration of DALBK or HOE-140 (peptidic B(1) and B(2) receptor antagonists, respectively; each at 3nmol in 10microl) onto the exposed infraorbital nerve of mice at the moment of surgery delayed the development of the thermal hyperalgesia. Systemic treatment on Day 5 (mice) or Day 4 (rats) with Des-Arg(9), Leu(8)-Bradykinin (DALBK, B(1) receptor antagonist, 0.1-1micromol/kg, i.p.) or HOE-140 (B(2) receptor antagonist, 0.001-1micromol/kg, i.p.) transiently reduced heat hyperalgesia in both species. Due to the peptidic nature of DALBK and HOE-140, it is likely that their effects reported herein resulted from blockade of peripheral kinin receptors. Thus, mechanisms operated by kinin B(1) and B(2) receptors, contribute to orofacial heat hyperalgesia induced by CION in both mice and rats. Perhaps kinin B(1) and B(2) receptor antagonists might constitute effective preventive and curative treatments for orofacial thermal hyperalgesia induced by nerve injury.

  9. Chirality-Mediated Mechanical and Structural Properties of Oligopeptide Hydrogels

    SciTech Connect

    Taraban, Marc B.; Feng, Yue; Hammouda, Boualem; Hyland, Laura L.; Yu, Y. Bruce

    2012-10-29

    The origin and the effects of homochirality in the biological world continuously stimulate numerous hypotheses and much debate. This work attempts to look at the biohomochirality issue from a different angle - the mechanical properties of the bulk biomaterial and their relation to nanoscale structures. Using a pair of oppositely charged peptides that co-assemble into hydrogels, we systematically investigated the effect of chirality on the mechanical properties of these hydrogels through different combinations of syndiotactic and isotactic peptides. It was found that homochirality confers mechanical advantage, resulting in a higher elastic modulus and strain yield value. Yet, heterochirality confers kinetic advantage, resulting in faster gelation. Structurally, both homochiral and heterochiral hydrogels are made of fibers interconnected by lappet-like webs, but the homochiral peptide fibers are thicker and denser. These results highlight the possible role of biohomochirality in the evolution and/or natural selection of biomaterials.

  10. Signaling mechanisms that mediate invasion in prostate cancer cells.

    PubMed

    Bonaccorsi, L; Marchiani, S; Muratori, M; Carloni, V; Forti, G; Baldi, E

    2004-12-01

    Recent evidence indicates that androgen-sensitive prostate cancer cells have a less malignant phenotype characterized by reduced migration and invasion. We investigated whether the presence of the androgen receptor could affect EGFR-mediated signaling by evaluating autotransphosphorylation of the receptor as well as activation of the downstream signaling pathway PI3K/AKT. Immunoprecipitation studies demonstrated a reduction of EGF-induced tyrosine phosphorylation of EGFR in PC3-AR cells. In addition, EGF-stimulated PI3K activity, a key signaling pathway for invasion of these cells, was decreased in PC3-AR cells and further reduced by treatment with R1881, indicating decreased functionality of EGFR. Our results suggest that the expression of androgen receptors by transfection in PC3 cells confers a less malignant phenotype by interfering with EGFR autophosphorylation and signaling leading to invasion in response to EGF. We used the selective tyrosine kinase inhibitor of the EGFR gefitinib (also known as Iressa or ZD1839) to further investigate the role of EGFR in the invasion and growth of PC cells. We demonstrate that in the androgen-insensitive cell lines PC3 and DU145 this compound was able to decrease in vitro invasion of Matrigel by inhibiting EGFR autotransphosphorylation and subsequent PI3K activation. Gefitinib may be useful in the treatment of androgen-independent prostate cancer to limit not only the proliferation but also the invasion of these tumors.

  11. Mechanism for Hypocretin-mediated sleep-to-wake transitions.

    PubMed

    Carter, Matthew E; Brill, Julia; Bonnavion, Patricia; Huguenard, John R; Huerta, Ramon; de Lecea, Luis

    2012-09-25

    Current models of sleep/wake regulation posit that Hypocretin (Hcrt)-expressing neurons in the lateral hypothalamus promote and stabilize wakefulness by projecting to subcortical arousal centers. However, the critical downstream effectors of Hcrt neurons are unknown. Here we use optogenetic, pharmacological, and computational tools to investigate the functional connectivity between Hcrt neurons and downstream noradrenergic neurons in the locus coeruleus (LC) during nonrapid eye movement (NREM) sleep. We found that photoinhibiting LC neurons during Hcrt stimulation blocked Hcrt-mediated sleep-to-wake transitions. In contrast, when LC neurons were optically stimulated to increase membrane excitability, concomitant photostimulation of Hcrt neurons significantly increased the probability of sleep-to-wake transitions compared with Hcrt stimulation alone. We also built a conductance-based computational model of Hcrt-LC circuitry that recapitulates our behavioral results using LC neurons as the main effectors of Hcrt signaling. These results establish the Hcrt-LC connection as a critical integrator-effector circuit that regulates NREM sleep/wake behavior during the inactive period. This coupling of distinct neuronal systems can be generalized to other hypothalamic integrator nuclei with downstream effector/output populations in the brain.

  12. Mechanism of human antibody-mediated neutralization of Marburg virus.

    PubMed

    Flyak, Andrew I; Ilinykh, Philipp A; Murin, Charles D; Garron, Tania; Shen, Xiaoli; Fusco, Marnie L; Hashiguchi, Takao; Bornholdt, Zachary A; Slaughter, James C; Sapparapu, Gopal; Klages, Curtis; Ksiazek, Thomas G; Ward, Andrew B; Saphire, Erica Ollmann; Bukreyev, Alexander; Crowe, James E

    2015-02-26

    The mechanisms by which neutralizing antibodies inhibit Marburg virus (MARV) are not known. We isolated a panel of neutralizing antibodies from a human MARV survivor that bind to MARV glycoprotein (GP) and compete for binding to a single major antigenic site. Remarkably, several of the antibodies also bind to Ebola virus (EBOV) GP. Single-particle EM structures of antibody-GP complexes reveal that all of the neutralizing antibodies bind to MARV GP at or near the predicted region of the receptor-binding site. The presence of the glycan cap or mucin-like domain blocks binding of neutralizing antibodies to EBOV GP, but not to MARV GP. The data suggest that MARV-neutralizing antibodies inhibit virus by binding to infectious virions at the exposed MARV receptor-binding site, revealing a mechanism of filovirus inhibition.

  13. Mechanism of Human Antibody-Mediated Neutralization of Marburg Virus

    PubMed Central

    Flyak, Andrew I.; Ilinykh, Philipp A.; Murin, Charles D.; Garron, Tania; Shen, Xiaoli; Fusco, Marnie L.; Hashiguchi, Takao; Bornholdt, Zachary A.; Slaughter, James C.; Sapparapu, Gopal; Klages, Curtis; Ksiazek, Thomas G.; Ward, Andrew B.; Saphire, Erica Ollmann; Bukreyev, Alexander; Crowe, James E.

    2015-01-01

    Summary The mechanisms by which neutralizing antibodies inhibit Marburg virus (MARV) are not known. We isolated a panel of neutralizing antibodies from a human MARV survivor that bind to MARV glycoprotein (GP) and compete for binding to a single major antigenic site. Remarkably, several of the antibodies also bind to Ebola virus (EBOV) GP. Single-particle EM structures of Antibody-GP complexes reveals that all of the neutralizing antibodies bind to MARV GP at or near the predicted region of the receptor-binding site. The presence of the glycan cap or mucin-like domain blocks binding of neutralizing antibodies to EBOV GP but not to MARV GP. The data suggest that MARV neutralizing antibodies inhibit virus by binding to infectious virions at the exposed MARV receptor-binding site, revealing a mechanism of filovirus inhibition. PMID:25723164

  14. Sigma-1 receptor-mediated increase in spinal p38 MAPK phosphorylation leads to the induction of mechanical allodynia in mice and neuropathic rats.

    PubMed

    Moon, Ji-Young; Roh, Dae-Hyun; Yoon, Seo-Yeon; Kang, Suk-Yun; Choi, Sheu-Ran; Kwon, Soon-Gu; Choi, Hoon-Seong; Han, Ho-Jae; Beitz, Alvin J; Lee, Jang-Hern

    2013-09-01

    The direct activation of the spinal sigma-1 receptor (Sig-1R) produces mechanical allodynia (MA) and thermal hyperalgesia (TH) in mice. In addition, the blockade of the spinal Sig-1R prevents the induction of MA, but not TH in chronic constriction injury (CCI)-induced neuropathic rats. The present study was designed to investigate whether the increase in spinal p38 MAPK phosphorylation (p-p38 MAPK) mediates Sig-1R-induced MA or TH in mice and the induction of MA in neuropathic rats. MA and TH were evaluated using von Frey filaments and a hot-plate apparatus, respectively. Neuropathic pain was produced by CCI of the right sciatic nerve in rats. Western blot assay and immunohistochemistry were performed to determine the changes of p-p38 MAPK expression in the spinal cord. Intrathecal (i.t.) injection of PRE084, a selective Sig-1R agonist, into naïve mice time-dependently increased the expression of p-p38 MAPK, which was blocked by pretreatment with BD1047, a Sig-1R antagonist. I.t. pretreatment with SB203580, a p38 MAPK inhibitor also dose-dependently inhibited PRE084-induced MA, whereas TH induction was not affected. In CCI rats, i.t. injection of BD1047 during the induction phase (postoperative days 0 to 5) reduced the CCI-induced increase in p-p38 MAPK. In addition, i.t. SB203580 treatment during the induction phase also suppressed the development of CCI-induced MA, but not TH. Conversely, i.t. SB203580 treatment during the maintenance phase (postoperative days 15 to 20) had no effect on CCI-induced MA or TH. These results demonstrate that the increase in spinal p-p38 MAPK is closely associated with the induction of Sig-1R mediated MA, but not TH. Sigma-1 receptor modulation of p-p38 MAPK also plays an important role in the induction, but not the maintenance, of MA in neuropathic pain.

  15. Caloric restriction as a mechanism mediating resistance to environmental disease.

    PubMed Central

    Frame, L T; Hart, R W; Leakey, J E

    1998-01-01

    It has been observed that susceptibility to many degenerative diseases increases concurrently with industrialization and rising living standards. Although epidemiologic studies suggest that specific environmental and dietary factors may be important, caloric intake alone (as reflected in body size) may account for much of the differential risk observed among diverse human populations. It has been suggested from animal studies that caloric intake may be the primary effector for many hormonal, metabolic, physiologic, and behavioral responses that coordinate reproductive strategy to apparent availability of food. When caloric intake is excessive, particularly at critical developmental stages, physiologic priorities are set for body growth and fecundity rather than for endurance and longevity. The converse occurs during periods of famine, thus increasing the probability that sufficient individuals survive to restore the population when conditions improve. Calorically restricted rodents have significantly longer reproductive and total life spans than their ad libitum-fed controls and exhibit a spectrum of biochemical and physiologic alterations that characterize their adaptation to reduced intake. These include reduced stature, hypercorticism in the absence of elevated adrenocorticotropic hormone levels, increased metabolic efficiency, decreased mitogenic response coupled with increased rates of apoptosis, reduced inflammatory response, induction of stress proteins and DNA repair enzymes, altered drug-metabolizing enzyme expression, and modified cell-mediated immune function. The overall profile of these changes is one of improved defense against environmental stress. This has been suggested as the mechanistic basis for the protective effects of low body weight on radiation and chemically induced cancers in experimental animals. It may also explain the significantly higher thresholds of acute toxicity observed when calorically restricted rodents are exposed to certain

  16. Venlafaxine Attenuates Heat Hyperalgesia Independent of Adenosine or Opioid System in a Rat Model of Peripheral Neuropathy

    PubMed Central

    Abed, Alireza; Hajhashemi, Valiollah; Banafshe, Hamid Reza; Minaiyan, Mohsen; Mesdaghinia, Azam

    2015-01-01

    Primarily opioidergic and adenosine mechanisms are considered to be involved in the antinociceptive effects of antidepressants. This study was designed to determine the efficacy of acute venlafaxine administration in alleviating symptoms of neuropathic pain and the role of endogenous adenosine and opioid systems in this effect of venlafaxine. We have evaluated the effect of caffeine, a non-selective adenosine A1 and A2 receptor antagonist and naloxone as an antagonist of opioid receptors on the antinociceptive effects of venlafaxine. Chronic constriction injury of the sciatic nerve resulted in thermal hyperalgesia, mechanical and cold allodynia in the rats. Animals were received on the 7th day after surgery, when the model had been fully established, venlafaxine (20 and 40 mg/Kg i.p.), or venlafaxine (40 mg/Kg) in combination with caffeine (5 mg/Kg i.p.) or naloxone (1 mg/Kg s.c.). Rats were tested for thermal reaction latencies, mechanical and cold allodynia 45 min after drug injection. Acute venlafaxine (40 mg/Kg i.p.) administration consistently decreased the thermal hyperalgesia and this effect was not blocked by concomitant caffeine or naloxone administration. There was no effect by either drug or the drug combination on the tactile and cold allodynia. The results of this study indicate that venlafaxine (40 mg/Kg i.p.) is effective in alleviating thermal hyperalgesia and this effect is independent through manipulation of adenosine or opioid system. This observation demonstrates that venlafaxine, which is a mixed inhibitor of norepinephrine and serotonin reuptake, differs from the other antidepressants in the mechanism of its antinociception action. PMID:26330872

  17. Mechanisms of abscisic acid-mediated control of stomatal aperture.

    PubMed

    Munemasa, Shintaro; Hauser, Felix; Park, Jiyoung; Waadt, Rainer; Brandt, Benjamin; Schroeder, Julian I

    2015-12-01

    Drought stress triggers an increase in the level of the plant hormone abscisic acid (ABA), which initiates a signaling cascade to close stomata and reduce water loss. Recent studies have revealed that guard cells control cytosolic ABA concentration through the concerted actions of biosynthesis, catabolism as well as transport across membranes. Substantial progress has been made at understanding the molecular mechanisms of how the ABA signaling core module controls the activity of anion channels and thereby stomatal aperture. In this review, we focus on our current mechanistic understanding of ABA signaling in guard cells including the role of the second messenger Ca(2+) as well as crosstalk with biotic stress responses.

  18. Repeated 100 Hz TENS for the Treatment of Chronic Inflammatory Hyperalgesia and Suppression of Spinal Release of Substance P in Monoarthritic Rats

    PubMed Central

    Liu, Hong-Xiang; Tian, Jin-Bin; Luo, Fei; Jiang, Yu-Hui; Deng, Zu-Guo; Xiong, Liang; Liu, Cheng; Wang, Jin-Shu

    2007-01-01

    Transcutaneous electrical nerve stimulation (TENS) has been shown to be an effective measure for pain relief. The aim of the present study was to determine the optimal intensity and interval of repeated 100 Hz TENS for the treatment of chronic inflammatory hyperalgesia in a monoarthritic pain model of the rat, and to assess the changes of the spinal substance P (SP) release in response to TENS treatment. A reliable, reproducible chronic monoarthritic pain model was produced by intra-articular injection of complete Freund's adjuvant (CFA) at single ankle joint. The efficacy of 100 Hz TENS treatments with different frequencies and intensities was compared. In the acute period (within 3 weeks) of monoarthritis, twice-a-week schedule of TENS reduced the swelling of the inflamed ankle significantly. In the stable period (4–9 weeks), however, once-a-week schedule produced a significantly better therapeutic effect on both inflammation and arthritic hyperalgesia than that of twice- or five-times-a-week schedule. Using three levels of intensity of TENS, we found that the weaker (1-1-2 mA) stimulation produced significantly better therapeutic effects. Repeated TENS produced a reduction of SP content in spinal perfusate in parallel with the progressive reduction of the arthritic pain scores. Our results suggest that (i) consecutive TENS treatments produced cumulative effect for chronic hyperalgesia, (ii) for chronic inflammatory hyperalgesia, a weaker intensity and more sparsely arranged treatment schedule may produce better therapeutic effect and (iii) a decrease in SP release may serve as one of the possible neurochemical mechanisms underlying the therapeutic effects of multiple TENS treatments on chronic inflammatory hyperalgesia. PMID:17342243

  19. Morphogenesis of protrusions from confined lipid bilayers mediated by mechanics

    NASA Astrophysics Data System (ADS)

    Arroyo, Marino; Staykova, Margarita; Rahimi, Mohammad; Stone, Howard A.

    2012-02-01

    Biological membranes adopt a wide range of shapes that structure and give functionality to cells, compartmentalizing the cytosol, forming organelles, or regulating their area. The formation, stabilization, and remodeling of these structures is generally attributed to localized forces or to biochemical processes (insertion of proteins, active compositional regulation). Noting that in the crowded intra and extra-cellular environments membranes are highly constrained, we explore to what extent can mechanics explain the shape of protrusions out of confined membranes. For this purpose, we developed an in-vitro system coupling a lipid bilayer to the strain-controlled deformation of an elastic sheet (Staykova et al, PNAS 108, 2011). We show that upon contracting the elastic support, tubular or spherical protrusions grow out of the adhered membrane, which can be reversibly controlled with strain and osmolarity without resorting to localized forces or chemical alterations of the bilayer. The morphologies produced by our minimal system are ubiquitous in cells, suggesting mechanics may be a simple and generic organizing principle. We can understand most of our observations in terms of a phase diagram accounting for elasticity, adhesion, and the limited amount of area and volume available.

  20. Molecular mechanisms of CRISPR-mediated microbial immunity.

    PubMed

    Gasiunas, Giedrius; Sinkunas, Tomas; Siksnys, Virginijus

    2014-02-01

    Bacteriophages (phages) infect bacteria in order to replicate and burst out of the host, killing the cell, when reproduction is completed. Thus, from a bacterial perspective, phages pose a persistent lethal threat to bacterial populations. Not surprisingly, bacteria evolved multiple defense barriers to interfere with nearly every step of phage life cycles. Phages respond to this selection pressure by counter-evolving their genomes to evade bacterial resistance. The antagonistic interaction between bacteria and rapidly diversifying viruses promotes the evolution and dissemination of bacteriophage-resistance mechanisms in bacteria. Recently, an adaptive microbial immune system, named clustered regularly interspaced short palindromic repeats (CRISPR) and which provides acquired immunity against viruses and plasmids, has been identified. Unlike the restriction–modification anti-phage barrier that subjects to cleavage any foreign DNA lacking a protective methyl-tag in the target site, the CRISPR–Cas systems are invader-specific, adaptive, and heritable. In this review, we focus on the molecular mechanisms of interference/immunity provided by different CRISPR–Cas systems.

  1. GABAergic Transmission in Rat Pontine Reticular Formation Regulates the Induction Phase of Anesthesia and Modulates Hyperalgesia Caused by Sleep Deprivation

    PubMed Central

    Vanini, Giancarlo; Nemanis, Kriste; Baghdoyan, Helen A.; Lydic, Ralph

    2014-01-01

    The oral part of the pontine reticular formation (PnO) contributes to the regulation of sleep, anesthesia, and pain. The role of PnO GABA in modulating these states remains incompletely understood. The present study used time to Loss and time to Resumption of Righting Response (LoRR and RoRR) as surrogate measures of loss and resumption of consciousness. This study tested three hypotheses: (1) pharmacologically manipulating GABA levels in rat PnO alters LoRR, RoRR, and nociception; (2) propofol decreases GABA levels in the PnO; and (3) inhibiting GABA synthesis in the PnO blocks hyperalgesia caused by sleep deprivation. Administering a GABA synthesis inhibitor (3-MPA) or a GABA uptake inhibitor (NPA) into rat PnO significantly altered LoRR caused by propofol. 3-MPA significantly decreased LoRR for propofol (−18%). NPA significantly increased LoRR during administration of propofol (36%). Neither 3-MPA nor NPA altered RoRR following cessation of propofol or isoflurane delivery. The finding that LoRR was decreased by 3-MPA and increased by NPA is consistent with measures showing that extracellular GABA levels in the PnO were decreased (41%) by propofol. Thermal nociception was significantly decreased by 3-MPA and increased by NPA, and 3-MPA blocked the hyperalgesia caused by sleep deprivation. The results demonstrate that GABA levels in the PnO regulate the time for loss of consciousness caused by propofol, extend the concept that anesthetic induction and emergence are not inverse processes, and suggest that GABAergic transmission in the PnO mediates hyperalgesia caused by sleep loss. PMID:24674578

  2. GABAergic transmission in rat pontine reticular formation regulates the induction phase of anesthesia and modulates hyperalgesia caused by sleep deprivation.

    PubMed

    Vanini, Giancarlo; Nemanis, Kriste; Baghdoyan, Helen A; Lydic, Ralph

    2014-07-01

    The oral part of the pontine reticular formation (PnO) contributes to the regulation of sleep, anesthesia and pain. The role of PnO γ-aminobutyric acid (GABA) in modulating these states remains incompletely understood. The present study used time to loss and time to resumption of righting response (LoRR and RoRR) as surrogate measures of loss and resumption of consciousness. This study tested three hypotheses: (i) pharmacologically manipulating GABA levels in rat PnO alters LoRR, RoRR and nociception; (ii) propofol decreases GABA levels in the PnO; and (iii) inhibiting GABA synthesis in the PnO blocks hyperalgesia caused by sleep deprivation. Administering a GABA synthesis inhibitor [3-mercaptopropionic acid (3-MPA)] or a GABA uptake inhibitor [nipecotic acid (NPA)] into rat PnO significantly altered LoRR caused by propofol. 3-MPA significantly decreased LoRR for propofol (-18%). NPA significantly increased LoRR during administration of propofol (36%). Neither 3-MPA nor NPA altered RoRR following cessation of propofol or isoflurane delivery. The finding that LoRR was decreased by 3-MPA and increased by NPA is consistent with measures showing that extracellular GABA levels in the PnO were decreased (41%) by propofol. Thermal nociception was significantly decreased by 3-MPA and increased by NPA, and 3-MPA blocked the hyperalgesia caused by sleep deprivation. The results demonstrate that GABA levels in the PnO regulate the time for loss of consciousness caused by propofol, extend the concept that anesthetic induction and emergence are not inverse processes, and suggest that GABAergic transmission in the PnO mediates hyperalgesia caused by sleep loss.

  3. Intra-cerebral and intra-nasal melanocortin-4 receptor antagonist blocks withdrawal hyperalgesia in alcohol-dependent rats.

    PubMed

    Roltsch Hellard, Emily A; Impastato, Renata A; Gilpin, Nicholas W

    2016-01-24

    Humans diagnosed with alcohol use disorder are more sensitive to painful stimuli during withdrawal, which suggests that excessive alcohol drinking worsens pain outcomes. Alcohol-dependent rats exhibit increases in nociceptive sensitivity during withdrawal. Data from animal models suggest that brain melanocortin-4 receptors (MC4Rs) mediate alcohol drinking and nociception. Here we tested: (1) the effect of alcohol dependence on thermal nociception in rats, and (2) the ability of acute alcohol and (3) MC4R antagonists to reverse hyperalgesia during withdrawal in alcohol-dependent rats. Rats were trained to self-administer operant alcohol and were tested for baseline thermal nociception. Half of the rats were made dependent on alcohol, then all rats were cannulated in the lateral ventricle. We tested the effects of acute alcohol drinking, acute fixed-dose alcohol, intra-ventricular agouti-related protein (endogenous MC4R antagonist), intra-ventricular HS014 (synthetic MC4R antagonist) and intra-nasal HS014 on hyperalgesia during withdrawal in alcohol-dependent rats, relative to non-dependent drinkers and alcohol-naïve controls. Alcohol-dependent rats exhibit thermal hyperalgesia that is abolished by alcohol drinking, bolus alcohol and intra-ventricular and intra-nasal MC4R antagonists. These manipulations did not affect thermal nociception in non-dependent drinkers and alcohol-naïve controls, suggesting that alcohol dependence produces neuroadaptations in brain MC4R systems. These results suggest that brain MC4R systems may be an effective therapeutic target for reducing nociception in the alcohol-dependent organism.

  4. Molecular mechanisms of ETS transcription factor-mediated tumorigenesis.

    PubMed

    Kar, Adwitiya; Gutierrez-Hartmann, Arthur

    2013-01-01

    The E26 transformation-specific (ETS) family of transcription factors is critical for development, differentiation, proliferation and also has a role in apoptosis and tissue remodeling. Changes in expression of ETS proteins therefore have a significant impact on normal physiology of the cell. Transcriptional consequences of ETS protein deregulation by overexpression, gene fusion, and modulation by RAS/MAPK signaling are linked to alterations in normal cell functions, and lead to unlimited increased proliferation, sustained angiogenesis, invasion and metastasis. Existing data show that ETS proteins control pathways in epithelial cells as well as stromal compartments, and the crosstalk between the two is essential for normal development and cancer. In this review, we have focused on ETS factors with a known contribution in cancer development. Instead of focusing on a prototype, we address cancer associated ETS proteins and have highlighted the diverse mechanisms by which they affect carcinogenesis. Finally, we discuss strategies for ETS factor targeting as a potential means for cancer therapeutics.

  5. Gap junction-mediated electrical transmission: regulatory mechanisms and plasticity

    PubMed Central

    Pereda, Alberto E.; Curti, Sebastian; Hoge, Gregory; Cachope, Roger; Flores, Carmen E.; Rash, John E.

    2012-01-01

    The term synapse applies to cellular specializations that articulate the processing of information within neural circuits by providing a mechanism for the transfer of information between two different neurons. There are two main modalities of synaptic transmission: chemical and electrical. While most efforts have been dedicated to the understanding of the properties and modifiability of chemical transmission, less is still known regarding the plastic properties of electrical synapses, whose structural correlate is the gap junction. A wealth of data indicates that, rather than passive intercellular channels, electrical synapses are more dynamic and modifiable than was generally perceived. This article will discuss the factors determining the strength of electrical transmission and review current evidence demonstrating its dynamic properties. Like their chemical counterparts, electrical synapses can also be plastic and modifiable. PMID:22659675

  6. Mechanically Induced Actin-mediated Rocketing of Phagosomes

    PubMed Central

    Müller-Taubenberger, Annette; Anderson, Kurt I.; Engel, Ulrike; Gerisch, Günther

    2006-01-01

    Actin polymerization can be induced in Dictyostelium by compressing the cells to bring phagosomes filled with large particles into contact with the plasma membrane. Asymmetric actin assembly results in rocketing movement of the phagosomes. We show that the compression-induced assembly of actin at the cytoplasmic face of the plasma membrane involves the Arp2/3 complex. We also identify two other proteins associated with the mechanically induced actin assembly. The class I myosin MyoB accumulates at the plasma membrane–phagosome interface early during the initiation of the response, and coronin is recruited as the actin filaments are disassembling. The forces generated by rocketing phagosomes are sufficient to push the entire microtubule apparatus forward and to dislocate the nucleus. PMID:16971511

  7. Molecular mechanisms of ETS transcription factor mediated tumorigenesis

    PubMed Central

    Kar, Adwitiya; Gutierrez-Hartmann, Arthur

    2014-01-01

    The ETS family of transcription factors is critical for development, differentiation, proliferation and also has a role in apoptosis and tissue remodeling. Changes in expression of ETS proteins therefore have a significant impact on normal physiology of the cell. Transcriptional consequences of ETS protein deregulation by overexpression, gene fusion, and modulation by RAS/MAPK signaling are linked to alterations in normal cell functions, and lead to unlimited increased proliferation, sustained angiogenesis, invasion and metastasis. Existing data show that ETS proteins control pathways in epithelial cells as well as stromal compartments, and the crosstalk between the two is essential for normal development and cancer. In this review we have focused on ETS factors with a known contribution in cancer development. Instead of focusing on a prototype, we address cancer associated ETS proteins and have highlighted the diverse mechanisms by which they affect carcinogenesis. Finally, we discuss strategies for ETS factor targeting as a potential means for cancer therapeutics. PMID:24066765

  8. Effect of painless diabetic neuropathy on pressure pain hypersensitivity (hyperalgesia) after acute foot trauma

    PubMed Central

    Wienemann, Tobias; Chantelau, Ernst A.; Koller, Armin

    2014-01-01

    Introduction and objective Acute injury transiently lowers local mechanical pain thresholds at a limb. To elucidate the impact of painless (diabetic) neuropathy on this post-traumatic hyperalgesia, pressure pain perception thresholds after a skeletal foot trauma were studied in consecutive persons without and with neuropathy (i.e. history of foot ulcer or Charcot arthropathy). Design and methods A case–control study was done on 25 unselected clinical routine patients with acute unilateral foot trauma (cases: elective bone surgery; controls: sprain, toe fracture). Cases were 12 patients (11 diabetic subjects) with severe painless neuropathy and chronic foot pathology. Controls were 13 non-neuropathic persons. Over 1 week after the trauma, cutaneous pressure pain perception threshold (CPPPT) and deep pressure pain perception threshold (DPPPT) were measured repeatedly, adjacent to the injury and at the opposite foot (pinprick stimulators, Algometer II®). Results In the control group, post-traumatic DPPPT (but not CPPPT) at the injured foot was reduced by about 15–25%. In the case group, pre- and post-operative CPPPT and DPPPT were supranormal. Although DPPPT fell post-operatively by about 15–20%, it remained always higher than the post-traumatic DPPPT in the control group: over musculus abductor hallucis 615 kPa (kilopascal) versus 422 kPa, and over metatarsophalangeal joint 518 kPa versus 375 kPa (medians; case vs. control group); CPPPT did not decrease post-operatively. Conclusion Physiological nociception and post-traumatic hyperalgesia to pressure are diminished at the foot with severe painless (diabetic) neuropathy. A degree of post-traumatic hypersensitivity required to ‘pull away’ from any one, even innocuous, mechanical impact in order to avoid additional damage is, therefore, lacking. PMID:25397867

  9. δ-Opioid receptor agonists inhibit migraine-related hyperalgesia, aversive state and cortical spreading depression in mice

    PubMed Central

    Pradhan, Amynah A; Smith, Monique L; Zyuzin, Jekaterina; Charles, Andrew

    2014-01-01

    Background and Purpose Migraine is an extraordinarily common brain disorder for which treatment options continue to be limited. Agonists that activate the δ-opioid receptor may be promising for the treatment of migraine as they are highly effective for the treatment of chronic rather than acute pain, do not induce hyperalgesia, have low abuse potential and have anxiolytic and antidepressant properties. The aim of this study was to investigate the therapeutic potential of δ-opioid receptor agonists for migraine by characterizing their effects in mouse migraine models. Experimental Approach Mechanical hypersensitivity was assessed in mice treated with acute and chronic doses of nitroglycerin (NTG), a known human migraine trigger. Conditioned place aversion to NTG was also measured as a model of migraine-associated negative affect. In addition, we assessed evoked cortical spreading depression (CSD), an established model of migraine aura, in a thinned skull preparation. Key Results NTG evoked acute and chronic mechanical and thermal hyperalgesia in mice, as well as conditioned place aversion. Three different δ-opioid receptor agonists, SNC80, ARM390 and JNJ20788560, significantly reduced NTG-evoked hyperalgesia. SNC80 also abolished NTG-induced conditioned place aversion, suggesting that δ-opioid receptor activation may also alleviate the negative emotional state associated with migraine. We also found that SNC80 significantly attenuated CSD, a model that is considered predictive of migraine preventive therapies. Conclusions and Implications These data show that δ-opioid receptor agonists modulate multiple basic mechanisms associated with migraine, indicating that δ-opioid receptors are a promising therapeutic target for this disorder. PMID:24467301

  10. Successful treatment of hydromorphone-induced neurotoxicity and hyperalgesia.

    PubMed

    Chung, Keun Sam; Carson, Shawn; Glassman, David; Vadivelu, Nalini

    2004-10-01

    There has been an increase in opioid consumption world wide in the last decade. There has also been a disturbing increase in the number of reports of neuroexcitatory opioid-related side effects observed in patients receiving large doses of systemically administered morphine and its structural analogue, hydromorphone. It is now becoming clearer that patients receiving long-term opioid therapy can develop unexpected pain. We describe an interesting case of successful management of hydromorphone-induced neurotoxicity and hyperalgesia produced by short-term therapy with rapidly escalating doses of systemic hydromorphone.

  11. The lateral prefrontal cortex mediates the hyperalgesic effects of negative cognitions in chronic pain patients

    PubMed Central

    Loggia, Marco L.; Berna, Chantal; Kim, Jieun; Cahalan, Christine M.; Martel, Marc-Olivier; Gollub, Randy L.; Wasan, Ajay D.; Napadow, Vitaly; Edwards, Robert R.

    2015-01-01

    While high levels of negative affect and cognitions have been associated in chronic pain conditions with greater pain sensitivity, the neural mechanisms mediating the hyperalgesic effect of psychological factors in patients with pain disorders are largely unknown. In this cross-sectional study, we hypothesized that 1) catastrophizing modulates brain responses to pain anticipation, and that 2) anticipatory brain activity mediates the hyperalgesic effect of different levels of catastrophizing, in fibromyalgia (FM) patients. Using functional Magnetic Resonance Imaging, we scanned the brains of 31 FM patients exposed to visual cues anticipating the onset of moderately intense deep-tissue pain stimuli. Our results indicated the existence of a negative association between catastrophizing and pain-anticipatory brain activity, including in the right lateral prefrontal cortex (IPFC). A bootstrapped mediation analysis revealed that pain-anticipatory activity in lateral prefrontal cortex (IPFC) mediates the association between catastrophizing and pain sensitivity. These findings highlight the role of IPFC in the pathophysiology of FM related hyperalgesia, and suggest that deficits in the recruitment of pain-inhibitory brain circuitry during pain-anticipatory periods may play an important contributory role in the association between various degrees of widespread hyperalgesia in FM and levels of catastrophizing, a well validated measure of negative cognitions and psychological distress. Perspective This article highlights the presence of alterations in pain-anticipatory brain activity in FM. These findings provide the rationale for the development of psychological or neurofeedback-based techniques aimed at modifying patients' negative affect and cognitions towards pain. PMID:25937162

  12. Cellular mechanisms of estradiol-mediated masculinization of the brain.

    PubMed

    Schwarz, Jaclyn M; McCarthy, Margaret M

    2008-04-01

    The sexual differentiation of reproductive physiology and behavior in the rodent brain is largely determined by estradiol aromatized from testicular androgens. The cellular mechanisms by which estradiol masculinizes the brain are beginning to emerge and revealing novel features of brain development that are highly region-specific. In the preoptic area, the major site controlling male sexual behavior, estradiol increases the level of the COX-2 enzyme and its product, prostaglandin E2 which promotes dendritic spine synaptogenesis. In the ventromedial nucleus of the hypothalamus, the major site controlling female reproductive behavior, estradiol promotes glutamate release from synaptic terminals, activating NMDA receptors and the MAP kinase pathway. In the arcuate nucleus, a major regulator of anterior pituitary function, estradiol increases GABA synthesis, altering the morphology of neighboring astrocytes and reducing formation of dendritic spines synapses. Glutamate, GABA and the importance of neuronal-astrocytic cross-talk are emerging as common aspects of masculinization. Advances are also being made in the mechanistic basis of female brain development, although the challenges are far greater.

  13. Toxin-mediated gene regulatory mechanism in Staphylococcus aureus

    PubMed Central

    Joo, Hwang-Soo; Otto, Michael

    2016-01-01

    The dangerous human pathogen Staphylococcus aureus relies heavily on toxins to cause disease, but toxin production can put a strong burden on the bacteria’s energy balance. Thus, controlling the synthesis of proteins solely needed in times of toxin production represents a way for the bacteria to avoid wasting energy. One hypothetical manner to accomplish this sort of regulation is by gene regulatory functions of the toxins themselves. There have been several reports about gene regulation by toxins in S. aureus, but these were never verified on the molecular level. In our study published in MBio [Joo et al., 7(5). pii: e01579-16], we show that phenol-soluble modulins (PSMs), important peptide toxins of S. aureus, release a repressor from the promoter of the operon encoding the toxin export system, thereby enabling toxin secretion. This study describes the first molecular regulatory mechanism exerted by an S. aureus toxin, setting a paradigmatic example of how S. aureus toxins may influence cell functions to adjust them to times of toxin production.

  14. Mechanisms underlying heterologous skin scaffold-mediated tissue remodeling

    PubMed Central

    Mimura, Kallyne K. O.; Moraes, Andréia R.; Miranda, Aline C.; Greco, Rebecca; Ansari, Tahera; Sibbons, Paul; Greco, Karin V.; Oliani, Sonia M.

    2016-01-01

    Biocompatibility of two newly developed porcine skin scaffolds was assessed after 3, 14, 21 and 90 days of implantation in rats. Both scaffolds showed absence of cells, preservation of ECM and mechanical properties comparable to non-decellularised skin before implantation. Host cell infiltration was much prominent on both scaffolds when compared to Permacol (surgical control). At day 3, the grafts were surrounded by polymorphonuclear cells, which were replaced by a notable number of IL-6-positive cells at day 14. Simultaneously, the number of pro-inflammatory M1-macrophage was enhanced. Interestingly, a predominant pro-remodeling M2 response, with newly formed vessels, myofibroblasts activation and a shift on the type of collagen expression was sequentially delayed (around 21 days). The gene expression of some trophic factors involved in tissue remodeling was congruent with the cellular events. Our findings suggested that the responsiveness of macrophages after non-crosslinked skin scaffolds implantation seemed to intimately affect various cell responses and molecular events; and this range of mutually reinforcing actions was predictive of a positive tissue remodeling that was essential for the long-standing success of the implants. Furthermore, our study indicates that non-crosslinked biologic scaffold implantation is biocompatible to the host tissue and somehow underlying molecular events involved in tissue repair. PMID:27725772

  15. Mechanisms of Gadographene-Mediated Proton Spin Relaxation

    PubMed Central

    Hung, Andy H.; Duch, Matthew C.; Parigi, Giacomo; Rotz, Matthew W.; Manus, Lisa M.; Mastarone, Daniel J.; Dam, Kevin T.; Gits, Colton C.; MacRenaris, Keith W.; Luchinat, Claudio; Hersam, Mark C.; Meade, Thomas J.

    2013-01-01

    Gd(III) associated with carbon nanomaterials relaxes water proton spins at an effectiveness that approaches or exceeds the theoretical limit for a single bound water molecule. These Gd(III)-labeled materials represent a potential breakthrough in sensitivity for Gd(III)-based contrast agents used for magnetic resonance imaging (MRI). However, their mechanism of action remains unclear. A gadographene library encompassing GdCl3, two different Gd(III)-complexes, graphene oxide (GO), and graphene suspended by two different surfactants and subjected to varying degrees of sonication was prepared and characterized for their relaxometric properties. Gadographene was found to perform comparably to other Gd(III)-carbon nanomaterials; its longitudinal (r1) and transverse (r2) relaxivity is modulated between 12–85 mM−1s−1 and 24–115 mM−1s−1, respectively, depending on the Gd(III)-carbon backbone combination. The unusually large relaxivity and its variance can be understood under the modified Florence model incorporating the Lipari-Szabo approach. Changes in hydration number (q), water residence time (τM), molecular tumbling rate (τR), and local motion (τfast) sufficiently explain most of the measured relaxivities. Furthermore, results implicated the coupling between graphene and Gd(III) as a minor contributor to proton spin relaxation. PMID:24298299

  16. Gabapentin reduces allodynia and hyperalgesia in painful diabetic neuropathy rats by decreasing expression level of Nav1.7 and p-ERK1/2 in DRG neurons.

    PubMed

    Zhang, Jun-Long; Yang, Jan-Ping; Zhang, Ji-Ru; Li, Rui-Qin; Wang, Jing; Jan, Jin-Jin; Zhuang, Qing

    2013-02-01

    It has been confirmed that gabapentin (GBP) induced a inhibition of the voltage-gated persistent sodium current in chronically compressed dorsal root ganglion (DRG) neurons. The persistent sodium current is found in excitable DRG neurons of painful diabetic neuropathy (PDN) rats where it is mediated by tetrodotoxin (TTX) sensitive sodium channels. Recently, many groups have used models of neurological disorder to explore the mechanism of GBP in neuropathic pain. There is no evidence, however, to explain the particular mechanism of GBP, including its analgesic actions in PDN rats. These issues were addressed in the present study. Using behavioral testing, we found that diabetes leads to mechanical allodynia and thermal hyperalgesia and these effects were reversed by a continuous GBP injection. To investigate the mechanism of GBP's reduction in neural excitability, we systematically analyzed the expression of Nav1.7 and p-ERK1/2 and tested the effect of GBP on these proteins. Diabetes significantly increased the excitability of DRG neurons and the expression of Nav1.7 and p-ERK1/2, and GBP significantly inhibited these changes. These results suggest that the inhibitory effect of GBP on the expression of Nav1.7 and p-ERK1/2 might be one of the analgesic mechanisms of action of GBP. This may partially explain the antinociceptive action of GBP in the PDN rats.

  17. Surface charge of gold nanoparticles mediates mechanism of toxicity

    NASA Astrophysics Data System (ADS)

    Schaeublin, Nicole M.; Braydich-Stolle, Laura K.; Schrand, Amanda M.; Miller, John M.; Hutchison, Jim; Schlager, John J.; Hussain, Saber M.

    2011-02-01

    Recently gold nanoparticles (Au NPs) have shown promising biological and military applications due to their unique electronic and optical properties. However, little is known about their biocompatibility in the event that they come into contact with a biological system. In the present study, we have investigated whether modulating the surface charge of 1.5 nm Au NPs induced changes in cellular morphology, mitochondrial function, mitochondrial membrane potential (MMP), intracellular calcium levels, DNA damage-related gene expression, and of p53 and caspase-3 expression levels after exposure in a human keratinocyte cell line (HaCaT). The evaluation of three different Au NPs (positively charged, neutral, and negatively charged) showed that cell morphology was disrupted by all three NPs and that they demonstrated a dose-dependent toxicity; the charged Au NPs displayed toxicity as low as 10 µg ml-1 and the neutral at 25 µg ml-1. Furthermore, there was significant mitochondrial stress (decreases in MMP and intracellular Ca2+ levels) following exposure to the charged Au NPs, but not the neutral Au NPs. In addition to the differences observed in the MMP and Ca2+ levels, up or down regulation of DNA damage related gene expression suggested a differential cell death mechanism based on whether or not the Au NPs were charged or neutral. Additionally, increased nuclear localization of p53 and caspase-3 expression was observed in cells exposed to the charged Au NPs, while the neutral Au NPs caused an increase in both nuclear and cytoplasmic p53 expression. In conclusion, these results indicate that surface charge is a major determinant of how Au NPs impact cellular processes, with the charged NPs inducing cell death through apoptosis and neutral NPs leading to necrosis.Recently gold nanoparticles (Au NPs) have shown promising biological and military applications due to their unique electronic and optical properties. However, little is known about their biocompatibility in the

  18. Benzoxazinone-Mediated Triazine Degradation: A Proposed Reaction Mechanism.

    PubMed

    Willett, C D; Lerch, R N; Lin, C-H; Goyne, K W; Leigh, N D; Roberts, C A

    2016-06-22

    The role of benzoxazinones (Bx, 2-hydroxy-2H-1,4-benzoxazin-3(4H)-one) in triazine resistance in plants has been studied for over half a century. In this research, fundamental parameters of the reaction between DIBOA-Glc (2-β-d-glucopyranosyloxy-4-hydroxy-1,4-benzoxazin-3-one) and atrazine (ATR, 6-chloro-N-ethyl-N'-(1-methylethyl)-1,3,5-triazine-2,4-diamine) were examined. Through a series of experiments employing a variety of chromatographic and spectroscopic techniques, the DIBOA-Glc/ATR reaction was characterized in terms of reactant and product kinetics, stoichiometry, identification of a reaction intermediate, and reaction products formed. Results of these experiments demonstrated that the reaction mechanism proceeds via nucleophilic attack of the hydroxamic acid moiety of DIBOA-Glc at the C-2 position of the triazine ring to form hydroxyatrazine (HA, 2-hydroxy-4-ethylamino-6-isopropylamino-s-triazine), with associated degradation of DIBOA-Glc. Degradation of reactants followed first-order kinetics with a noncatalytic role of DIBOA-Glc. A reaction intermediate was identified as a DIBOA-Glc-HA conjugate, indicating a 1:1 DIBOA-Glc:ATR stoichiometry. Reaction products included HA and Cl(-), but definitive identification of DIBOA-Glc reaction product(s) was not attained. With these reaction parameters elucidated, DIBOA-Glc can be evaluated in terms of its potential for a myriad of applications, including its use to address the problem of widespread ATR contamination of soil and water resources.

  19. Transcutaneous electric acupoint stimulation alleviates remifentanil-induced hyperalgesia in patients undergoing thyroidectomy: a randomized controlled trial

    PubMed Central

    Chen, Yanqing; Yao, Yusheng; Wu, Yihuan; Dai, Dongsheng; Zhao, Qiuyan; Qiu, Liangcheng

    2015-01-01

    Background: In this prospective, randomized, double-blind study, we verified the hypothesis that TEAS can alleviate remifentanil-induced hyperalgesia in patients undergoing thyroidectomy. Methods: 60 American Society of Anesthesiologists physical status (ASA) I-IIpatients, aged 18-60 year, scheduled for thyroidectomy were randomly allocated to TEAS or sham groups. TEAS consisted of 30 min of stimulation (6-9 mA, 2/10 Hz) on the Hegu (LI4) and Neiguan (PC6) before anesthesia. Anesthesia was maintained with sevoflurane adjusted to bispectral index (40-60) and target remifentanil 5.0 ng/ml. Mechanical pain thresholds were assessed using electronic von Frey. The primary outcome was mechanical pain thresholds. Secondary outcomes included postoperative pain scores, the time to first rescue analgesic, cumulative number of rescue analgesia, and side effects, including postoperative nausea and vomiting (PONV), dizziness and shivering in 24 h postoperatively. Results: Baseline mechanical pain thresholds were similar between the groups. The analysis revealed the decrease in mechanical threshold was greater in the sham group than the TEAS group (P < 0.001). Postoperative pain scores and cumulative number of rescue analgesia were lower in the TEAS group (P < 0.05). In addition, TEAS group patients reduced the incidence of PONV and shivering. Conclusion: Preoperative TEAS can attenuate remifentanil-induced hyperalgesia in patients undergoing thyroidectomy. PMID:26131165

  20. Downregulation of the spinal NMDA receptor NR2B subunit during electro-acupuncture relief of chronic visceral hyperalgesia.

    PubMed

    Liu, Hongping; Zhang, Yuhua; Qi, Debo; Li, Weimin

    2017-01-01

    The involvement of spinal NR2B, a N-methyl-D-aspartate (NMDA) receptor subunit, in the therapeutic effect of electro-acupuncture (EA) on chronic visceral hyperalgesia was investigated. Chronic visceral hyperalgesia was induced using an irritable bowel syndrome (IBS) model in rats. Graded colorectal distention (CRD) stimuli at strengths of 20, 40, 60 and 80 mmHg were applied, and behavioral tests were performed to measure the abdominal withdrawal reflex (AWR) in response to the CRD stimuli and assess the severity of the visceral hyperalgesia. Rats were randomly divided into four groups: normal intact (control) group, IBS model (model) group, EA-treated IBS rats (EA) group and sham EA-treated IBS rats (sham EA) group. For the EA treatment, electric stimuli were applied through needles inserted into two acupoints [Zu-san-li (ST-36) and Shang-ju-xu (ST-37)] in both hind limbs, while the sham EA treatment consisted of only the insertion of needles into these same acupoints without an application of electric stimuli. Our results showed that AWR scores of the model group responding to CRD stimuli of 20, 40, 60 and 80 mmHg were significantly increased. These increased scores subsequently decreased following EA treatment (P < 0.05) compared with those for the other groups. The expression of NR2B in the superficial laminae (SDH, laminae I and II), nucleus proprius (NP, laminae III and IV), neck of the dorsal horn (NECK, laminae V and VI) and central canal region (lamina X) at thoracolumbar (T13-L2) and lumbosacral (L6-S2) segmental level significantly increased in the model group versus the control group (P < 0.05) and significantly decreased after EA treatment (P < 0.05). There were no significant changes in neither AWR scores nor expression of the NR2B subunit in these spinal regions after the sham EA treatment. These results confirm that EA can relieve chronic visceral hyperalgesia in IBS model rats and suggest that such an effect is possibly mediated through the

  1. Mediating Mechanisms for the Intergenerational Transmission of Constructive Parenting: A Prospective Longitudinal Study

    PubMed Central

    Chen, Zeng-yin; Liu, Ruth X.; Kaplan, Howard B.

    2010-01-01

    Based on a prospective longitudinal panel data set that was collected at three developmental stages—early adolescence, young adulthood, and middle adulthood— this study investigates marital satisfaction and educational attainment as mediating mechanisms as well as gender's moderating effect for the intergenerational transmission of constructive parenting (N = 1,560). The results show that perceived satisfying experiences with parents during early adolescence are positively related to marital satisfaction and educational attainment in young adulthood, which, in turn, are positively related to individuals' utilization of constructive parenting in middle adulthood. The two mediating mechanisms account for most of the direct effect of the intergenerational transmission of constructive parenting. Furthermore, the mediating effect of marital relationship is stronger for current fathers than for mothers because of a stronger association between marital satisfaction and constructive parenting for men. The implications are discussed. PMID:20357901

  2. The Mediated MIMIC Model for Understanding the Underlying Mechanism of DIF

    ERIC Educational Resources Information Center

    Cheng, Ying; Shao, Can; Lathrop, Quinn N.

    2016-01-01

    Due to its flexibility, the multiple-indicator, multiple-causes (MIMIC) model has become an increasingly popular method for the detection of differential item functioning (DIF). In this article, we propose the mediated MIMIC model method to uncover the underlying mechanism of DIF. This method extends the usual MIMIC model by including one variable…

  3. Mother-Child Attachment and Cognitive Performance in Middle Childhood: An Examination of Mediating Mechanisms

    ERIC Educational Resources Information Center

    West, Katara K.; Mathews, Brittany L.; Kerns, Kathryn A.

    2013-01-01

    Although mother-child attachment has been shown to predict cognitive performance, there has been a lack of attention to the mediating mechanisms that explain these associations. In the present study, we investigated relations of early mother-child attachment and cognitive performance in middle childhood (the latter in terms of both academic…

  4. Microwave-Mediated Synthesis of Lophine: Developing a Mechanism to Explain a Product

    ERIC Educational Resources Information Center

    Crouch, R. David; Howard, Jessica L.; Zile, Jennifer L.; Barker, Kathryn H.

    2006-01-01

    The microwave-mediated preparation of lophine (2,4,5-triphenylimidazole) is described. This experiment allows for an introduction to the emerging technology of microwave-assisted organic synthesis while providing an opportunity for students to employ the principles of carbonyl chemistry in devising a mechanism to explain the formation of the…

  5. Rheological and mechanical properties of polyamide 6 modified by electron-beam initiated mediation process

    NASA Astrophysics Data System (ADS)

    Shin, Boo Young; Kim, Jae Hong

    2015-07-01

    Polyamide (PA6) has been modified by electron-beam initiated mediator process to improve drawbacks of PA6. Glycidyl methacrylate (GMA) was chosen as a reactive mediator for modification process of PA6. The mixture of the PA6 and GMA was prepared by using a twin-screw extruder, and then the mixture was exposed to electron-beam irradiation at various doses at room temperature. The modified PA6 were characterized by observing rheological and mechanical properties and compared virgin PA6. Thermal properties, water absorption, and gel fraction were also investigated. Tight gel was not found even when PA6 was irradiated at 200 kGy. Complex viscosity and storage modulus of PA6 were remarkably increased by electron-beam irradiation with medium of GMA. Maximum increase in complex viscosity was 75 times higher than virgin PA6 at 0.1 rad/s when it was irradiated at 200 kGy with the GMA. Mechanical properties were also improved without scarifying of processability. The reaction mechanisms for the mediation process with the reactive mediator of GMA were estimated to elucidate the cause of significantly enhanced rheological and mechanical properties without loss of thermoplasticity.

  6. Crosstalk between Activated Microglia and Neurons in the Spinal Dorsal Horn Contributes to Stress-induced Hyperalgesia

    PubMed Central

    Qi, Jian; Chen, Chen; Meng, Qing-Xi; Wu, Yan; Wu, Haitao; Zhao, Ting-Bao

    2016-01-01

    Stress has been shown to enhance pain sensitivity resulting in stress-induced hyperalgesia. However, the underlying mechanisms have yet to be elucidated. Using single-prolonged stress combined with Complete Freund’s Adjuvant injection model, we explored the reciprocal regulatory relationship between neurons and microglia, which is critical for the maintenance of posttraumatic stress disorder (PTSD)-induced hyperalgesia. In our assay, significant mechanical allodynia was observed. Additionally, activated neurons in spinal dorsal horn were observed by analysis of Fos expression. And, microglia were also significantly activated with the presence of increased Iba-1 expression. Intrathecal administration of c-fos antisense oligodeoxynucleotides (ASO) or minocycline (a specific microglia inhibitor) attenuated mechanical allodynia. Moreover, intrathecal administration of c-fos ASO significantly suppressed the activation of neurons and microglia. Interestingly, inhibition of microglia activation by minocycline significantly suppressed the activation of both neurons and microglia in spinal dorsal horn. P38 inhibitor SB203580 suppressed IL-6 production, and inhibition of IL-6 receptor (IL-6R) activation by tocilizumab suppressed Fos expression. Together, our data suggest that the presence of a “crosstalk” between activated microglia and neurons in the spinal dorsal horn, which might contribute to the stress-induced hyperactivated state, leading to an increased pain sensitivity. PMID:27995982

  7. Mechanisms of Biased β-Arrestin-Mediated Signaling Downstream from the Cannabinoid 1 Receptor

    PubMed Central

    Delgado-Peraza, Francheska; Ahn, Kwang H.; Nogueras-Ortiz, Carlos; Mungrue, Imran N.; Mackie, Ken; Kendall, Debra A.

    2016-01-01

    Activation of G protein-coupled receptors results in multiple waves of signaling that are mediated by heterotrimeric G proteins and the scaffolding proteins β-arrestin 1/2. Ligands can elicit full or subsets of cellular responses, a concept defined as ligand bias or functional selectivity. However, our current understanding of β-arrestin-mediated signaling is still very limited. Here we provide a comprehensive view of β-arrestin-mediated signaling from the cannabinoid 1 receptor (CB1R). By using a signaling biased receptor, we define the cascades, specific receptor kinases, and molecular mechanism underlying β-arrestin-mediated signaling: We identify the interaction kinetics of CB1R and β-arrestin 1 during their endocytic trafficking as directly proportional to its efficacy. Finally, we demonstrate that signaling results in the control of genes clustered around prosurvival and proapoptotic functions among others. Together, these studies constitute a comprehensive description of β-arrestin-mediated signaling from CB1Rs and suggest modulation of receptor endocytic trafficking as a therapeutic approach to control β-arrestin-mediated signaling. PMID:27009233

  8. Lycopene ameliorates thermal hyperalgesia and cold allodynia in STZ-induced diabetic rat.

    PubMed

    Kuhad, Anurag; Chopra, Kanwaljit

    2008-02-01

    Peripheral neuropathy is one of the common complications of diabetes mellitus. It is frequently associated with debilitating pain. The present study was designed to investigate effect of Lycopene, a carotenoid found in tomatoes, on hyperalgesia and cold allodynia in streptozotocin (STZ) induced diabetic rats. After 4-weeks of STZ injection, diabetic mice exhibited a significant thermal hyperalgesia cold allodynia, hyperglycemia and loss of body weights as compared with control rats. Chronic treatment of lycopene for 4 weeks significantly attenuated the cold allodynia and thermal hyperalgesia. The results emphasize the role of antioxidant such as lycopene as an adjuvant therapy in the treatment of diabetic neuropathy.

  9. The Relation of Perceived Neighborhood Danger to Childhood Aggression: A Test of Mediating Mechanisms1

    PubMed Central

    Colder, Craig R.; Mott, Joshua; Levy, Susan; Flay, Brian

    2008-01-01

    In the current study, two mediational mechanisms, parenting practices and children’s beliefs about aggression, were hypothesized to account for the relationship between perceived neighborhood danger and childhood aggression. Using structural equation modeling, data were analyzed from an inner-city school-based sample of 732 predominantly African American 5th graders. Results suggested that perceived neighborhood danger was associated with strong positive beliefs about aggression, which in turn was associated with high levels of aggression. The hypothesized mediating role of parenting practices (restrictive discipline, parental monitoring, and parental involvement) on the relation between perceived neighborhood danger and child aggression was not supported. However, the current findings suggest that children’s positive beliefs about aggression mediated the relationship between restrictive discipline and aggression. Directions for future research are discussed. PMID:10824275

  10. CLASSICAL CONDITIONING AND PAIN: CONDITIONED ANALGESIA AND HYPERALGESIA

    PubMed Central

    Miguez, Gonzalo; Laborda, Mario A.; Miller, Ralph R.

    2013-01-01

    This article reviews situations in which stimuli produce an increase or a decrease in nociceptive responses through basic associative processes and provides an associative account of such changes. Specifically, the literature suggests that cues associated with stress can produce conditioned analgesia or conditioned hyperalgesia, depending on the properties of the conditioned stimulus (e.g., contextual cues and audiovisual cues vs. gustatory and olfactory cues, respectively) and the proprieties of the unconditioned stimulus (e.g., appetitive, aversive, or analgesic, respectively). When such cues are associated with reducers of exogenous pain (e.g., opiates), they typically increase sensitivity to pain. Overall, the evidence concerning conditioned stress-induced analgesia, conditioned hyperalagesia, conditioned tolerance to morphine, and conditioned reduction of morphine analgesia suggests that selective associations between stimuli underlie changes in pain sensitivity. PMID:24269884

  11. Gabapentin alleviates facet-mediated pain in the rat through reduced neuronal hyperexcitability and astrocytic activation in the spinal cord

    PubMed Central

    Dong, Ling; Crosby, Nathan D.; Winkelstein, Beth A.

    2013-01-01

    Although joint pain is common, its mechanism(s) remain undefined, with little known about the spinal neuronal responses that contribute to this type of pain. Afferent activity and sustained spinal neuronal hyperexcitability correlate to facet joint loading and the extent of behavioral sensitivity induced after painful facet injury, suggesting spinal neuronal plasticity is induced in association with facet-mediated pain. This study used a rat model of painful C6/C7 facet joint stretch, together with intrathecal administration of gabapentin, to investigate the effects of one aspect of spinal neuronal function on joint pain. Gabapentin or saline vehicle was given via lumbar puncture prior to and at 1 day after painful joint distraction. Mechanical hyperalgesia was measured in the forepaw for 7 days. Extracellular recordings of neuronal activity and astrocytic and microglial activation in the cervical spinal cord were evaluated at day 7. Gabapentin significantly (p=0.0001) attenuated mechanical hyperalgesia and the frequency of evoked neuronal firing also significantly decreased (p<0.047) with gabapentin treatment. Gabapentin also decreased (p<0.04) spinal GFAP expression. Although spinal Iba1 expression was doubled over sham, gabapentin did not reduce it. Facet joint-mediated pain appears to be sustained through spinal neuronal modifications that are also associated with astrocytic activation. PMID:24094695

  12. Chrysin induces hyperalgesia via the GABAA receptor in mice.

    PubMed

    Zhai, Kui; Hu, Li; Chen, Juan; Fu, Cai-Yun; Chen, Qiang

    2008-08-01

    Chrysin (5,7-dihydroxyflavone) is a natural flavone commonly found in many plants including PASSIFLORA COERULEA L. Researchers have performed extensive and detailed investigations on the behavioral and pharmacological effects of chrysin IN VIVO, but there was little information available on the effect of chrysin on nociception. Therefore, the present study was undertaken to investigate the effect of chrysin on the nociceptive threshold using the tail-immersion test. Intraperitoneal ( I. P.) injection of chrysin (10, 25, 50, 75, 100 mg/kg) dose- and time-dependently induced a pronounced decrease of the tail withdrawal latencies (TWL), thus characterizing a hyperalgesic effect (ED50 = 65.59 mg/kg). The following results showed that GABAA receptors were involved in the hyperalgesic effects of chrysin. 1) The hyperalgesia induced by chrysin was significantly and dose-dependently blocked by pretreatment with flumazenil (0.75, 1 mg/kg, I. P.), a specific antagonist for benzodiazepine sites associated with GABAA receptors. 2) Bicuculline (2, 4 mg/kg, I. P.), a GABAA receptor antagonist, markedly antagonized the hyperalgesic effect of chrysin in a dose-dependent manner. 3) Picrotoxin (2 mg/kg, I. P.), a chloride channel blocker, could also notably antagonize the hyperalgesia of chrysin. Oral administration of chrysin (75 mg/kg) also produced a hyperalgesic effect in the tail-immersion test. In addition, diazepam (1 mg/kg, I. P.) showed a marked antinociceptive effect, which was completely blocked by flumazenil (1 mg/kg, I. P.). In conclusion, it can be summarized that both I. P. and oral administration of chrysin produced a significant hyperalgesic effect in the tail-immersion test and that the hyperalgesic effect of chrysin may be associated with GABAA receptors.

  13. Inflammation and hyperalgesia in rats neonatally treated with capsaicin: effects on two classes of nociceptive neurons in the superficial dorsal horn.

    PubMed

    Ren, K; Williams, G M; Ruda, M A; Dubner, R

    1994-11-01

    To address the mechanisms of hyperalgesia and dorsal horn plasticity following peripheral tissue inflammation, the effects of adjuvant-induced inflammation of the rat hindpaw on behavioral nociception and nociceptive neuronal activity in the superficial dorsal horn were examined in neonatally capsaicin-treated rats 6-8 weeks of age. Capsaicin treatment resulted in an 82% loss of unmyelinated fibers in L5 dorsal roots, a dramatic reduction of substance P-like immunoreactivity in the spinal cord, and a significant decrease in the percentage of dorsal horn nociceptive neurons that responded to C-fiber stimulation and noxious heating of the skin. The thermal nociceptive threshold was significantly increased in capsaicin-treated rats, but behavioral hyperalgesia to thermal stimuli still developed in response to inflammation. Following inflammation, there was a significant decrease in mechanical threshold and an increase in response duration to mechanical stimuli in both vehicle- and capsaicin-treated rats, suggesting that a state of mechanical hyperalgesia was also induced. The capsaicin treatment appears to have differential effects on nociceptive specific (NS) and wide-dynamic-range (WDR) neurons in inflamed rats. Expansion of the receptive fields of nociceptive neurons, a measure of the effect of inflammation-induced CNS plasticity, was less extensive for NS than for WDR neurons in capsaicin-treated rats. Compared to vehicle-treated rats, a smaller population of NS neurons, but a similar percentage of WDR neurons, had background activity in inflamed capsaicin-treated rats. C-fiber strength electrical stimulation of the sciatic nerve produced expansion of the receptive fields in a greater portion of NS neurons (53%, P < 0.05) in capsaicin- than in vehicle-treated rats (32%). There was no difference in stimulation-induced expansion of the receptive fields for WDR neurons between vehicle- or capsaicin-treated rats. An N-methyl-D-aspartate receptor antagonist, MK-801

  14. Molecular mechanism of protopanaxadiol saponin fraction-mediated anti-inflammatory actions

    PubMed Central

    Yang, Yanyan; Lee, Jongsung; Rhee, Man Hee; Yu, Tao; Baek, Kwang-Soo; Sung, Nak Yoon; Kim, Yong; Yoon, Keejung; Kim, Ji Hye; Kwak, Yi-Seong; Hong, Sungyoul; Kim, Jong-Hoon; Cho, Jae Youl

    2014-01-01

    Background Korean Red Ginseng (KRG) is a representative traditional herbal medicine with many different pharmacological properties including anticancer, anti-atherosclerosis, anti-diabetes, and anti-inflammatory activities. Only a few studies have explored the molecular mechanism of KRG-mediated anti-inflammatory activity. Methods We investigated the anti-inflammatory mechanisms of the protopanaxadiol saponin fraction (PPD-SF) of KRG using in vitro and in vivo inflammatory models. Results PPD-SF dose-dependently diminished the release of inflammatory mediators [nitric oxide (NO), tumor necrosis factor-α, and prostaglandin E2], and downregulated the mRNA expression of their corresponding genes (inducible NO synthase, tumor necrosis factor-α, and cyclooxygenase-2), without altering cell viability. The PPD-SF-mediated suppression of these events appeared to be regulated by a blockade of p38, c-Jun N-terminal kinase (JNK), and TANK (TRAF family member-associated NF-kappa-B activator)-binding kinase 1 (TBK1), which are linked to the activation of activating transcription factor 2 (ATF2) and interferon regulatory transcription factor 3 (IRF3). Moreover, this fraction also ameliorated HCl/ethanol/-induced gastritis via suppression of phospho-JNK2 levels. Conclusion These results strongly suggest that the anti-inflammatory action of PPD-SF could be mediated by a reduction in the activation of p38-, JNK2-, and TANK-binding-kinase-1-linked pathways and their corresponding transcription factors (ATF2 and IRF3). PMID:25535478

  15. Response to mechanical stress is mediated by the TRPA channel painless in the Drosophila heart.

    PubMed

    Sénatore, Sébastien; Rami Reddy, Vatrapu; Sémériva, Michel; Perrin, Laurent; Lalevée, Nathalie

    2010-09-02

    Mechanotransduction modulates cellular functions as diverse as migration, proliferation, differentiation, and apoptosis. It is crucial for organ development and homeostasis and leads to pathologies when defective. However, despite considerable efforts made in the past, the molecular basis of mechanotransduction remains poorly understood. Here, we have investigated the genetic basis of mechanotransduction in Drosophila. We show that the fly heart senses and responds to mechanical forces by regulating cardiac activity. In particular, pauses in heart activity are observed under acute mechanical constraints in vivo. We further confirm by a variety of in situ tests that these cardiac arrests constitute the biological force-induced response. In order to identify molecular components of the mechanotransduction pathway, we carried out a genetic screen based on the dependence of cardiac activity upon mechanical constraints and identified Painless, a TRPA channel. We observe a clear absence of in vivo cardiac arrest following inactivation of painless and further demonstrate that painless is autonomously required in the heart to mediate the response to mechanical stress. Furthermore, direct activation of Painless is sufficient to produce pauses in heartbeat, mimicking the pressure-induced response. Painless thus constitutes part of a mechanosensitive pathway that adjusts cardiac muscle activity to mechanical constraints. This constitutes the first in vivo demonstration that a TRPA channel can mediate cardiac mechanotransduction. Furthermore, by establishing a high-throughput system to identify the molecular players involved in mechanotransduction in the cardiovascular system, our study paves the way for understanding the mechanisms underlying a mechanotransduction pathway.

  16. Conservation of miRNA-mediated silencing mechanisms across 600 million years of animal evolution

    PubMed Central

    Mauri, Marta; Kirchner, Marieluise; Aharoni, Reuven; Ciolli Mattioli, Camilla; van den Bruck, David; Gutkovitch, Nadya; Modepalli, Vengamanaidu; Selbach, Matthias; Moran, Yehu; Chekulaeva, Marina

    2017-01-01

    Our current knowledge about the mechanisms of miRNA silencing is restricted to few lineages such as vertebrates, arthropods, nematodes and land plants. miRNA-mediated silencing in bilaterian animals is dependent on the proteins of the GW182 family. Here, we dissect the function of GW182 protein in the cnidarian Nematostella, separated by 600 million years from other Metazoa. Using cultured human cells, we show that Nematostella GW182 recruits the CCR4-NOT deadenylation complexes via its tryptophan-containing motifs, thereby inhibiting translation and promoting mRNA decay. Further, similarly to bilaterians, GW182 in Nematostella is recruited to the miRNA repression complex via interaction with Argonaute proteins, and functions downstream to repress mRNA. Thus, our work suggests that this mechanism of miRNA-mediated silencing was already active in the last common ancestor of Cnidaria and Bilateria. PMID:27604873

  17. Conservation of miRNA-mediated silencing mechanisms across 600 million years of animal evolution.

    PubMed

    Mauri, Marta; Kirchner, Marieluise; Aharoni, Reuven; Ciolli Mattioli, Camilla; van den Bruck, David; Gutkovitch, Nadya; Modepalli, Vengamanaidu; Selbach, Matthias; Moran, Yehu; Chekulaeva, Marina

    2017-01-25

    Our current knowledge about the mechanisms of miRNA silencing is restricted to few lineages such as vertebrates, arthropods, nematodes and land plants. miRNA-mediated silencing in bilaterian animals is dependent on the proteins of the GW182 family. Here, we dissect the function of GW182 protein in the cnidarian Nematostella, separated by 600 million years from other Metazoa. Using cultured human cells, we show that Nematostella GW182 recruits the CCR4-NOT deadenylation complexes via its tryptophan-containing motifs, thereby inhibiting translation and promoting mRNA decay. Further, similarly to bilaterians, GW182 in Nematostella is recruited to the miRNA repression complex via interaction with Argonaute proteins, and functions downstream to repress mRNA. Thus, our work suggests that this mechanism of miRNA-mediated silencing was already active in the last common ancestor of Cnidaria and Bilateria.

  18. The mechanism of neutral red-mediated microbial electrosynthesis in Escherichia coli: menaquinone reduction.

    PubMed

    Harrington, Timothy D; Tran, Vi N; Mohamed, Abdelrhman; Renslow, Ryan; Biria, Saeid; Orfe, Lisa; Call, Douglas R; Beyenal, Haluk

    2015-09-01

    The aim of this work was to elucidate the mechanism of mediated microbial electrosynthesis via neutral red from an electrode to fermenting Escherichia coli cultures in a bioelectrochemical system. Chemical reduction of NAD(+) by reduced neutral red did not occur as predicted. Instead, neutral red was shown to reduce the menaquinone pool in the inner bacterial membrane. The reduced menaquinone pool altered fermentative metabolite production via the arcB redox-sensing cascade in the absence of terminal electron acceptors. When the acceptors DMSO, fumarate, or nitrate were provided, as many as 19% of the electrons trapped in the reduced acceptors were derived from the electrode. These results demonstrate the mechanism of neutral red-mediated microbial electrosynthesis during fermentation as well as how neutral red enables microbial electrosynthesis of reduced terminal electron acceptors.

  19. The mechanism of neutral red-mediated microbial electrosynthesis in Escherichia coli: menaquinone reduction

    PubMed Central

    Harrington, Timothy D.; Tran, Vi N.; Mohamed, Abdelrhman; Renslow, Ryan; Biria, Saeid; Orfe, Lisa; Call, Douglas R.; Beyenal, Haluk

    2015-01-01

    The aim of this work was to elucidate the mechanism of mediated microbial electrosynthesis via neutral red from an electrode to fermenting Escherichia coli cultures in a bioelectrochemical system. Chemical reduction of NAD+ by reduced neutral red did not occur as predicted. Instead, neutral red was shown to reduce the menaquinone pool in the inner bacterial membrane. The reduced menaquinone pool altered fermentative metabolite production via the arcB redoxsensing cascade in the absence of terminal electron acceptors. When the acceptors DMSO, fumarate, or nitrate were provided, as many as 19% of the electrons trapped in the reduced acceptors were derived from the electrode. These results demonstrate the mechanism of neutral red-mediated microbial electrosynthesis during fermentation as well as how neutral red enables microbial electrosynthesis of reduced terminal electron acceptors. PMID:26094195

  20. Fucoidan attenuates the existing allodynia and hyperalgesia in a rat model of neuropathic pain.

    PubMed

    Hu, Chuanyin; Zhang, Guoping; Zhao, Yun-Tao

    2014-06-13

    Fucoidan is an active constituent found in brown seaweeds, which have potential neuroprotection. The current study aimed to investigate the effects of fucoidan on the maintenance of neuropathic pain induced by L5 spinal nerve ligation (SNL) and the underlying mechanism related to the spinal neuroimmune responses. Animals were randomized into 5 groups: sham-operation with vehicle and SNL with vehicle or fucoidan (15, 50, and 100mg/kg). Different doses of fucoidan or vehicle were administered intrathecally once daily from postoperative day (POD) 11-20. Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) was measured on 1 day before operation and days 10, 20, 22, 24, 26, 28, 30 after operation. Glial activation markers such as glial fibrillary acidic protein (GFAP) and macrophage antigen complex-1 (mac-1), inflammatory cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6 activation, and extracellular signalregulated protein kinase (ERK) activation in the lumbar spinal cord were determined on day 30 after operation. The results showed that fucoidan caused dose-dependently attenuation of mechanical allodynia and thermal hyperalgesia. Furthermore, fucoidan could markedly inhibit neuroimmune activation characterized by glial activation, production of cytokines as well as ERK activation. The analgesic effect of intrathecal fucoidan in rats receiving SNL might partly attribute to the inhibition of neuroimmune activation associated with the maintenance of neuropathic pain.

  1. Nanoscale imaging and mechanical analysis of Fc receptor-mediated macrophage phagocytosis against cancer cells.

    PubMed

    Li, Mi; Liu, Lianqing; Xi, Ning; Wang, Yuechao; Xiao, Xiubin; Zhang, Weijing

    2014-02-18

    Fc receptor-mediated macrophage phagocytosis against cancer cells is an important mechanism in the immune therapy of cancers. Traditional research about macrophage phagocytosis was based on optical microscopy, which cannot reveal detailed information because of the 200-nm-resolution limit. Quantitatively investigating the macrophage phagocytosis at micro- and nanoscale levels is still scarce. The advent of atomic force microscopy (AFM) offers an excellent analytical instrument for quantitatively investigating the biological processes at single-cell and single-molecule levels under native conditions. In this work, we combined AFM and fluorescence microscopy to visualize and quantify the detailed changes in cell morphology and mechanical properties during the process of Fc receptor-mediated macrophage phagocytosis against cancer cells. Lymphoma cells were discernible by fluorescence staining. Then, the dynamic process of phagocytosis was observed by time-lapse optical microscopy. Next, AFM was applied to investigate the detailed cellular behaviors during macrophage phagocytosis under the guidance of fluorescence recognition. AFM imaging revealed the distinct features in cellular ultramicrostructures for the different steps of macrophage phagocytosis. AFM cell mechanical property measurements indicated that the binding of cancer cells to macrophages could make macrophages become stiffer. The experimental results provide novel insights in understanding the Fc-receptor-mediated macrophage phagocytosis.

  2. The effect of corporal punishment and verbal abuse on delinquency: mediating mechanisms.

    PubMed

    Evans, Sara Z; Simons, Leslie Gordon; Simons, Ronald L

    2012-08-01

    While the link between parenting and delinquency is well established, there is less consensus among scholars with regards to the processes that account for this link. The current study had two objectives. The first was to disentangle the effects of African American parents' use of corporal punishment and verbal abuse on the conduct problems of their preteen children. The second was to investigate the mechanisms that explain this relationship, such as having low self-control or a hostile view of relationships, whereby these harsh parenting practices increase a youth's involvement in problem behavior. Further, we are interested in specifically addressing how these mechanisms may operate differently for males versus females. Analyses utilized structural equation modeling and longitudinal data spanning approximately 2.5 years from a sample of 704 (54.2 % female) African American children ages 10-12. The results indicated that verbal abuse was a more important predictor of conduct problems than corporal punishment. Additionally, we found that the mechanisms that mediated the impact of verbal abuse and corporal punishment on conduct problems varied by gender. For males, most of the effect of verbal abuse was mediated by low self-control, whereas anger/frustration was the primary mediator for females. Implications of these results and directions for future study are also discussed.

  3. Multiple mechanisms mediate glucose repression of the yeast GAL1 gene.

    PubMed Central

    Lamphier, M S; Ptashne, M

    1992-01-01

    Several mechanisms contribute to the glucose repression of the GAL1 gene in Saccharomyces cerevisiae. We show that one mechanism involves the transcriptional down-regulation of the GAL4 gene and a second requires the GAL80 gene. We also examine the contribution of cis-acting negative elements in the GAL1 promoter to glucose repression. In an otherwise wild-type strain disruption of any one of these three mechanisms alleviates repression of GAL1 only 2- to 4-fold. However, in the absence of the other two mechanisms the transcriptional down-regulation of GAL4 is sufficient to repress GAL1 expression 40- to 60-fold and the GAL80-dependent mechanism is sufficient to repress GAL1 expression 20- to 30-fold. These first two mechanisms constitute a functionally redundant system of repression and both must be disrupted in order to abolish glucose repression of GAL1. In contrast, negative elements in the GAL1 promoter are effective in repressing GAL1 expression 2- to 4-fold in glucose medium only when at least one of the other two mechanisms of repression is present. Thus, glucose repression of GAL1 is mediated primarily by the first two mechanisms, whereas the third mechanism supplements repression severalfold. PMID:1631075

  4. Influence of structural load-bearing scaffolds on mechanical load- and BMP-2-mediated bone regeneration.

    PubMed

    McDermott, Anna M; Mason, Devon E; Lin, Angela S P; Guldberg, Robert E; Boerckel, Joel D

    2016-09-01

    A common design constraint in functional tissue engineering is that scaffolds intended for use in load-bearing sites possess similar mechanical properties to the replaced tissue. Here, we tested the hypothesis that in vivo loading would enhance bone morphogenetic protein-2 (BMP-2)-mediated bone regeneration in the presence of a load-bearing PLDL scaffold, whose pores and central core were filled with BMP-2-releasing alginate hydrogel. First, we evaluated the effects of in vivo mechanical loading on bone regeneration in the structural scaffolds. Second, we compared scaffold-mediated bone regeneration, independent of mechanical loading, with alginate hydrogel constructs, without the structural scaffold, that have been shown previously to facilitate in vivo mechanical stimulation of bone formation. Contrary to our hypothesis, mechanical loading had no effect on bone formation, distribution, or biomechanical properties in structural scaffolds. Independent of loading, the structural scaffolds reduced bone formation compared to non-structural alginate, particularly in regions in which the scaffold was concentrated, resulting in impaired functional regeneration. This is attributable to a combination of stress shielding by the scaffold and inhibition of cellular infiltration and tissue ingrowth. Collectively, these data question the necessity of scaffold similarity to mature tissue at the time of implantation and emphasize development of an environment conducive to cellular activation of matrix production and ultimate functional regeneration.

  5. Chronic Oral Pelargonidin Alleviates Streptozotocin-Induced Diabetic Neuropathic Hyperalgesia in Rat: Involvement of Oxidative Stress

    PubMed Central

    Mirshekar, Mohammadali; Roghani, Mehrdad; Khalili, Mohsen; Baluchnejadmojarad, Tourandokht; Arab Moazzen, Saiedeh

    2010-01-01

    Background: Diabetes mellitus in some clinical cases is accompanied with hyperalgesia. In this study, we evaluated the possible beneficial effect of chronic pelargonidin (PG) treatment on hyperalgesia in streptozotocin (STZ)-diabetic neuropathic rat. Methods: Male Wistar rats (n = 56) were divided into seven groups, i.e. control, diabetic, PG-treated control, PG (single- and multiple-dose)-treated diabetic, and sodium salicylate-treated control and diabetics. For induction of diabetes, STZ was injected i.p. at a single dose of 60 mg/kg. PG was orally administered at a dose of 10 mg/kg once and/or on alternate days for 8 weeks; 1 week after diabetes induction. After two months, hyperalgesia was assessed using standard formalin and hot tail immersion tests. Meanwhile, markers of oxidative stress in brain were measured. One-way analysis of variance was used for statistical analysis of the data. Results: Diabetic rats showed a marked chemical and thermal hyperalgesia, indicating that development of diabetic neuropathy and PG treatment (especially multiple-doses) significantly ameliorated the alteration in hyperalgesia (P<0.05-0.01) in diabetic rats as compared to untreated diabetics. PG (multiple doses) also significantly decreased diabetes-induced thiobarbituric acid reactive substances formation and non-significantly reversed elevation of nitrite level and reduction of antioxidant defensive enzyme superoxide dismutase. Conclusion: These results clearly suggest that PG prevents diabetic neuropathic hyperalgesia through attenuation of oxidative stress. PMID:20683496

  6. Is the Volume of the Caudate Nuclei Associated With Area of Secondary Hyperalgesia? – Protocol for a 3-Tesla MRI Study of Healthy Volunteers

    PubMed Central

    Asghar, Mohammad Sohail; Wetterslev, Jørn; Pipper, Christian Bressen; Johan Mårtensson, Johan; Becerra, Lino; Christensen, Anders; Nybing, Janus Damm; Havsteen, Inger; Boesen, Mikael; Dahl, Jørgen Berg

    2016-01-01

    Background Experience and development of pain may be influenced by a number of physiological, psychological, and psychosocial factors. In a previous study we found differences in neuronal activation to noxious stimulation, and microstructural neuroanatomical differences, when comparing healthy volunteers with differences in size of the area of secondary hyperalgesia following a standardized burn injury. Objective We aim to investigate the degree of association between the volume of pain-relevant structures in the brain and the size of the area of secondary hyperalgesia following brief thermal sensitization. Methods The study consists of one experimental day, in which whole-brain magnetic resonance imaging (MRI) scans will be conducted including T1-weighed three-dimensional anatomy scan, diffusion tensor imaging, and resting state functional MRI. Before the experimental day, all included participants will undergo experimental pain testing in a parallel study (Clinicaltrials.gov Identifier: NCT02527395). Results from this experimental pain testing, as well as the size of the area of secondary hyperalgesia from the included participants, will be extracted from this parallel study. Results The association between the volume of pain-relevant structures in the brain and the area of secondary hyperalgesia will be investigated by linear regression of the estimated best linear unbiased predictors on the individual volumes of the pain relevant brain structures. Conclusions We plan to investigate the association between experimental pain testing parameters and the volume, connectivity, and resting state activity of pain-relevant structures in the brain. These results may improve our knowledge of the mechanisms responsible for the development of acute and chronic pain. ClinicalTrial Danish Research Ethics Committee (identifier: H-15010473). Danish Data Protection Agency (identifier: RH-2015-149). Clinicaltrials.gov NCT02567318; http://clinicaltrials.gov/ct2/show/NCT02567318

  7. Evidence that the human cutaneous venoarteriolar response is not mediated by adrenergic mechanisms

    NASA Technical Reports Server (NTRS)

    Crandall, C. G.; Shibasaki, M.; Yen, T. C.

    2002-01-01

    The venoarteriolar response causes vasoconstriction to skin and muscle via local mechanisms secondary to venous congestion. The purpose of this project was to investigate whether this response occurs through alpha-adrenergic mechanisms. In supine individuals, forearm skin blood flow was monitored via laser-Doppler flowmetry over sites following local administration of terazosin (alpha(1)-antagonist), yohimbine (alpha(2)-antagonist), phentolamine (non-selective alpha-antagonist) and bretylium tosylate (inhibits neurotransmission of adrenergic nerves) via intradermal microdialysis or intradermal injection. In addition, skin blood flow was monitored over an area of forearm skin that was locally anaesthetized via application of EMLA (2.5 % lidocaine (lignocaine) and 2.5 % prilocaine) cream. Skin blood flow was also monitored over adjacent sites that received the vehicle for the specified drug. Each trial was performed on a minimum of seven subjects and on separate days. The venoarteriolar response was engaged by lowering the subject's arm from heart level such that the sites of skin blood flow measurement were 34 +/- 1 cm below the heart. The arm remained in this position for 2 min. Selective and non-selective alpha-adrenoceptor antagonism and presynaptic inhibition of adrenergic neurotransmission did not abolish the venoarteriolar response. However, local anaesthesia blocked the venoarteriolar response without altering alpha-adrenergic mediated vasoconstriction. These data suggest that the venoarteriolar response does not occur through adrenergic mechanisms as previously reported. Rather, the venoarteriolar response may due to myogenic mechanisms associated with changes in vascular pressure or is mediated by a non-adrenergic, but neurally mediated, local mechanism.

  8. Development of mechanisms associated with neurogenic-mediated skin inflammation during the growth of rats.

    PubMed

    Ohshima, Mihoko; Miyake, Mio; Takeda, Masanori; Muto, Taichiro; Ueda, Norishi; Ito, Komei; Sakamoto, Tatsuo

    2010-04-01

    Neurogenic-mediated inflammation may be associated with several inflammatory skin diseases including atopic dermatitis. However, age-dependent differences in neurogenic-mediated skin responses are not fully understood. We compared skin plasma leakage in rats aged 2 and 8 wk, which was induced by topical capsaicin, topical formalin, and intracutaneous substance P, whose effects are mediated via tachykinin NK1 receptors. Evans blue dye extravasation served as an index of the increase in skin vascular permeability. Capsaicin, formalin, and substance P caused a skin response in a dose-dependent manner in both age groups. However, the skin response was much greater in adults than in pups. In addition, the localization of sensory C-fibers and tachykinin NK1 receptors in the skin was investigated by immunofluorescent staining with antisubstance P and antitachykinin NK1 receptor antibodies, respectively. Substance P-immunoreactive nerves were detected throughout the dermis and tachykinin NK1 receptors were mainly detected in blood vessel walls in the dermis in both age groups. However, they were more sparsely distributed in pups. In conclusion, the weak neurogenic-mediated skin inflammation in pups is probably because of immature neural mechanisms associated with skin inflammation such as reduced innervation of sensory C-fibers and low expression of tachykinin NK1 receptors.

  9. Chemically- and mechanically-mediated influences on the transport and mechanical characteristics of rock fractures

    SciTech Connect

    Min, K.-B.; Rutqvist, J.; Elsworth, D.

    2009-02-01

    A model is presented to represent changes in the mechanical and transport characteristics of fractured rock that result from coupled mechanical and chemical effects. The specific influence is the elevation of dissolution rates on contacting asperities, which results in a stress- and temperature-dependent permanent closure. A model representing this pressure-dissolution-like behavior is adapted to define the threshold and resulting response in terms of fundamental thermodynamic properties of a contacting fracture. These relations are incorporated in a stress-stiffening model of fracture closure to define the stress- and temperature-dependency of aperture loss and behavior during stress and temperature cycling. These models compare well with laboratory and field experiments, representing both decoupled isobaric and isothermal responses. The model was applied to explore the impact of these responses on heated structures in rock. The result showed a reduction in ultimate induced stresses over the case where chemical effects were not incorporated, with permanent reduction in final stresses after cooling to ambient conditions. Similarly, permeabilities may be lower than they were in the case where chemical effects were not considered, with a net reduction apparent even after cooling to ambient temperature. These heretofore-neglected effects may have a correspondingly significant impact on the performance of heated structures in rock, such as repositories for the containment of radioactive wastes.

  10. Investigating dynamic structural and mechanical changes of neuroblastoma cells associated with glutamate-mediated neurodegeneration

    NASA Astrophysics Data System (ADS)

    Fang, Yuqiang; Iu, Catherine Y. Y.; Lui, Cathy N. P.; Zou, Yukai; Fung, Carmen K. M.; Li, Hung Wing; Xi, Ning; Yung, Ken K. L.; Lai, King W. C.

    2014-11-01

    Glutamate-mediated neurodegeneration resulting from excessive activation of glutamate receptors is recognized as one of the major causes of various neurological disorders such as Alzheimer's and Huntington's diseases. However, the underlying mechanisms in the neurodegenerative process remain unidentified. Here, we investigate the real-time dynamic structural and mechanical changes associated with the neurodegeneration induced by the activation of N-methyl-D-aspartate (NMDA) receptors (a subtype of glutamate receptors) at the nanoscale. Atomic force microscopy (AFM) is employed to measure the three-dimensional (3-D) topography and mechanical properties of live SH-SY5Y cells under stimulus of NMDA receptors. A significant increase in surface roughness and stiffness of the cell is observed after NMDA treatment, which indicates the time-dependent neuronal cell behavior under NMDA-mediated neurodegeneration. The present AFM based study further advance our understanding of the neurodegenerative process to elucidate the pathways and mechanisms that govern NMDA induced neurodegeneration, so as to facilitate the development of novel therapeutic strategies for neurodegenerative diseases.

  11. Estrogens facilitate memory processing through membrane mediated mechanisms and alterations in spine density

    PubMed Central

    Luine, Victoria N.; Frankfurt, Maya

    2012-01-01

    Estrogens exert sustained, genomically mediated effects on memory throughout the female life cycle, but here we review new studies documenting rapid effects of estradiol on memory, which are exerted through membrane-mediated mechanisms. Use of recognition memory tasks in rats, shows that estrogens enhance memory consolidation within one hour. 17α-estradiol is more potent than 17β-estradiol, and the dose response relationship between estrogens and memory is an inverted U shape. Use of specific estrogen receptor (ER) agonists suggests mediation by an ERβ-like membrane receptor. Enhanced memory is associated with increased spine density and altered noradrenergic activity in the medial prefrontal cortex and hippocampus within 30 min. of administration. The environmental chemical, bisphenol-A, rapidly antagonizes enhancements in memory in both sexes possibly through actions on spines. Thus, estradiol and related compounds exert rapid alterations in cognition through non-genomic mechanisms, a finding which may provide a basis for better understanding and treating memory impairments. PMID:22981654

  12. Empowerment beliefs and intention to uptake cervical cancer screening: three psychosocial mediating mechanisms.

    PubMed

    Luszczynska, Aleksandra; Durawa, Alicja B; Scholz, Urte; Knoll, Nina

    2012-01-01

    Three studies tested if the associations between women's empowerment beliefs and intentions to attend cervical cancer screening could be explained by mediating psychological mechanisms: control-related beliefs, well being-related beliefs, and beliefs and evaluations referring to social functioning. Data were collected from January to March 2011 in the rural and urban areas across regions of Poland. Study 1 (N = 386) indicated that women with strong empowerment harbored stronger self-efficacy and beliefs that screening participation would make them feel in control of their own health and body. These two types of cognitions were, in turn, associated with stronger cervical cancer screening intentions. Results of Study 2 (N = 527) confirmed three significant well being-related mediators in the relationship between empowerment beliefs and cervical cancer screening: perceived benefits of screening related to well being, appearance satisfaction, discomfort- and shame-related barriers for screening. Finally, Study 3 (N = 424) showed that empowerment enabled receiving higher social support for cervical cancer screening, promoted perceiving fewer barriers for cervical cancer screening-related communication and more social benefits of engaging in cervical cancer screening. Support for cervical cancer screening, social barriers, and benefits were, in turn, related to screening intentions. Across the studies similar shares of intention variance were explained, and thus the hypothesized mediating mechanisms may have similar explanatory power.

  13. CXCL12/CXCR4 chemokine signaling in spinal glia induces pain hypersensitivity through MAPKs-mediated neuroinflammation in bone cancer rats.

    PubMed

    Hu, Xue-Ming; Liu, Yan-Nan; Zhang, Hai-Long; Cao, Shou-Bin; Zhang, Ting; Chen, Li-Ping; Shen, Wen

    2015-02-01

    The activation of MAPK pathways in spinal cord and subsequent production of proinflammatory cytokines in glial cells contribute to the development of spinal central sensitization, the basic mechanism underlying bone cancer pain (BCP). Our previous study showed that spinal CXCL12 from astrocytes mediates BCP generation by binding to CXCR4 in both astrocyters and microglia. Here, we verified that CXCL12/CXCR4 signaling contributed to BCP through a MAPK-mediated mechanism. In naïve rats, a single intrathecal administration of CXCL12 considerably induced pain hyperalgesia and phosphorylation expression of spinal MAPK members (including extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase), which could be partially prevented by pre-treatment with CXCR4 inhibitor AMD3100. This CXCL12-induced hyperalgesia was also reduced by MAPK inhibitors. In bone cancer rats, tumor cell inoculation into the tibial cavity caused prominent and persistent pain hyperalgesia, and associated with up-regulation of CXCL12 and CXCR4, activation of glial cells, phosphorylation of MAPKs, and production of proinflammatory cytokines in the spinal cord. These tumor cell inoculation-induced behavioral and neurochemical alterations were all suppressed by blocking CXCL12/CXCR4 signaling or MAPK pathways. Taken together, these results demonstrate that spinal MAPK pathways mediated CXCL12/CXCR4-induced pain hypersensitivity in bone cancer rats, which could be druggable targets for alleviating BCP and glia-derived neuroinflammation. Following tumor cell inoculation, chemokine CXCL12 from astrocytes spreads around the spinal environment, resulting in functional activation of CXCR4-expressing astrocytes and microglia. Once glia are activated, they may initiate MAPK (mitogen-activated protein kinase) pathways, and subsequently produce proinflammatory cytokines and chemokines. Among them, CXCL12 could reinforce the astrocytic and microglial activation in autocrine and paracrine manners

  14. Mechanisms of coronavirus cell entry mediated by the viral spike protein.

    PubMed

    Belouzard, Sandrine; Millet, Jean K; Licitra, Beth N; Whittaker, Gary R

    2012-06-01

    Coronaviruses are enveloped positive-stranded RNA viruses that replicate in the cytoplasm. To deliver their nucleocapsid into the host cell, they rely on the fusion of their envelope with the host cell membrane. The spike glycoprotein (S) mediates virus entry and is a primary determinant of cell tropism and pathogenesis. It is classified as a class I fusion protein, and is responsible for binding to the receptor on the host cell as well as mediating the fusion of host and viral membranes-A process driven by major conformational changes of the S protein. This review discusses coronavirus entry mechanisms focusing on the different triggers used by coronaviruses to initiate the conformational change of the S protein: receptor binding, low pH exposure and proteolytic activation. We also highlight commonalities between coronavirus S proteins and other class I viral fusion proteins, as well as distinctive features that confer distinct tropism, pathogenicity and host interspecies transmission characteristics to coronaviruses.

  15. CSRP's Impact on low-income preschoolers' preacademic skills: self-regulation as a mediating mechanism.

    PubMed

    Raver, C Cybele; Jones, Stephanie M; Li-Grining, Christine; Zhai, Fuhua; Bub, Kristen; Pressler, Emily

    2011-01-01

    Based on theoretically driven models, the Chicago School Readiness Project (CSRP) targeted low-income children's school readiness through the mediating mechanism of self-regulation. The CSRP is a multicomponent, cluster-randomized efficacy trial implemented in 35 Head Start-funded classrooms (N = 602 children). The analyses confirm that the CSRP improved low-income children's self-regulation skills (as indexed by attention/impulse control and executive function) from fall to spring of the Head Start year. Analyses also suggest significant benefits of CSRP for children's preacademic skills, as measured by vocabulary, letter-naming, and math skills. Partial support was found for improvement in children's self-regulation as a hypothesized mediator for children's gains in academic readiness. Implications for programs and policies that support young children's behavioral health and academic success are discussed.

  16. Structural mechanism of nuclear transport mediated by importin β and flexible amphiphilic proteins.

    PubMed

    Yoshimura, Shige H; Kumeta, Masahiro; Takeyasu, Kunio

    2014-12-02

    Karyopherin β family proteins mediate the nuclear/cytoplasmic transport of various proteins through the nuclear pore complex (NPC), although they are substantially larger than the size limit of the NPC.To elucidate the molecular mechanism underlying this paradoxical function, we focused on the unique structures called HEAT repeats, which consist of repetitive amphiphilic α helices. An in vitro transport assay and FRAP analyses demonstrated that not only karyopherin β family proteins but also other proteins with HEAT repeats could pass through the NPC by themselves, and serve as transport mediators for their binding partners. Biochemical and spectroscopic analyses and molecular dynamics simulations of purified HEAT-rich proteins revealed that they interact with hydrophobic groups, including phenyl and alkyl groups, and undergo reversible conformational changes in tertiary structures, but not in secondary structures. These results show that conformational changes in the flexible amphiphilic motifs play a critical role in translocation through the NPC.

  17. Clathrin-mediated endocytosis is the dominant mechanism of vesicle retrieval at hippocampal synapses.

    PubMed

    Granseth, Björn; Odermatt, Benjamin; Royle, Stephen J; Lagnado, Leon

    2006-09-21

    The maintenance of synaptic transmission requires that vesicles be recycled after releasing neurotransmitter. Several modes of retrieval have been proposed to operate at small synaptic terminals of central neurons, including a fast "kiss-and-run" mechanism that releases neurotransmitter through a fusion pore. Using an improved fluorescent reporter comprising pHluorin fused to synaptophysin, we find that only a slow mode of endocytosis (tau = 15 s) operates at hippocampal synapses when vesicle fusion is triggered by a single nerve impulse or short burst. This retrieval mechanism is blocked by overexpression of the C-terminal fragment of AP180 or by knockdown of clathrin using RNAi, and it is associated with the movement of clathrin and vesicle proteins out of the synapse. These results indicate that clathrin-mediated endocytosis is the major, if not exclusive, mechanism of vesicle retrieval after physiological stimuli.

  18. Chirality-Induced Budding: A Raft-Mediated Mechanism for Endocytosis and Morphology of Caveolae?

    PubMed Central

    Sarasij, R. C.; Mayor, Satyajit; Rao, Madan

    2007-01-01

    The formation of transport carriers (spherical vesicles and tubules) involves membrane budding, growth, and ultimately fission. We propose a mechanism of membrane budding, wherein the tilt and chirality of constituent molecules, confined to a patch of area A, induces buds of ∼50–100 nm that are comparable to vesicles involved in endocytosis. Because such chiral and tilted lipid molecules are likely to exist in “rafts”, we suggest the involvement of this mechanism in generating membrane buds in the clathrin and dynamin-independent, raft-component mediated endocytosis of glycosylphosphatidylinositol-anchored proteins. We argue that caveolae, permanent cell surface structures with characteristic morphology and enriched in raft constituents, are also likely to be formed by this mechanism. Thus, molecular chirality and tilt, and its expression over large spatial scales may be a common organizing principle in membrane budding of transport carriers. PMID:17237196

  19. Sensitization of neonatal rat lumbar motoneuron by the inflammatory pain mediator bradykinin

    PubMed Central

    Bouhadfane, Mouloud; Kaszás, Attila; Rózsa, Balázs; Harris-Warrick, Ronald M; Vinay, Laurent; Brocard, Frédéric

    2015-01-01

    Bradykinin (Bk) is a potent inflammatory mediator that causes hyperalgesia. The action of Bk on the sensory system is well documented but its effects on motoneurons, the final pathway of the motor system, are unknown. By a combination of patch-clamp recordings and two-photon calcium imaging, we found that Bk strongly sensitizes spinal motoneurons. Sensitization was characterized by an increased ability to generate self-sustained spiking in response to excitatory inputs. Our pharmacological study described a dual ionic mechanism to sensitize motoneurons, including inhibition of a barium-sensitive resting K+ conductance and activation of a nonselective cationic conductance primarily mediated by Na+. Examination of the upstream signaling pathways provided evidence for postsynaptic activation of B2 receptors, G protein activation of phospholipase C, InsP3 synthesis, and calmodulin activation. This study questions the influence of motoneurons in the assessment of hyperalgesia since the withdrawal motor reflex is commonly used as a surrogate pain model. DOI: http://dx.doi.org/10.7554/eLife.06195.001 PMID:25781633

  20. Mechanisms of acetylcholine-mediated vasodilatation in young and aged human skin

    PubMed Central

    Holowatz, Lacy A; Thompson, Caitlin S; Minson, Christopher T; Kenney, W Larry

    2005-01-01

    Thermoregulatory cutaneous vasodilatation (VD) is attenuated in aged skin. While acetylcholine (ACh) plays a role in thermally mediated VD, the precise mechanisms through which ACh-mediated VD acts and whether those downstream mechanisms change with ageing are unclear. We tested the hypotheses that both nitric oxide (NO)- and prostanoid-mediated pathways contribute to exogenous ACh-mediated VD, and that both are attenuated with advanced age. Twelve young (Y: 23 ± 1 years) and 10 older (O: 69 ± 1 years) subjects underwent infusions of 137.5 μm ACh at four intradermal microdialysis sites: control (C, Ringer solution), NO synthase inhibited (NOS-I, 10 mml-NAME), cyclooxygenase inhibited (COX-I, 10 mm ketorolac) and NOS-I + COX-I. Red blood cell flux was monitored using laser-Doppler flowmetry, and cutaneous vascular conductance (CVC) was calculated (laser-Doppler flux/mean arterial pressure) and normalized to maximal CVC (%CVCmax) (28 mm sodium nitroprusside + local heating to 43°C). Baseline %CVCmax was increased in the O at COX-I sites (COX-I 16 ± 1, NOS-I + COX-I 16 ± 2 versus C 10 ± 1%CVCmax; P < 0.001) but not in the young, suggesting an age-related shift toward COX vasoconstrictors contributing to basal cutaneous vasomotor tone. There was no difference in peak %CVCmax during ACh infusion between age groups, and the response was unchanged by NOS-I (O: NOS-I 35 ± 5 versus C 38 ± 5%CVCmax; P = 0.84) (Y: NOS-I 41 ± 4 versus C 39 ± 4%CVCmax; P = 0.67). COX-I and NOS-I + COX-I attenuated the peak CVC response to ACh in both groups (COX-I O: 29 ± 3, Y: 22 ± 2%CVCmaxversus C; P < 0.001 both groups; NOS-I + COX-I O: 32 ± 3 versus Y: 29 ± 2%CVCmax; versus C; P < 0.001 both groups). ACh mediates cutaneous VD through prostanoid and non-NO-, non-prostanoid-dependent pathways. Further, older subjects have a diminished prostanoid contribution to ACh-mediated VD. PMID:15661816

  1. Crotalphine desensitizes TRPA1 ion channels to alleviate inflammatory hyperalgesia.

    PubMed

    Bressan, Elisangela; Touska, Filip; Vetter, Irina; Kistner, Katrin; Kichko, Tatjana I; Teixeira, Nathália B; Picolo, Gisele; Cury, Yara; Lewis, Richard J; Fischer, Michael J M; Zimmermann, Katharina; Reeh, Peter W

    2016-11-01

    Crotalphine is a structural analogue to a novel analgesic peptide that was first identified in the crude venom from the South American rattlesnake Crotalus durissus terrificus. Although crotalphine's analgesic effect is well established, its direct mechanism of action remains unresolved. The aim of the present study was to investigate the effect of crotalphine on ion channels in peripheral pain pathways. We found that picomolar concentrations of crotalphine selectively activate heterologously expressed and native TRPA1 ion channels. TRPA1 activation by crotalphine required intact N-terminal cysteine residues and was followed by strong and long-lasting desensitization of the channel. Homologous desensitization of recombinant TRPA1 and heterologous desensitization in cultured dorsal root ganglia neurons was observed. Likewise, crotalphine acted on peptidergic TRPA1-expressing nerve endings ex vivo as demonstrated by suppression of calcitonin gene-related peptide release from the trachea and in vivo by inhibition of chemically induced and inflammatory hypersensitivity in mice. The crotalphine-mediated desensitizing effect was abolished by the TRPA1 blocker HC030031 and absent in TRPA1-deficient mice. Taken together, these results suggest that crotalphine is the first peptide to mediate antinociception selectively and at subnanomolar concentrations by targeting TRPA1 ion channels.

  2. Critical Role for Telomerase in the Mechanism of Flow-Mediated Dilation in the Human Microcirculation

    PubMed Central

    Freed, Julie K.; Durand, Matthew J.; Riedel, Michael; Ait-Aissa, Karima; Green, Paula; Hockenberry, Joseph C.; Morgan, R. Garret; Donato, Anthony J.; Peleg, Refael; Gasparri, Mario; Rokkas, Chris K.; Santos, Janine H.; Priel, Esther; Gutterman, David D.

    2016-01-01

    Rationale: Telomerase is a nuclear regulator of telomere elongation with recent reports suggesting a role in regulation of mitochondrial reactive oxygen species. Flow-mediated dilation in patients with cardiovascular disease is dependent on the formation of reactive oxygen species. Objective: We examined the hypothesis that telomerase activity modulates microvascular flow-mediated dilation, and loss of telomerase activity contributes to the change of mediator from nitric oxide to mitochondrial hydrogen peroxide in patients with coronary artery disease (CAD). Methods and Results: Human coronary and adipose arterioles were isolated for videomicroscopy. Flow-mediated dilation was measured in vessels pretreated with the telomerase inhibitor BIBR-1532 or vehicle. Statistical differences between groups were determined using a 2-way analysis of variance repeated measure (n≥4; P<0.05). L-NAME (Nω-nitro-L-arginine methyl ester; nitric oxide synthase inhibitor) abolished flow-mediated dilation in arterioles from subjects without CAD, whereas polyethylene glycol-catalase (PEG-catalase; hydrogen peroxide scavenger) had no effect. After exposure to BIBR-1532, arterioles from non-CAD subjects maintained the magnitude of dilation but changed the mediator from nitric oxide to mitochondrial hydrogen peroxide (% max diameter at 100 cm H2O: vehicle 74.6±4.1, L-NAME 37.0±2.0*, PEG-catalase 82.1±2.8; BIBR-1532 69.9±4.0, L-NAME 84.7±2.2, PEG-catalase 36.5±6.9*). Conversely, treatment of microvessels from CAD patients with the telomerase activator AGS 499 converted the PEG-catalase-inhibitable dilation to one mediated by nitric oxide (% max diameter at 100 cm H2O: adipose, AGS 499 78.5±3.9; L-NAME 10.9±17.5*; PEG-catalase 79.2±4.9). Endothelial-independent dilation was not altered with either treatment. Conclusions: We have identified a novel role for telomerase in re-establishing a physiological mechanism of vasodilation in arterioles from subjects with CAD. These findings

  3. Bnip3 Mediates Permeabilization of Mitochondria and Release of Cytochrome c via a Novel Mechanism

    PubMed Central

    Quinsay, Melissa N.; Lee, Youngil; Rikka, Shivaji; Sayen, M. Richard; Molkentin, Jeffery D.; Gottlieb, Roberta A.; Gustafsson, Åsa B.

    2010-01-01

    Bnip3 is a member of the BH3-only subfamily of pro-apoptotic Bcl-2 proteins and is associated with loss of cardiac myocytes after a myocardial infarction. Previous studies have demonstrated that Bnip3 induces mitochondrial dysfunction, but the mechanisms involved in this process remain unknown. In this study, we demonstrate that Bnip3 induces permeabilization of the mitochondria via a novel mechanism that is different from other BH3-only proteins. We found that Bnip3 induced mitochondrial swelling and cytochrome c release in isolated heart mitochondria in vitro. Another BH3-only protein, tBid, also caused release of cytochrome c but failed to induce swelling of mitochondria. Swelling of mitochondria is a characteristic of mitochondrial permeability transition pore (mPTP) opening, but Bnip3-mediated mitochondrial swelling was insensitive to cyclosporine A, an inhibitor of the mPTP and independent of cyclophilin D (cypD), an essential component of the mPTP. Bnip3 also induced permeabilization of the mitochondrial membranes as evident by calcein release from the matrix in both wild type (WT) and cypD deficient mouse embryonic fibroblasts (MEFs). Moreover, Bnip3 induced mitochondrial matrix remodeling and large amplitude swelling of the inner membrane, which led to disassembly of OPA1 complexes and release from the mitochondria. Thus, these studies suggest that Bnip3 mediates mitochondrial permeabilization by a novel mechanism that is different from other BH3-only proteins. PMID:20025887

  4. P-cadherin promotes collective cell migration via a Cdc42-mediated increase in mechanical forces

    PubMed Central

    Plutoni, Cédric; Bazellieres, Elsa; Le Borgne-Rochet, Maïlys; Comunale, Franck; Brugues, Agusti; Séveno, Martial; Planchon, Damien; Thuault, Sylvie; Morin, Nathalie; Bodin, Stéphane; Trepat, Xavier

    2016-01-01

    Collective cell migration (CCM) is essential for organism development, wound healing, and metastatic transition, the primary cause of cancer-related death, and it involves cell–cell adhesion molecules of the cadherin family. Increased P-cadherin expression levels are correlated with tumor aggressiveness in carcinoma and aggressive sarcoma; however, how P-cadherin promotes tumor malignancy remains unknown. Here, using integrated cell biology and biophysical approaches, we determined that P-cadherin specifically induces polarization and CCM through an increase in the strength and anisotropy of mechanical forces. We show that this mechanical regulation is mediated by the P-cadherin/β-PIX/Cdc42 axis; P-cadherin specifically activates Cdc42 through β-PIX, which is specifically recruited at cell–cell contacts upon CCM. This mechanism of cell polarization and migration is absent in cells expressing E- or R-cadherin. Thus, we identify a specific role of P-cadherin through β-PIX–mediated Cdc42 activation in the regulation of cell polarity and force anisotropy that drives CCM. PMID:26783302

  5. Mechanism of HSV infection through soluble adapter-mediated virus bridging to the EGF receptor

    SciTech Connect

    Nakano, Kenji; Kobayashi, Masatoshi; Nakamura, Kei-ichiro; Nakanishi, Takeshi; Asano, Ryutaro; Kumagai, Izumi; Tahara, Hideaki; Kuwano, Michihiko; Cohen, Justus B.; Glorioso, Joseph C.

    2011-04-25

    Herpes simplex virus entry into cells requires the binding of envelope glycoprotein D (gD) to an entry receptor. Depending on the cell, entry occurs by different mechanisms, including fusion at the cell surface or endocytosis. Here we examined the entry mechanism through a non-HSV receptor mediated by a soluble bi-specific adapter protein composed of recognition elements for gD and the EGF receptor (EGFR). Virus entered into endosomes using either EGF or an EGFR-specific single chain antibody (scFv) for receptor recognition. Infection was less efficient with the EGF adapter which could be attributed to its weaker binding to a viral gD. Infection mediated by the scFv adapter was pH sensitive, indicating that gD-EGFR bridging alone was insufficient for capsid release from endosomes. We also show that the scFv adapter enhanced infection of EGFR-expressing tumor tissue in vivo. Our results indicate that adapters may retarget HSV infection without drastically changing the entry mechanism.

  6. Two different mechanisms mediate chemotaxis to inorganic phosphate in Pseudomonas aeruginosa

    PubMed Central

    Rico-Jiménez, Miriam; Reyes-Darias, Jose Antonio; Ortega, Álvaro; Díez Peña, Ana Isabel; Morel, Bertrand; Krell, Tino

    2016-01-01

    Inorganic phosphate (Pi) is a central signaling molecule that modulates virulence in various pathogens. In Pseudomonas aeruginosa, low Pi concentrations induce transcriptional alterations that increase virulence. Also, under low Pi levels, P. aeruginosa exhibits Pi chemotaxis—a process mediated by the two non-paralogous receptors CtpH and CtpL. Here we show that the two receptors operate via different mechanisms. We demonstrate that the ligand binding domain (LBD) of CtpH but not CtpL binds Pi directly. We identify the periplasmic ligand binding protein PstS as the protein that binds in its Pi loaded state to CtpL, resulting in receptor stimulation. PstS forms part of the Pi transporter and has thus a double function in Pi transport and chemotaxis. The affinity of Pi for CtpH was modest whereas that for PstS very high, which may explain why CtpH and CtpL mediate chemotaxis to high and low Pi concentrations, respectively. The pstS/ctpH double mutant was almost devoid of Pi taxis, indicating that PstS is the only CtpL Pi-shuttle. Chemotaxis mechanisms based on indirect ligand recognition were unambiguously identified in enterobacteria. The discovery of a similar mechanism in a different bacterial order, involving a different chemoreceptor type and chemoeffector suggests that such systems are widespread. PMID:27353565

  7. Reduced expression IRF7 in nasal epithelial cells from smokers as a potential mechanism mediating enhanced susceptibility to influenza

    EPA Science Inventory

    Rationale: Smokers are more susceptible to viral infections, including influenza virus, yet the mechanisms mediating this effect are not known. Methods: We have established an in vitro model of differentiated nasal epithelial cells from smokers, which maintain enhanced levels...

  8. Targeting Opioid-Induced Hyperalgesia in Clinical Treatment: Neurobiological Considerations.

    PubMed

    Arout, Caroline A; Edens, Ellen; Petrakis, Ismene L; Sofuoglu, Mehmet

    2015-06-01

    Opioid analgesics have become a cornerstone in the treatment of moderate to severe pain, resulting in a steady rise of opioid prescriptions. Subsequently, there has been a striking increase in the number of opioid-dependent individuals, opioid-related overdoses, and fatalities. Clinical use of opioids is further complicated by an increasingly deleterious profile of side effects beyond addiction, including tolerance and opioid-induced hyperalgesia (OIH), where OIH is defined as an increased sensitivity to already painful stimuli. This paradoxical state of increased nociception results from acute and long-term exposure to opioids, and appears to develop in a substantial subset of patients using opioids. Recently, there has been considerable interest in developing an efficacious treatment regimen for acute and chronic pain. However, there are currently no well-established treatments for OIH. Several substrates have emerged as potential modulators of OIH, including the N-methyl-D-aspartate and γ-aminobutyric acid receptors, and most notably, the innate neuroimmune system. This review summarizes the neurobiology of OIH in the context of clinical treatment; specifically, we review evidence for several pathways that show promise for the treatment of pain going forward, as prospective adjuvants to opioid analgesics. Overall, we suggest that this paradoxical state be considered an additional target of clinical treatment for chronic pain.

  9. LTR-mediated retroposition as a mechanism of RNA-based duplication in metazoans

    PubMed Central

    Tan, Shengjun; Cardoso-Moreira, Margarida; Shi, Wenwen; Zhang, Dan; Huang, Jiawei; Mao, Yanan; Jia, Hangxing; Zhang, Yaqiong; Chen, Chunyan; Shao, Yi; Leng, Liang; Liu, Zhonghua; Huang, Xun; Long, Manyuan

    2016-01-01

    In a broad range of taxa, genes can duplicate through an RNA intermediate in a process mediated by retrotransposons (retroposition). In mammals, L1 retrotransposons drive retroposition, but the elements responsible for retroposition in other animals have yet to be identified. Here, we examined young retrocopies from various animals that still retain the sequence features indicative of the underlying retroposition mechanism. In Drosophila melanogaster, we identified and de novo assembled 15 polymorphic retrocopies and found that all retroposed loci are chimeras of internal retrocopies flanked by discontinuous LTR retrotransposons. At the fusion points between the mRNAs and the LTR retrotransposons, we identified shared short similar sequences that suggest the involvement of microsimilarity-dependent template switches. By expanding our approach to mosquito, zebrafish, chicken, and mammals, we identified in all these species recently originated retrocopies with a similar chimeric structure and shared microsimilarities at the fusion points. We also identified several retrocopies that combine the sequences of two or more parental genes, demonstrating LTR-retroposition as a novel mechanism of exon shuffling. Finally, we found that LTR-mediated retrocopies are immediately cotranscribed with their flanking LTR retrotransposons. Transcriptional profiling coupled with sequence analyses revealed that the sense-strand transcription of the retrocopies often lead to the origination of in-frame proteins relative to the parental genes. Overall, our data show that LTR-mediated retroposition is highly conserved across a wide range of animal taxa; combined with previous work from plants and yeast, it represents an ancient and ongoing mechanism continuously shaping gene content evolution in eukaryotes. PMID:27934698

  10. T-cell-mediated drug hypersensitivity: immune mechanisms and their clinical relevance

    PubMed Central

    Cai, Fenfen; Lee, Frederick J; Pichler, Werner J

    2016-01-01

    T-cell-mediated drug hypersensitivity represents a significant proportion of immune mediated drug hypersensitivity reactions. In the recent years, there has been an increase in understanding the immune mechanisms behind T-cell-mediated drug hypersensitivity. According to hapten mechanism, drug specific T-cell response is stimulated by drug-protein conjugate presented on major histocompatibility complex (MHC) as it is presented as a new antigenic determinant. On the other hand, p-i concept suggests that a drug can stimulate T cells via noncovalent direct interaction with T-cell receptor and/or peptide-MHC. The drug binding site is quite variable and this leads to several different mechanisms within p-i concept. Altered peptide repertoire can be regarded as an 'atypical' subset of p-i concept since the mode of the drug binding and the binding site are essentially identical to p-i concept. However, the intracellular binding of abacavir to HLA-B*57:01 additionally results in alteration in peptide repertoire. Furthermore the T-cell response to altered peptide repertoire model is only shown for abacavir and HLA-B*57:01 and therefore it may not be generalised to other drug hypersensitivity. Danger hypothesis has been postulated to play an important role in drug hypersensitivity by providing signal 2 but its experimental data is lacking at this point in time. Furthermore, the recently described allo-immune response suggests that danger signal may be unnecessary. Finally, in view of these new understanding, the classification and the definition of type B adverse drug reaction should be revised. PMID:27141480

  11. Tips, Stalks, Tubes: Notch-Mediated Cell Fate Determination and Mechanisms of Tubulogenesis during Angiogenesis

    PubMed Central

    Tung, Jennifer J.; Tattersall, Ian W.; Kitajewski, Jan

    2012-01-01

    Angiogenesis is the process of developing vascular sprouts from existing blood vessels. Luminal endothelial cells convert into “tip” cells that contribute to the development of a multicellular stalk, which then undergoes lumen formation. In this review, we consider a variety of cellular and molecular pathways that mediate these transitions. We focus first on Notch signaling in cell fate determination as a mechanism to define tip and stalk cells. We next discuss the current models of lumen formation and describe new players in this process, such as chloride intracellular channel proteins. Finally, we consider the possible medical therapeutic benefits of understanding these processes and acknowledge potential obstacles in drug development. PMID:22355796

  12. Mechanisms mediating plant distributions across estuarine landscapes in a low-latitude tidal estuary.

    PubMed

    Guo, Hongyu; Pennings, Steven C

    2012-01-01

    Understanding of how plant communities are organized and will respond to global changes requires an understanding of how plant species respond to multiple environmental gradients. We examined the mechanisms mediating the distribution patterns of tidal marsh plants along an estuarine gradient in Georgia (USA) using a combination of field transplant experiments and monitoring. Our results could not be fully explained by the "competition-to-stress hypothesis" (the current paradigm explaining plant distributions across estuarine landscapes). This hypothesis states that the upstream limits of plant distributions are determined by competition, and the downstream limits by abiotic stress. We found that competition was generally strong in freshwater and brackish marshes, and that conditions in brackish and salt marshes were stressful to freshwater marsh plants, results consistent with the competition-to-stress hypothesis. Four other aspects of our results, however, were not explained by the competition-to-stress hypothesis. First, several halophytes found the freshwater habitat stressful and performed best (in the absence of competition) in brackish or salt marshes. Second, the upstream distribution of one species was determined by the combination of both abiotic and biotic (competition) factors. Third, marsh productivity (estimated by standing biomass) was a better predictor of relative biotic interaction intensity (RII) than was salinity or flooding, suggesting that productivity is a better indicator of plant stress than salinity or flooding gradients. Fourth, facilitation played a role in mediating the distribution patterns of some plants. Our results illustrate that even apparently simple abiotic gradients can encompass surprisingly complex processes mediating plant distributions.

  13. A novel posttranscriptional mechanism for dietary cholesterol-mediated suppression of liver LDL receptor expression.

    PubMed

    Singh, Amar Bahadur; Kan, Chin Fung Kelvin; Shende, Vikram; Dong, Bin; Liu, Jingwen

    2014-07-01

    It is well-established that over-accumulation of dietary cholesterol in the liver inhibits sterol-regulatory element binding protein (SREBP)-mediated LDL receptor (LDLR) gene transcription leading to a reduced hepatic LDLR mRNA level in hypercholesterolemic animals. However, it is unknown whether elevated cholesterol levels can elicit a cellular response to increase LDLR mRNA turnover to further repress LDLR expression in liver tissue. In the current study, we examined the effect of a high cholesterol diet on the hepatic expression of LDLR mRNA binding proteins in three different animal models and in cultured hepatic cells. Our results demonstrate that high cholesterol feeding specifically elevates the hepatic expression of LDLR mRNA decay promoting factor heterogeneous nuclear ribonucleoprotein (HNRNP)D without affecting expressions of other LDLR mRNA binding proteins in vivo and in vitro. Employing the approach of adenovirus-mediated gene knockdown, we further show that depletion of HNRNPD in the liver results in a marked reduction of serum LDL-cholesterol and a substantial increase in liver LDLR expression in hyperlipidemic mice. Additional studies of gene knockdown in albumin-luciferase-untranslated region (UTR) transgenic mice provide strong evidence supporting the essential role of 3'UTR in HNRNPD-mediated LDLR mRNA degradation in liver tissue. Altogether, this work identifies a novel posttranscriptional regulatory mechanism by which dietary cholesterol inhibits liver LDLR expression via inducing HNRNPD to accelerate LDLR mRNA degradation.

  14. Curcumin attenuates CFA induced thermal hyperalgesia by modulation of antioxidant enzymes and down regulation of TNF-α, IL-1β and IL-6.

    PubMed

    Singh, Ajeet Kumar; Vinayak, Manjula

    2015-03-01

    Reactive oxygen species are signaling mediators of nociceptive pathways. Exogenous administrations of antioxidants show anti-hyperalgesic effect. However, very little is known about the role of endogenous antioxidant defense system in pain pathology. Curcumin is a dietary antioxidant which shows ameliorative effect on thermal hypersensitivity, however detailed study is lacking. Present study was aimed to analyze the changes in oxidative stress, modulation of antioxidant enzymes and pro-inflammatory cytokines in complete Freund's adjuvant induced inflammatory hyperalgesia and the effect of curcumin on antioxidant defense system and pro-inflammatory cytokines. Anti-hyperalgesic activity of curcumin was evidenced after 6 h of treatment. Oxidative stress was evidenced in paw skin and spinal cord of hyperalgesic rats by high level of lipid peroxidation. A decrease in activity of antioxidant enzymes like catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase and an increase in level of pro-inflammatory cytokines TNF-α, IL-1β and IL-6 in paw skin was observed as compared to normal rats. However, activity of antioxidant enzymes was enhanced in spinal cord. The changes were brought towards normal level after curcumin treatment. The results suggest that modulation of antioxidant defense system is early event in initiation of inflammatory hyperalgesia which might lead to initiation of other signaling pathways mediated by lipid peroxide, TNF-α, IL-1β and IL-6. Decrease in oxidative stress and down regulation of these cytokines by curcumin is suggested to be involved in its anti-hyperalgesic effect.

  15. Shear-mediated platelet activation in the free flow: Perspectives on the emerging spectrum of cell mechanobiological mechanisms mediating cardiovascular implant thrombosis.

    PubMed

    Slepian, Marvin J; Sheriff, Jawaad; Hutchinson, Marcus; Tran, Phat; Bajaj, Naing; Garcia, Joe G N; Scott Saavedra, S; Bluestein, Danny

    2017-01-04

    Shear-mediated platelet activation (SMPA) is central in thrombosis of implantable cardiovascular therapeutic devices. Despite the morbidity and mortality associated with thrombosis of these devices, our understanding of mechanisms operative in SMPA, particularly in free flowing blood, remains limited. Herein we present and discuss a range of emerging mechanisms for consideration for "free flow" activation under supraphysiologic shear. Further definition and manipulation of these mechanisms will afford opportunities for novel pharmacologic and mechanical strategies to limit SMPA and enhance overall implant device safety.

  16. Autophagy in pulmonary macrophages mediates lung inflammatory injury via NLRP3 inflammasome activation during mechanical ventilation.

    PubMed

    Zhang, Yang; Liu, Gongjian; Dull, Randal O; Schwartz, David E; Hu, Guochang

    2014-07-15

    The inflammatory response is a primary mechanism in the pathogenesis of ventilator-induced lung injury. Autophagy is an essential, homeostatic process by which cells break down their own components. We explored the role of autophagy in the mechanisms of mechanical ventilation-induced lung inflammatory injury. Mice were subjected to low (7 ml/kg) or high (28 ml/kg) tidal volume ventilation for 2 h. Bone marrow-derived macrophages transfected with a scrambled or autophagy-related protein 5 small interfering RNA were administered to alveolar macrophage-depleted mice via a jugular venous cannula 30 min before the start of the ventilation protocol. In some experiments, mice were ventilated in the absence and presence of autophagy inhibitors 3-methyladenine (15 mg/kg ip) or trichostatin A (1 mg/kg ip). Mechanical ventilation with a high tidal volume caused rapid (within minutes) activation of autophagy in the lung. Conventional transmission electron microscopic examination of lung sections showed that mechanical ventilation-induced autophagy activation mainly occurred in lung macrophages. Autophagy activation in the lungs during mechanical ventilation was dramatically attenuated in alveolar macrophage-depleted mice. Selective silencing of autophagy-related protein 5 in lung macrophages abolished mechanical ventilation-induced nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) inflammasome activation and lung inflammatory injury. Pharmacological inhibition of autophagy also significantly attenuated the inflammatory responses caused by lung hyperinflation. The activation of autophagy in macrophages mediates early lung inflammation during mechanical ventilation via NLRP3 inflammasome signaling. Inhibition of autophagy activation in lung macrophages may therefore provide a novel and promising strategy for the prevention and treatment of ventilator-induced lung injury.

  17. DNA sequence-dependent mechanics and protein-assisted bending in repressor-mediated loop formation

    NASA Astrophysics Data System (ADS)

    Boedicker, James Q.; Garcia, Hernan G.; Johnson, Stephanie; Phillips, Rob

    2013-12-01

    As the chief informational molecule of life, DNA is subject to extensive physical manipulations. The energy required to deform double-helical DNA depends on sequence, and this mechanical code of DNA influences gene regulation, such as through nucleosome positioning. Here we examine the sequence-dependent flexibility of DNA in bacterial transcription factor-mediated looping, a context for which the role of sequence remains poorly understood. Using a suite of synthetic constructs repressed by the Lac repressor and two well-known sequences that show large flexibility differences in vitro, we make precise statistical mechanical predictions as to how DNA sequence influences loop formation and test these predictions using in vivo transcription and in vitro single-molecule assays. Surprisingly, sequence-dependent flexibility does not affect in vivo gene regulation. By theoretically and experimentally quantifying the relative contributions of sequence and the DNA-bending protein HU to DNA mechanical properties, we reveal that bending by HU dominates DNA mechanics and masks intrinsic sequence-dependent flexibility. Such a quantitative understanding of how mechanical regulatory information is encoded in the genome will be a key step towards a predictive understanding of gene regulation at single-base pair resolution.

  18. Social, institutional, and knowledge mechanisms mediate diverse ecosystem service benefits from coral reefs.

    PubMed

    Hicks, Christina C; Cinner, Joshua E

    2014-12-16

    Ecosystem services are supplied by nature but, by definition, are received by people. Ecosystem service assessments, intended to influence the decisions people make regarding their interactions with nature, need to understand how people benefit from different ecosystem services. A critical question is therefore, What determines the distribution of ecosystem service benefits between different sections of society? Here, we use an entitlements approach to examine how people perceive ecosystem service benefits across 28 coral reef fishing communities in four countries. In doing so, we quantitatively show that bundles of benefits are mediated by key access mechanisms (e.g., rights-based, economic, knowledge, social, and institutional). We find that specific access mechanisms influence which ecosystem services people prioritize. Social, institutional, and knowledge mechanisms are associated with the largest number and diversity of benefits. However, local context strongly determines whether specific access mechanisms enable or constrain benefits. Local ecological knowledge enabled people to prioritize a habitat benefit in Kenya, but constrained people from prioritizing the same benefit in Madagascar. Ecosystem service assessments, and their resultant policies, need to include the broad suite of access mechanisms that enable different people to benefit from a supply of ecosystem services.

  19. Mycobacteria counteract a TLR-mediated nitrosative defense mechanism in a zebrafish infection model.

    PubMed

    Elks, Philip M; van der Vaart, Michiel; van Hensbergen, Vincent; Schutz, Esther; Redd, Michael J; Murayama, Emi; Spaink, Herman P; Meijer, Annemarie H

    2014-01-01

    Pulmonary tuberculosis (TB), caused by the intracellular bacterial pathogen Mycobacterium tuberculosis (Mtb), is a major world health problem. The production of reactive nitrogen species (RNS) is a potent cytostatic and cytotoxic defense mechanism against intracellular pathogens. Nevertheless, the protective role of RNS during Mtb infection remains controversial. Here we use an anti-nitrotyrosine antibody as a readout to study nitration output by the zebrafish host during early mycobacterial pathogenesis. We found that recognition of Mycobacterium marinum, a close relative of Mtb, was sufficient to induce a nitrosative defense mechanism in a manner dependent on MyD88, the central adaptor protein in Toll like receptor (TLR) mediated pathogen recognition. However, this host response was attenuated by mycobacteria via a virulence mechanism independent of the well-characterized RD1 virulence locus. Our results indicate a mechanism of pathogenic mycobacteria to circumvent host defense in vivo. Shifting the balance of host-pathogen interactions in favor of the host by targeting this virulence mechanism may help to alleviate the problem of infection with Mtb strains that are resistant to multiple drug treatments.

  20. Social, institutional, and knowledge mechanisms mediate diverse ecosystem service benefits from coral reefs

    PubMed Central

    Cinner, Joshua E.

    2014-01-01

    Ecosystem services are supplied by nature but, by definition, are received by people. Ecosystem service assessments, intended to influence the decisions people make regarding their interactions with nature, need to understand how people benefit from different ecosystem services. A critical question is therefore, What determines the distribution of ecosystem service benefits between different sections of society? Here, we use an entitlements approach to examine how people perceive ecosystem service benefits across 28 coral reef fishing communities in four countries. In doing so, we quantitatively show that bundles of benefits are mediated by key access mechanisms (e.g., rights-based, economic, knowledge, social, and institutional). We find that specific access mechanisms influence which ecosystem services people prioritize. Social, institutional, and knowledge mechanisms are associated with the largest number and diversity of benefits. However, local context strongly determines whether specific access mechanisms enable or constrain benefits. Local ecological knowledge enabled people to prioritize a habitat benefit in Kenya, but constrained people from prioritizing the same benefit in Madagascar. Ecosystem service assessments, and their resultant policies, need to include the broad suite of access mechanisms that enable different people to benefit from a supply of ecosystem services. PMID:25453100

  1. URB597, an inhibitor of fatty acid amide hydrolase, reduces hyperalgesia in diabetic rats.

    PubMed

    Hasanein, Parisa; Parviz, Mohsen; Keshavarz, Mansoor; Roohbakhsh, Ali

    2009-06-01

    Diabetic rats display increased pain responses after injection of formalin into the paw or thermal stimulation of the tail, suggesting the presence of hyperalgesia. In this study, we investigated the efficacy of URB597 (0.1, 0.3, and 0.5 mg/kg, i.p.), an inhibitor of endocannabinoids metabolism, on 2 models of experimental hyperalgesia in streptozotocin (STZ)-induced diabetic rats. Animals were divided into control, URB597-treated control (0.1, 0.3, and 0.5 mg/kg), diabetic, and URB597-treated diabetic (0.1, 0.3, and 0.5 mg/kg) groups. Formalin and tail-flick tests were performed 4 and 8 weeks after the onset of hyperglycemia, respectively. Diabetes caused significant hyperalgesia during these tests. URB597 (0.3 and 0.5 mg/kg) reversed chemical and thermal hyperalgesia in diabetic rats. Administration of URB597 at a dose of 0.1 mg/kg did not alter pain-related behaviors in control and diabetic groups compared with those of the respective control groups. URB597 treatment did not affect body weight or plasma glucose level of treated animals compared with nontreated animals. This study shows that increasing endocannabinoid neurotransmission with URB597 displays efficacy in chemical and thermal models of diabetic hyperalgesia. It also suggests that URB597 is a promising tool for treatment of painful diabetic neuropathy.

  2. Neuropeptide-mediated excitability: a key triggering mechanism for seizure generation in the developing brain

    PubMed Central

    Baram, Tallie Z.; Hatalski, Carolyn G.

    2012-01-01

    Most human seizures occur early in life, consistent with established excitability-promoting features of the developing brain. Surprisingly, the majority of developmental seizures are not spontaneous but are provoked by injurious or stressful stimuli. What mechanisms mediate ‘triggering’ of seizures and limit such reactive seizures to early postnatal life? Recent evidence implicates the excitatory neuropeptide, corticotropin-releasing hormone (CRH). Stress activates expression of the CRH gene in several limbic regions, and CRH-expressing neurons are strategically localized in the immature rat hippocampus, in which this neuropeptide increases the excitability of pyramidal cells in vitro. Indeed, in vivo, activation of CRH receptors – maximally expressed in hippocampus and amygdala during the developmental period which is characterized by peak susceptibility to ‘provoked’ convulsions – induces severe, age-dependent seizures. Thus, converging data indicate that activation of expression of CRH constitutes an important mechanism for generating developmentally regulated, triggered seizures, with considerable clinical relevance. PMID:9829688

  3. A resonance mechanism of efficient energy transfer mediated by Fenna-Matthews-Olson complex

    NASA Astrophysics Data System (ADS)

    Alicki, Robert; Miklaszewski, Wiesław

    2012-04-01

    The Wigner-Weisskopf-type model developed by Alicki and Giraldi [J. Phys. B 44, 154020 (2011)], 10.1088/0953-4075/44/15/154020 is applied to the biological process of energy transfer from a large peripheral light harvesting antenna to the reaction center. This process is mediated by the Fenna-Matthews-Olson (FMO) photosynthetic complex with a remarkably high efficiency. The proposed model provides a simple resonance mechanism of this phenomenon employing exciton coherent motion and is described by analytical formulas. A coupling to the vibrational environment is a necessary component of this mechanism as well as a fine-tuning of the FMO complex Hamiltonian. The role of the relatively strong coupling to the energy sink in achieving the resonance condition and the absence of heating of the vibrational environment are emphasized.

  4. An electrostatic mechanism for Ca2+-mediated regulation of gap junction channels

    PubMed Central

    Bennett, Brad C.; Purdy, Michael D.; Baker, Kent A.; Acharya, Chayan; McIntire, William E.; Stevens, Raymond C.; Zhang, Qinghai; Harris, Andrew L.; Abagyan, Ruben; Yeager, Mark

    2016-01-01

    Gap junction channels mediate intercellular signalling that is crucial in tissue development, homeostasis and pathologic states such as cardiac arrhythmias, cancer and trauma. To explore the mechanism by which Ca2+ blocks intercellular communication during tissue injury, we determined the X-ray crystal structures of the human Cx26 gap junction channel with and without bound Ca2+. The two structures were nearly identical, ruling out both a large-scale structural change and a local steric constriction of the pore. Ca2+ coordination sites reside at the interfaces between adjacent subunits, near the entrance to the extracellular gap, where local, side chain conformational rearrangements enable Ca2+chelation. Computational analysis revealed that Ca2+-binding generates a positive electrostatic barrier that substantially inhibits permeation of cations such as K+ into the pore. Our results provide structural evidence for a unique mechanism of channel regulation: ionic conduction block via an electrostatic barrier rather than steric occlusion of the channel pore. PMID:26753910

  5. Oxidative Stress-Mediated Skeletal Muscle Degeneration: Molecules, Mechanisms, and Therapies

    PubMed Central

    Choi, Min Hee; Ow, Jin Rong; Yang, Nai-Di

    2016-01-01

    Oxidative stress is a loss of balance between the production of reactive oxygen species during cellular metabolism and the mechanisms that clear these species to maintain cellular redox homeostasis. Increased oxidative stress has been associated with muscular dystrophy, and many studies have proposed mechanisms that bridge these two pathological conditions at the molecular level. In this review, the evidence indicating a causal role of oxidative stress in the pathogenesis of various muscular dystrophies is revisited. In particular, the mediation of cellular redox status in dystrophic muscle by NF-κB pathway, autophagy, telomere shortening, and epigenetic regulation are discussed. Lastly, the current stance of targeting these pathways using antioxidant therapies in preclinical and clinical trials is examined. PMID:26798425

  6. Dynamic interactions mediated by nonredundant signaling mechanisms couple circadian clock neurons.

    PubMed

    Evans, Jennifer A; Leise, Tanya L; Castanon-Cervantes, Oscar; Davidson, Alec J

    2013-11-20

    Interactions among suprachiasmatic nucleus (SCN) neurons are required for robust circadian rhythms entrained to local time. To investigate these signaling mechanisms, we developed a functional coupling assay that uniquely captures the dynamic process by which SCN neurons interact. As a population, SCN neurons typically display synchronized rhythms with similar peak times, but will peak 6-12 hr apart after in vivo exposure to long days. Once they are removed from these conditions, SCN neurons resynchronize through a phase-dependent coupling process mediated by both vasoactive intestinal polypeptide (VIP) and GABAA signaling. Notably, GABAA signaling contributes to coupling when the SCN network is in an antiphase configuration, but opposes synchrony under steady-state conditions. Further, VIP acts together with GABAA signaling to couple the network in an antiphase configuration, but promotes synchrony under steady-state conditions by counteracting the actions of GABAA signaling. Thus, SCN neurons interact through nonredundant coupling mechanisms influenced by the state of the network.

  7. Vinpocetine Reduces Carrageenan-Induced Inflammatory Hyperalgesia in Mice by Inhibiting Oxidative Stress, Cytokine Production and NF-κB Activation in the Paw and Spinal Cord

    PubMed Central

    Ruiz-Miyazawa, Kenji W.; Zarpelon, Ana C.; Pinho-Ribeiro, Felipe A.; Pavão-de-Souza, Gabriela F.; Casagrande, Rubia; Verri, Waldiceu A.

    2015-01-01

    Vinpocetine is a safe nootropic agent used for neurological and cerebrovascular diseases. The anti-inflammatory activity of vinpocetine has been shown in cell based assays and animal models, leading to suggestions as to its utility in analgesia. However, the mechanisms regarding its efficacy in inflammatory pain treatment are still not completely understood. Herein, the analgesic effect of vinpocetine and its anti-inflammatory and antioxidant mechanisms were addressed in murine inflammatory pain models. Firstly, we investigated the protective effects of vinpocetine in overt pain-like behavior induced by acetic acid, phenyl-p-benzoquinone (PBQ) and formalin. The intraplantar injection of carrageenan was then used to induce inflammatory hyperalgesia. Mechanical and thermal hyperalgesia were evaluated using the electronic von Frey and the hot plate tests, respectively, with neutrophil recruitment to the paw assessed by a myeloperoxidase activity assay. A number of factors were assessed, both peripherally and in the spinal cord, including: antioxidant capacity, reduced glutathione (GSH) levels, superoxide anion, tumor necrosis factor alpha (TNF-α) and interleukin 1 beta (IL-1β) levels, as well as nuclear factor kappa B (NF-κB) activation. Vinpocetine inhibited the overt pain-like behavior induced by acetic acid, PBQ and formalin (at both phases), as well as the carrageenan-induced mechanical and thermal hyperalgesia and associated neutrophil recruitment. Both peripherally and in the spinal cord, vinpocetine also inhibited: antioxidant capacity and GSH depletion; increased superoxide anion; IL-1β and TNF-α levels; and NF-κB activation. As such, vinpocetine significantly reduces inflammatory pain by targeting oxidative stress, cytokine production and NF-κB activation at both peripheral and spinal cord levels. PMID:25822523

  8. Ultrasound Microbubble Treatment Enhances Clathrin-Mediated Endocytosis and Fluid-Phase Uptake through Distinct Mechanisms

    PubMed Central

    Fekri, Farnaz; Delos Santos, Ralph Christian; Karshafian, Raffi

    2016-01-01

    Drug delivery to tumors is limited by several factors, including drug permeability of the target cell plasma membrane. Ultrasound in combination with microbubbles (USMB) is a promising strategy to overcome these limitations. USMB treatment elicits enhanced cellular uptake of materials such as drugs, in part as a result of sheer stress and formation of transient membrane pores. Pores formed upon USMB treatment are rapidly resealed, suggesting that other processes such as enhanced endocytosis may contribute to the enhanced material uptake by cells upon USMB treatment. How USMB regulates endocytic processes remains incompletely understood. Cells constitutively utilize several distinct mechanisms of endocytosis, including clathrin-mediated endocytosis (CME) for the internalization of receptor-bound macromolecules such as Transferrin Receptor (TfR), and distinct mechanism(s) that mediate the majority of fluid-phase endocytosis. Tracking the abundance of TfR on the cell surface and the internalization of its ligand transferrin revealed that USMB acutely enhances the rate of CME. Total internal reflection fluorescence microscopy experiments revealed that USMB treatment altered the assembly of clathrin-coated pits, the basic structural units of CME. In addition, the rate of fluid-phase endocytosis was enhanced, but with delayed onset upon USMB treatment relative to the enhancement of CME, suggesting that the two processes are distinctly regulated by USMB. Indeed, vacuolin-1 or desipramine treatment prevented the enhancement of CME but not of fluid phase endocytosis upon USMB, suggesting that lysosome exocytosis and acid sphingomyelinase, respectively, are required for the regulation of CME but not fluid phase endocytosis upon USMB treatment. These results indicate that USMB enhances both CME and fluid phase endocytosis through distinct signaling mechanisms, and suggest that strategies for potentiating the enhancement of endocytosis upon USMB treatment may improve targeted

  9. Central mechanisms mediating the hypophagic effects of oleoylethanolamide and N-acylphosphatidylethanolamines: different lipid signals?

    PubMed Central

    Romano, Adele; Tempesta, Bianca; Provensi, Gustavo; Passani, Maria B.; Gaetani, Silvana

    2015-01-01

    The spread of “obesity epidemic” and the poor efficacy of many anti-obesity therapies in the long-term highlight the need to develop novel efficacious therapy. This necessity stimulates a large research effort to find novel mechanisms controlling feeding and energy balance. Among these mechanisms a great deal of attention has been attracted by a family of phospholipid-derived signaling molecules that play an important role in the regulation of food-intake. They include N-acylethanolamines (NAEs) and N-acylphosphatidylethanolamines (NAPEs). NAPEs have been considered for a long time simply as phospholipid precursors of the lipid mediator NAEs, but increasing body of evidence suggest a role in many physiological processes including the regulation of feeding behavior. Several observations demonstrated that among NAEs, oleoylethanolamide (OEA) acts as a satiety signal, which is generated in the intestine, upon the ingestion of fat, and signals to the central nervous system. At this level different neuronal pathways, including oxytocinergic, noradrenergic, and histaminergic neurons, seem to mediate its hypophagic action. Similarly to NAEs, NAPE (with particular reference to the N16:0 species) levels were shown to be regulated by the fed state and this finding was initially interpreted as fluctuations of NAE precursors. However, the observation that exogenously administered NAPEs are able to inhibit food intake, not only in normal rats and mice but also in mice lacking the enzyme that converts NAPEs into NAEs, supported the hypothesis of a role of NAPE in the regulation of feeding behavior. Indirect observations suggest that the hypophagic action of NAPEs might involve central mechanisms, although the molecular target remains unknown. The present paper reviews the role that OEA and NAPEs play in the mechanisms that control food intake, further supporting this group of phospholipids as optimal candidate for the development of novel anti-obesity treatments. PMID

  10. Molecular Mechanisms Responsible for Neuron-Derived Conditioned Medium (NCM)-Mediated Protection of Ischemic Brain

    PubMed Central

    Lin, Chi-Hsin; Wang, Chen-Hsuan; Hsu, Shih-Lan; Liao, Li-Ya; Lin, Ting-An; Hsueh, Chi-Mei

    2016-01-01

    The protective value of neuron-derived conditioned medium (NCM) in cerebral ischemia and the underlying mechanism(s) responsible for NCM-mediated brain protection against cerebral ischemia were investigated in the study. NCM was first collected from the neuronal culture growing under the in vitro ischemic condition (glucose-, oxygen- and serum-deprivation or GOSD) for 2, 4 or 6 h. Through the focal cerebral ischemia (bilateral CCAO/unilateral MCAO) animal model, we discovered that ischemia/reperfusion (I/R)-induced brain infarction was significantly reduced by NCM, given directly into the cistern magna at the end of 90 min of CCAO/MCAO. Immunoblocking and chemical blocking strategies were applied in the in vitro ischemic studies to show that NCM supplement could protect microglia, astrocytes and neurons from GOSD-induced cell death, in a growth factor (TGFβ1, NT-3 and GDNF) and p-ERK dependent manner. Brain injection with TGFβ1, NT3, GDNF and ERK agonist (DADS) alone or in combination, therefore also significantly decreased the infarct volume of ischemic brain. Moreover, NCM could inhibit ROS but stimulate IL-1β release from GOSD-treated microglia and limit the infiltration of IL-β-positive microglia into the core area of ischemic brain, revealing the anti-oxidant and anti-inflammatory activities of NCM. In overall, NCM-mediated brain protection against cerebral ischemia has been demonstrated for the first time in S.D. rats, due to its anti-apoptotic, anti-oxidant and potentially anti-glutamate activities (NCM-induced IL-1β can inhibit the glutamate-mediated neurotoxicity) and restriction upon the infiltration of inflammatory microglia into the core area of ischemic brain. The therapeutic potentials of NCM, TGFβ1, GDNF, NT-3 and DADS in the control of cerebral ischemia in human therefore have been suggested and require further investigation. PMID:26745377

  11. Improvement of Transmembrane Transport Mechanism Study of Imperatorin on P-Glycoprotein-Mediated Drug Transport.

    PubMed

    Liao, Zheng-Gen; Tang, Tao; Guan, Xue-Jing; Dong, Wei; Zhang, Jing; Zhao, Guo-Wei; Yang, Ming; Liang, Xin-Li

    2016-11-24

    P-glycoprotein (P-gp) affects the transport of many drugs; including puerarin and vincristine. Our previous study demonstrated that imperatorin increased the intestinal absorption of puerarin and vincristine by inhibiting P-gp-mediated drug efflux. However; the underlying mechanism was not known. The present study investigated the mechanism by which imperatorin promotes P-gp-mediated drug transport. We used molecular docking to predict the binding force between imperatorin and P-gp and the effect of imperatorin on P-gp activity. P-gp efflux activity and P-gp ATPase activity were measured using a rhodamine 123 (Rh-123) accumulation assay and a Pgp-Glo™ assay; respectively. The fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH) was used to assess cellular membrane fluidity in MDCK-MDR1 cells. Western blotting was used to analyze the effect of imperatorin on P-gp expression; and P-gp mRNA levels were assessed by qRT-PCR. Molecular docking results demonstrated that the binding force between imperatorin and P-gp was much weaker than the force between P-gp and verapamil (a P-gp substrate). Imperatorin activated P-gp ATPase activity; which had a role in the inhibition of P-gp activity. Imperatorin promoted Rh-123 accumulation in MDCK-MDR1 cells and decreased cellular membrane fluidity. Western blotting demonstrated that imperatorin inhibited P-gp expression; and qRT-PCR revealed that imperatorin down-regulated P-gp (MDR1) gene expression. Imperatorin decreased P-gp-mediated drug efflux by inhibiting P-gp activity and the expression of P-gp mRNA and protein. Our results suggest that imperatorin could down-regulate P-gp expression to overcome multidrug resistance in tumors.

  12. Molecular Mechanisms Responsible for Neuron-Derived Conditioned Medium (NCM)-Mediated Protection of Ischemic Brain.

    PubMed

    Lin, Chi-Hsin; Wang, Chen-Hsuan; Hsu, Shih-Lan; Liao, Li-Ya; Lin, Ting-An; Hsueh, Chi-Mei

    2016-01-01

    The protective value of neuron-derived conditioned medium (NCM) in cerebral ischemia and the underlying mechanism(s) responsible for NCM-mediated brain protection against cerebral ischemia were investigated in the study. NCM was first collected from the neuronal culture growing under the in vitro ischemic condition (glucose-, oxygen- and serum-deprivation or GOSD) for 2, 4 or 6 h. Through the focal cerebral ischemia (bilateral CCAO/unilateral MCAO) animal model, we discovered that ischemia/reperfusion (I/R)-induced brain infarction was significantly reduced by NCM, given directly into the cistern magna at the end of 90 min of CCAO/MCAO. Immunoblocking and chemical blocking strategies were applied in the in vitro ischemic studies to show that NCM supplement could protect microglia, astrocytes and neurons from GOSD-induced cell death, in a growth factor (TGFβ1, NT-3 and GDNF) and p-ERK dependent manner. Brain injection with TGFβ1, NT3, GDNF and ERK agonist (DADS) alone or in combination, therefore also significantly decreased the infarct volume of ischemic brain. Moreover, NCM could inhibit ROS but stimulate IL-1β release from GOSD-treated microglia and limit the infiltration of IL-β-positive microglia into the core area of ischemic brain, revealing the anti-oxidant and anti-inflammatory activities of NCM. In overall, NCM-mediated brain protection against cerebral ischemia has been demonstrated for the first time in S.D. rats, due to its anti-apoptotic, anti-oxidant and potentially anti-glutamate activities (NCM-induced IL-1β can inhibit the glutamate-mediated neurotoxicity) and restriction upon the infiltration of inflammatory microglia into the core area of ischemic brain. The therapeutic potentials of NCM, TGFβ1, GDNF, NT-3 and DADS in the control of cerebral ischemia in human therefore have been suggested and require further investigation.

  13. Contribution of C5-mediated mechanisms to host defence against Echinococcus granulosus hydatid infection.

    PubMed

    Ferreira, A M; Breijo, M; Sim, R B; Nieto, A

    2000-09-01

    The aim of this work was to investigate the contribution of complement C5-mediated mechanisms, with an emphasis on inflammation, to host defences against Echinococcus granulosus hydatid disease. Thus, we compared the systemic and local inflammatory responses induced by the parasite, and the outcome of infection, between congenic C5-sufficient (B10.D2 n/SnJ) and C5-deficient (B10.D2 o/SnJ) mice challenged with protoscoleces. Indirect evidence of in-vivo complement activation during the establishment phase was obtained; infection induced serum amyloid P and eosinophil responses which were dependent on C5. Early recruitment of polymorphonuclear cells was not dependent on the presence of C5. The higher capacity of C5-sufficient mice to recruit eosinophils was also observed during the cystic phase of infection, and mice recruiting more eosinophils developed lower parasite masses. Analysis of the outcome of infection after 8 months showed that C5-sufficient mice were more resistant to infection than C5-deficient mice in terms of individuals with no cysts; this trend was not statistically significant. In addition, C5-deficient mice developed higher numbers of large (> 5 mm in diameter) cysts and higher cyst weights than C5-sufficient mice indicating that C5-mediated mechanisms are detrimental for parasite growth. Taken together, our results suggest that complement, through C5-mediated effectors, contributes to host defences by both restricting the establishment of infection and controlling the growth of established cysts. This contribution may, at least partially, be associated with the ability of C5a to promote eosinophil infiltration.

  14. Separating behavioral and physiological mechanisms in testosterone-mediated trade-offs.

    PubMed

    Mougeot, François; Redpath, Stephen M; Piertney, Stuart B; Hudson, Peter J

    2005-08-01

    Testosterone often mediates trade-offs between reproduction and other life-history traits, which are usually investigated using testosterone implants. However, this approach does not distinguish between the physiological and behavioral effects of testosterone. We studied a wild game bird, the red grouse Lagopus lagopus scoticus, and took a new approach to investigate mechanisms linking elevated testosterone to increased parasite intensity. We caught males in autumn, removed their parasites, implanted them with the antiandrogen flutamide in combination with an aromatase inhibitor (FA males) or with empty implants (control males), and challenged them with parasites. The FA treatment increased testosterone concentration and physiological stress, but without enhancing testosterone-dependent behaviors, because testosterone receptors were blocked. FA males ended up with more parasites than the control males the following autumn, an effect similar to that of a testosterone treatment reported elsewhere. However, and unlike the testosterone treatment, the FA treatment did not affect home range, pairing, or breeding success. The results supported a physiological mechanism (increased susceptibility) linking elevated testosterone and increased parasite intensity. The FA treatment provided a new way of investigating testosterone-mediated trade-offs whereby testosterone concentration was increased while the effects on behavior were blocked, resulting in physiological costs without phenotypic benefits.

  15. Direct and quinone-mediated palladium reduction by Geobacter sulfurreducens: mechanisms and modeling.

    PubMed

    Pat-Espadas, Aurora M; Razo-Flores, Elías; Rangel-Mendez, J Rene; Cervantes, Francisco J

    2014-01-01

    Palladium(II) reduction to Pd(0) nanoparticles by Geobacter sulfurreducens was explored under conditions of neutral pH, 30 °C and concentrations of 25, 50, and 100 mg of Pd(II)/L aiming to investigate the effect of solid species of palladium on their microbial reduction. The influence of anthraquinone-2,6-disulfonate was reported to enhance the palladium reaction rate in an average of 1.7-fold and its addition is determining to achieve the reduction of solid species of palladium. Based on the obtained results two mechanisms are proposed: (1) direct, which is fully described considering interactions of amide, sulfur, and phosphoryl groups associated to proteins from bacteria on palladium reduction reaction, and (2) quinone-mediated, which implies multiheme c-type cytochromes participation. Speciation analysis and kinetic results were considered and integrated into a model to fit the experimental data that explain both mechanisms. This work provides elements for a better understanding of direct and quinone-mediated palladium reduction by G. sulfurreducens, which could facilitate metal recovery with concomitant formation of valuable palladium nanoparticles in industrial processes.

  16. In vivo mechanical loading rapidly activates β-catenin signaling in osteocytes through a prostaglandin mediated mechanism.

    PubMed

    Lara-Castillo, N; Kim-Weroha, N A; Kamel, M A; Javaheri, B; Ellies, D L; Krumlauf, R E; Thiagarajan, G; Johnson, M L

    2015-07-01

    The response of the skeleton to loading appears to be mediated through the activation of the Wnt/β-catenin signaling pathway and osteocytes have long been postulated to be the primary mechanosensory cells in bone. To examine the kinetics of the mechanoresponse of bone and cell types involved in vivo, we performed forearm loading of 17-week-old female TOPGAL mice. β-catenin signaling was observed only in embedded osteocytes, not osteoblasts, at 1h post-loading, spreading to additional osteocytes and finally to cells on the bone surface by 24h. This early activation at 1h appeared to be independent of receptor (Lrp5/6) mediated activation as it occurred in the presence of the inhibitors sclerostin and/or Dkk1. The COX-2 inhibitor, Carprofen, blocked the activation of β-catenin signaling and decline in sclerostin positive osteocytes post-loading implying an important role for prostaglandin. In vitro, PI3K/Akt activation was shown to be required for β-catenin nuclear translocation downstream from prostaglandin in MLO-Y4 osteocyte-like cells supporting this mechanism. Downstream targets of β-catenin signaling, sclerostin and Dkk1, were also examined and found to be significantly downregulated in osteocytes in vivo at 24h post-loading. The pattern of initially activated osteocytes appeared random and in order to understand this heterogeneous expression, a novel finite element model of the strain field in the ulna was developed, which predicts highly variable local magnitudes of strain experienced by osteocytes. In summary, both in vivo and in vitro models show the rapid activation of β-catenin in response to load through the early release of prostaglandin and that strain fields in the bone are extremely heterogeneous resulting in heterogeneous activation of the β-catenin pathway in osteocytes in vivo.

  17. In Vivo Mechanical Loading Rapidly Activates β–catenin Signaling in Osteocytes through a Prostaglandin Mediated Mechanism

    PubMed Central

    Lara-Castillo, N; Kim-Weroha, NA; Kamel, MA; Javaheri, B; Ellies, DL; Krumlauf, RE; Thiagarajan, G; Johnson, ML

    2015-01-01

    The response of the skeleton to loading appears to be mediated through the activation of the Wnt/β-catenin signaling pathway and osteocytes have long been postulated to be the primary mechanosensory cells in bone. To examine the kinetics of the mechanoresponse of bone and cell types involved in the in vivo, we performed forearm loading of 17-week-old female TOPGAL mice. β-catenin signaling was observed only in embedded osteocytes, not osteoblasts, at 1 hour post loading, spreading to additional osteocytes and finally to cells on the bone surface by 24 hrs. This early activation at 1 hour appeared to be independent of receptor (Lrp5/6) mediated activation as it occurred in the presence of the inhibitors sclerostin and/or Dkk1. The COX-2 inhibitor, Carprofen, blocked the activation of β-catenin signaling and decline in sclerostin positive osteocytes post-loading implying an important role for prostaglandin. In vitro, PI3K/Akt activation was shown to be required for β-catenin nuclear translocation downstream from prostaglandin in MLO-Y4 osteocyte-like cells supporting this mechanism. Downstream targets of β-catenin signaling, sclerostin and Dkk1, were also examined and found to be significantly down regulated in osteocytes in vivo at 24 hours post-loading. The pattern of initially activated osteocytes appeared random and in order to understand this heterogeneous expression, a novel finite element model of the strain field in the ulna was developed, which predicts highly variable local magnitudes of strain experienced by osteocytes. In summary, both in vivo and in vitro models show the rapid activation of β-catenin in response to load through the early release of prostaglandin and that strain fields in the bone are extremely heterogeneous resulting in heterogeneous activation of the β-catenin pathway in osteocytes in vivo. PMID:25836764

  18. EP2 receptor antagonism reduces peripheral and central hyperalgesia in a preclinical mouse model of endometriosis.

    PubMed

    Greaves, Erin; Horne, Andrew W; Jerina, Helen; Mikolajczak, Marta; Hilferty, Lisa; Mitchell, Rory; Fleetwood-Walker, Sue M; Saunders, Philippa T K

    2017-03-10

    Endometriosis is an incurable gynecological disorder characterized by debilitating pain and the establishment of innervated endometriosis lesions outside the uterus. In a preclinical mouse model of endometriosis we demonstrated overexpression of the PGE2-signaling pathway (including COX-2, EP2, EP4) in endometriosis lesions, dorsal root ganglia (DRG), spinal cord, thalamus and forebrain. TRPV1, a PGE2-regulated channel in nociceptive neurons was also increased in the DRG. These findings support the concept that an amplification process occurs along the pain neuroaxis in endometriosis. We then tested TRPV1, EP2, and EP4 receptor antagonists: The EP2 antagonist was the most efficient analgesic, reducing primary hyperalgesia by 80% and secondary hyperalgesia by 40%. In this study we demonstrate reversible peripheral and central hyperalgesia in mice with induced endometriosis.

  19. EP2 receptor antagonism reduces peripheral and central hyperalgesia in a preclinical mouse model of endometriosis

    PubMed Central

    Greaves, Erin; Horne, Andrew W.; Jerina, Helen; Mikolajczak, Marta; Hilferty, Lisa; Mitchell, Rory; Fleetwood-Walker, Sue M.; Saunders, Philippa T. K.

    2017-01-01

    Endometriosis is an incurable gynecological disorder characterized by debilitating pain and the establishment of innervated endometriosis lesions outside the uterus. In a preclinical mouse model of endometriosis we demonstrated overexpression of the PGE2-signaling pathway (including COX-2, EP2, EP4) in endometriosis lesions, dorsal root ganglia (DRG), spinal cord, thalamus and forebrain. TRPV1, a PGE2-regulated channel in nociceptive neurons was also increased in the DRG. These findings support the concept that an amplification process occurs along the pain neuroaxis in endometriosis. We then tested TRPV1, EP2, and EP4 receptor antagonists: The EP2 antagonist was the most efficient analgesic, reducing primary hyperalgesia by 80% and secondary hyperalgesia by 40%. In this study we demonstrate reversible peripheral and central hyperalgesia in mice with induced endometriosis. PMID:28281561

  20. Signalling mechanisms mediating Zn2+-induced TRPM2 channel activation and cell death in microglial cells

    PubMed Central

    Mortadza, Sharifah Syed; Sim, Joan A.; Stacey, Martin; Jiang, Lin-Hua

    2017-01-01

    Excessive Zn2+ causes brain damage via promoting ROS generation. Here we investigated the role of ROS-sensitive TRPM2 channel in H2O2/Zn2+-induced Ca2+ signalling and cell death in microglial cells. H2O2/Zn2+ induced concentration-dependent increases in cytosolic Ca2+ concentration ([Ca2+]c), which was inhibited by PJ34, a PARP inhibitor, and abolished by TRPM2 knockout (TRPM2-KO). Pathological concentrations of H2O2/Zn2+ induced substantial cell death that was inhibited by PJ34 and DPQ, PARP inhibitors, 2-APB, a TRPM2 channel inhibitor, and prevented by TRPM2-KO. Further analysis indicate that Zn2+ induced ROS production, PARP-1 stimulation, increase in the [Ca2+]c and cell death, all of which were suppressed by chelerythrine, a protein kinase C inhibitor, DPI, a NADPH-dependent oxidase (NOX) inhibitor, GKT137831, a NOX1/4 inhibitor, and Phox-I2, a NOX2 inhibitor. Furthermore, Zn2+-induced PARP-1 stimulation, increase in the [Ca2+]c and cell death were inhibited by PF431396, a Ca2+-sensitive PYK2 inhibitor, and U0126, a MEK/ERK inhibitor. Taken together, our study shows PKC/NOX-mediated ROS generation and PARP-1 activation as an important mechanism in Zn2+-induced TRPM2 channel activation and, TRPM2-mediated increase in the [Ca2+]c to trigger the PYK2/MEK/ERK signalling pathway as a positive feedback mechanism that amplifies the TRPM2 channel activation. Activation of these TRPM2-depenent signalling mechanisms ultimately drives Zn2+-induced Ca2+ overloading and cell death. PMID:28322340

  1. Dysfunction of nodes of Ranvier: a mechanism for anti-ganglioside antibody-mediated neuropathies

    PubMed Central

    Susuki, Keiichiro; Yuki, Nobuhiro; Schafer, Dorothy P.; Hirata, Koichi; Zhang, Gang; Funakoshi, Kei; Rasband, Matthew N.

    2011-01-01

    Autoantibodies against gangliosides GM1 or GD1a are associated with acute motor axonal neuropathy (AMAN) and acute motor-sensory axonal neuropathy (AMSAN), whereas antibodies to GD1b ganglioside are detected in acute sensory ataxic neuropathy (ASAN). These neuropathies have been proposed to be closely related and comprise a continuous spectrum, although the underlying mechanisms, especially for sensory nerve involvement, are still unclear. Antibodies to GM1 and GD1a have been proposed to disrupt the nodes of Ranvier in motor nerves via complement pathway. We hypothesized that the disruption of nodes of Ranvier is a common mechanism whereby various anti-ganglioside antibodies found in these neuropathies lead to nervous system dysfunction. Here, we show that the IgG monoclonal anti-GD1a/GT1b antibody injected into rat sciatic nerves caused deposition of IgG and complement products on the nodal axolemma and disrupted clusters of nodal and paranodal molecules predominantly in motor nerves, and induced early reversible motor nerve conduction block. Injection of IgG monoclonal anti-GD1b antibody induced nodal disruption predominantly in sensory nerves. In an ASAN rabbit model associated with IgG anti-GD1b antibodies, complement-mediated nodal disruption was observed predominantly in sensory nerves. In an AMAN rabbit model associated with IgG anti-GM1 antibodies, complement attack of nodes was found primarily in motor nerves, but occasionally in sensory nerves as well. Periaxonal macrophages and axonal degeneration were observed in dorsal roots from ASAN rabbits and AMAN rabbits. Thus, nodal disruption may be a common mechanism in immune-mediated neuropathies associated with autoantibodies to gangliosides GM1, GD1a, or GD1b, providing an explanation for the continuous spectrum of AMAN, AMSAN, and ASAN. PMID:22178332

  2. Molecular mechanism underlying juvenile hormone-mediated repression of precocious larval–adult metamorphosis

    PubMed Central

    Kayukawa, Takumi; Jouraku, Akiya; Ito, Yuka; Shinoda, Tetsuro

    2017-01-01

    Juvenile hormone (JH) represses precocious metamorphosis of larval to pupal and adult transitions in holometabolous insects. The early JH-inducible gene Krüppel homolog 1 (Kr-h1) plays a key role in the repression of metamorphosis as a mediator of JH action. Previous studies demonstrated that Kr-h1 inhibits precocious larval–pupal transition in immature larva via direct transcriptional repression of the pupal specifier Broad-Complex (BR-C). JH was recently reported to repress the adult specifier gene Ecdysone-induced protein 93F (E93); however, its mechanism of action remains unclear. Here, we found that JH suppressed ecdysone-inducible E93 expression in the epidermis of the silkworm Bombyx mori and in a B. mori cell line. Reporter assays in the cell line revealed that the JH-dependent suppression was mediated by Kr-h1. Genome-wide ChIP-seq analysis identified a consensus Kr-h1 binding site (KBS, 14 bp) located in the E93 promoter region, and EMSA confirmed that Kr-h1 directly binds to the KBS. Moreover, we identified a C-terminal conserved domain in Kr-h1 essential for the transcriptional repression of E93. Based on these results, we propose a mechanism in which JH-inducible Kr-h1 directly binds to the KBS site upstream of the E93 locus to repress its transcription in a cell-autonomous manner, thereby preventing larva from bypassing the pupal stage and progressing to precocious adult development. These findings help to elucidate the molecular mechanisms regulating the metamorphic genetic network, including the functional significance of Kr-h1, BR-C, and E93 in holometabolous insect metamorphosis. PMID:28096379

  3. Evaluation of heat hyperalgesia and anxiety like-behaviors in a rat model of orofacial cancer.

    PubMed

    Gambeta, Eder; Kopruszinski, Caroline Machado; Dos Reis, Renata Cristiane; Zanoveli, Janaina Menezes; Chichorro, Juliana Geremias

    2016-04-21

    Pain and anxiety are commonly experienced by cancer patients and both significantly impair their quality of life. Some authors claim that there is a relationship between pain and anxiety, while others suggest that there is not a direct association. In any case, there is indeed a consensus that anxiety impairs the pain condition beyond be under diagnosed and undertreated in cancer pain patients. Herein we investigated if rats presenting heat hyperalgesia induced by orofacial cancer cell inoculation would display anxiety-like behaviors. In addition, we evaluated if pain blockade would result in alleviation of anxiety behaviors, as well as, if blockade of anxiety would result in pain relief. Orofacial cancer was induced in male Wistar rats by inoculation of Walker-256 cells into the right vibrissal pad. Heat facial hyperalgesia was assessed on day 6 after the inoculation, and on this time point rats were submitted to the elevated plus maze and the light-dark transition tests. The influence of lidocaine and midazolam on heat hyperalgesia and anxiety-like behaviors was assessed. The peak of facial heat hyperalgesia was detected 6 days after cancer cells inoculation, and at this time point, rats exhibited increased anxiety-like behaviors. Local treatment with lidocaine (2%/50μL) caused a marked reduction of heat hyperalgesia, but failed to affect the anxiety-like behaviors, while midazolam (0.5mg/kg, i.p.) treatment failed to change the heat threshold, but induced an anxiolytic-like effect. Altogether, our data demonstrated that rats with orofacial cancer present pain- and anxiety-like behaviors, but brief heat hyperalgesia relief does not affect the anxiety-like behaviors, and vice-versa, in our experimental conditions.

  4. Pressure Pain Sensitivity in Patients With Suspected Opioid-Induced Hyperalgesia

    PubMed Central

    Wasserman, Ronald A.; Hassett, Afton L.; Harte, Steven E.; Goesling, Jenna; Malinoff, Herbert L.; Berland, Daniel W.; Zollars, Jennifer; Moser, Stephanie E.; Brummett, Chad M.

    2015-01-01

    Background and Objectives This study was designed to test whether a brief quantitative sensory testing (QST) assessment could be used to detect hyperalgesia in patients with suspected opioid-induced hyperalgesia. Methods Twenty patients on long-term opioid therapy with suspected opioid-induced hyperalgesia were recruited along with and 20 healthy controls. Pressure pain threshold, Pain50, a measure of intermediate suprathreshold pressure pain sensitivity, and tolerance levels, were evaluated. As a secondary outcome, changes in pressure pain sensitivity following intravenous administration of placebo (saline) and fentanyl (1.5 μg/kg) were assessed. Results There were no significant differences in pain measures between healthy controls and patients. However, there was an association between higher doses of opioids and having a lower pain tolerance (r= -0.46, P=0.041) and lower Pain50 (r=-0.46, P = 0.044), which was consistent with the hypothesis. Patients on >100 mg oral morphine equivalents (OME) displayed decreased pressure pain tolerance compared to patients taking <100 mg OME (P = 0.042). In addition, male patients showed a hyperalgesic response to fentanyl administration, which was significant for the Pain50 measure (P=0.002). Conclusions Whereas there were no differences between patients suspected of having opioid-induced hyperalgesia and the healthy controls, the finding that higher doses of opioids were associated with more sensitivity suggests that dose might be an important factor in the development of hyperalgesia. In addition, male patients demonstrated a hyperalgesic response after a bolus of fentanyl. Future studies are needed to develop better diagnostics for detecting hyperalgesia in the clinical setting. PMID:26469365

  5. Divergence between organometallic and single-electron-transfer mechanisms in copper(II)-mediated aerobic C-H oxidation.

    PubMed

    Suess, Alison M; Ertem, Mehmed Z; Cramer, Christopher J; Stahl, Shannon S

    2013-07-03

    Copper(II)-mediated C-H oxidation is the subject of extensive interest in synthetic chemistry, but the mechanisms of many of these reactions are poorly understood. Here, we observe different products from Cu(II)-mediated oxidation of N-(8-quinolinyl)benzamide, depending on the reaction conditions. Under basic conditions, the benzamide group undergoes directed C-H methoxylation or chlorination. Under acidic conditions, the quinoline group undergoes nondirected chlorination. Experimental and computational mechanistic studies implicate an organometallic C-H activation/functionalization mechanism under the former conditions and a single-electron-transfer mechanism under the latter conditions. This rare observation of divergent, condition-dependent mechanisms for oxidation of a single substrate provides a valuable foundation for understanding Cu(II)-mediated C-H oxidation reactions.

  6. Opioid-induced myoclonus and hyperalgesia following a short course of low-dose oral morphine

    PubMed Central

    Woodward, Owen Bleddyn; Naraen, Sangeeta; Naraen, Akriti

    2016-01-01

    A 76-year-old man was admitted to hospital with a right-sided fractured neck of femur requiring repair via a cemented hemiarthroplasty. Intraoperatively he received 10 mg of intravenous morphine. Post-operatively he received a short course of low-dose oral opioids and subsequently developed myoclonic jerks and hyperalgesia. The opioids were discontinued and both adverse effects resolved. This case report discusses the concurrent development of myoclonus and hyperalgesia following a low dose of opioids and explores possible management options. PMID:28386402

  7. A new mechanism for spatial pattern formation via lateral and protrusion-mediated lateral signalling

    PubMed Central

    Hunter, Ginger L.; Baum, Buzz

    2016-01-01

    Tissue organization and patterning are critical during development when genetically identical cells take on different fates. Lateral signalling plays an important role in this process by helping to generate self-organized spatial patterns in an otherwise uniform collection of cells. Recent data suggest that lateral signalling can be mediated both by junctional contacts between neighbouring cells and via cellular protrusions that allow non-neighbouring cells to interact with one another at a distance. However, it remains unclear precisely how signalling mediated by these distinct types of cell–cell contact can physically contribute to the generation of complex patterns without the assistance of diffusible morphogens or pre-patterns. To explore this question, in this work we develop a model of lateral signalling based on a single receptor/ligand pair as exemplified by Notch and Delta. We show that allowing the signalling kinetics to differ at junctional versus protrusion-mediated contacts, an assumption inspired by recent data which show that the cleavage of Notch in several systems requires both Delta binding and the application of mechanical force, permits individual cells to act to promote both lateral activation and lateral inhibition. Strikingly, under this model, in which Delta can sequester Notch, a variety of patterns resembling those typical of reaction–diffusion systems is observed, together with more unusual patterns that arise when we consider changes in signalling kinetics, and in the length and distribution of protrusions. Importantly, these patterns are self-organizing—so that local interactions drive tissue-scale patterning. Together, these data show that protrusions can, in principle, generate different types of patterns in addition to contributing to long-range signalling and to pattern refinement. PMID:27807273

  8. Comparison of molecular mechanisms mediating cell contact phenomena in model developmental systems: an exploration of universality.

    PubMed

    Bowers-Morrow, Vivienne M; Ali, Sinan O; Williams, Keith L

    2004-08-01

    Are there universal molecular mechanisms associated with cell contact phenomena during metazoan ontogenesis? Comparison of adhesion systems in disparate model systems indicates the existence of unifying principles. Requirements for multicellularity are (a) the construction of three-dimensional structures involving a crucial balance between adhesiveness and motility; and (b) the establishment of integration at molecular, cellular, tissue, and organismal levels of organization. Mechanisms for (i) cell-cell and cell-substrate adhesion, (ii) cell movement, (iii) cell-cell communication, (iv) cellular responses, (v) regulation of these processes, and (vi) their integration with patterning, growth, and other developmental processes are all crucial to metazoan development, and must have been present for the emergence and radiation of Metazoa. The principal unifying themes of this review are the dynamics and regulation of cell contact phenomena. Our knowledge of the dynamic molecular mechanisms underlying cell contact phenomena remains fragmentary. Here we examine the molecular bases of cell contact phenomena using extant model developmental systems (representing a wide range of phyla) including the simplest i.e. sponges, and the eukaryotic protist Dictyostelium discoideum, the more complex Drosophila melanogaster, and vertebrate systems. We discuss cell contact phenomena in a broad developmental context. The molecular language of cell contact phenomena is complex; it involves a plethora of structurally and functionally diverse molecules, and diverse modes of intermolecular interactions mediated by protein and/or carbohydrate moieties. Reasons for this are presumably the necessity for a high degree of specificity of intermolecular interactions, the requirement for a multitude of different signals, and the apparent requirement for an increasingly large repertoire of cell contact molecules in more complex developmental systems, such as the developing vertebrate nervous

  9. Potential mechanisms by which polyphenol-rich grapes prevent obesity-mediated inflammation and metabolic diseases.

    PubMed

    Chuang, Chia-Chi; McIntosh, Michael K

    2011-08-21

    Obesity and metabolic disease-related health problems (e.g., type 2 diabetes, atherosclerosis, and hypertension) are the most prevalent nutrition-related issues in the United States. An emerging feature of obesity and type 2 diabetes is their linkage with chronic inflammation that begins in white adipose tissue and eventually becomes systemic. One potential strategy to reduce inflammation and insulin resistance is consumption of polyphenol-rich foods like grapes or their by-products, which have anti-inflammatory properties. Polyphenols commonly found in grape products have been reported to reduce inflammation by (a) acting as an antioxidant or increasing antioxidant gene or protein expression, (b) attenuating endoplasmic reticulum stress signaling, (c) blocking proinflammatory cytokines or endotoxin-mediated kinases and transcription factors involved in metabolic disease, (d) suppressing inflammatory- or inducing metabolic-gene expression via increasing histone deacetylase activity, or (e) activating transcription factors that antagonize chronic inflammation. Thus, polyphenol-rich grape products may reduce obesity-mediated chronic inflammation by multiple mechanisms, thereby preventing metabolic diseases.

  10. Neurobiology of microglial action in CNS injuries: receptor-mediated signaling mechanisms and functional roles

    PubMed Central

    Hu, Xiaoming; Liou, Anthony K.F.; Leak, Rehana K.; Xu, Mingyue; An, Chengrui; Suenaga, Jun; Shi, Yejie; Gao, Yanqin; Zheng, Ping; Chen, Jun

    2014-01-01

    Microglia are the first line of immune defense against central nervous system (CNS) injuries and disorders. These highly plastic cells play dualistic roles in neuronal injury and recovery and are known for their ability to assume diverse phenotypes. A broad range of surface receptors are expressed on microglia and mediate microglial ‘On’ or ‘Off’ responses to signals from other host cells as well as invading microorganisms. The integrated actions of these receptors result in tightly regulated biological functions, including cell mobility, phagocytosis, the induction of acquired immunity, and trophic factor/inflammatory mediator release. Over the last few years, significant advances have been made towards deciphering the signaling mechanisms related to these receptors and their specific cellular functions. In this review, we describe the current state of knowledge of the surface receptors involved in microglial activation, with an emphasis on their engagement of distinct functional programs and their roles in CNS injuries. It will become evident from this review that microglial homeostasis is carefully maintained by multiple counterbalanced strategies, including, but not limited to, ‘On’ and ‘Off’ receptor signaling. Specific regulation of theses microglial receptors may be a promising therapeutic strategy against CNS injuries. PMID:24923657

  11. Reactive oxygen species-mediated cardiac-reperfusion injury: Mechanisms and therapies.

    PubMed

    Bagheri, Fereshte; Khori, Vahid; Alizadeh, Ali Mohammad; Khalighfard, Solmaz; Khodayari, Saeed; Khodayari, Hamid

    2016-11-15

    Reperfusion injury is an inherent response to the restoration of blood flow after ischemia. It is a complex process involving numerous mechanisms occurring in the intracellular and extracellular environments, and it is mediated in part by reactive oxygen species (ROS). The imbalance between the cellular formation of free radicals and cells' capacity to defend against them can cause cardiac tissue injuries. In this context, ROS play an essential role in both the organ injury and repair processes. After reperfusion, infiltration into the myocardium of inflammatory leucocytes, such as macrophages and neutrophils, causes further ROS production beyond the initiation of the inflammatory cascade. In this case, ROS overproduction is crucial in cardiac injury, and it can increase the complications related to cardiac reperfusion. In myocardial tissue, ROS can be produced from several sources, such as xanthine oxidase, cytochrome oxidase, cyclooxygenase, mediated unsaturated fatty acid oxidation, oxidation of catecholamines, mitochondrial oxidation, activation of leukocyte nicotinamide adenine dinucleotide phosphate oxidase, iron release, and reduction-oxidation reaction cycling; all of these sources reduce molecular oxygen in the reperfused myocardium. This review discusses about the molecular and therapeutic aspects of cardiac-reperfusion injuries generated by ROS. Experimental and clinical evidence with respect to the use of ischemic preconditioning, Ca(2+), nitric oxide, and conventional antioxidants in cardiac-reperfusion injury are summarized, and causal therapy approaches with various antioxidants are discussed.

  12. IFNγ Regulates Activated Vδ2+ T Cells through a Feedback Mechanism Mediated by Mesenchymal Stem Cells

    PubMed Central

    Fechter, Karoline; Dorronsoro, Akaitz; Jakobsson, Emma; Ferrin, Izaskun; Lang, Valérie; Sepulveda, Pilar; Pennington, Daniel J.; Trigueros, César

    2017-01-01

    γδ T cells play a role in a wide range of diseases such as autoimmunity and cancer. The majority of circulating human γδ T lymphocytes express a Vγ9Vδ2+ (Vδ2+) T cell receptor (TCR) and following activation release pro-inflammatory cytokines. In this study, we show that IFNγ, produced by Vδ2+ cells, activates mesenchymal stem cell (MSC)-mediated immunosupression, which in turn exerts a negative feedback mechanism on γδ T cell function ranging from cytokine production to proliferation. Importantly, this modulatory effect is limited to a short period of time (<24 hours) post-T cell activation, after which MSCs can no longer exert their immunoregulatory capacity. Using genetically modified MSCs with the IFNγ receptor 1 constitutively silenced, we demonstrate that IFNγ is essential to this process. Activated γδ T cells induce expression of several factors by MSCs that participate in the depletion of amino acids. In particular, we show that indolamine 2,3-dioxygenase (IDO), an enzyme involved in L-tryptophan degradation, is responsible for MSC-mediated immunosuppression of Vδ2+ T cells. Thus, our data demonstrate that γδ T cell responses can be immuno-modulated by different signals derived from MSC. PMID:28076364

  13. Role of carbohydrate-mediated adherence in cytopathogenic mechanisms of Acanthamoeba.

    PubMed

    Cao, Z; Jefferson, D M; Panjwani, N

    1998-06-19

    Acanthamoeba keratitis is a vision-threatening corneal infection. The mannose-binding protein of Acanthamoeba is thought to mediate adhesion of parasites to host cells. We characterized the amoeba lectin with respect to its carbohydrate binding properties and the role in amoeba-induced cytopathic effect (CPE). Sugar inhibition assays revealed that the amoeba lectin has the highest affinity for alpha-Man and Man(alpha1-3)Man units. In vitro cytopathic assays indicated that mannose-based saccharides which inhibit amoeba adhesion to corneal epithelial cells were also potent inhibitors of amoeba-induced CPE. Another major finding was that N-acetyl-D-glucosamine (GlcNAc) which does not inhibit adhesion of amoeba to host cells is also an inhibitor of amoeba-induced CPE. The Acanthamoebae are thought to produce CPE by secreting cytotoxic proteinases. By zymography, one metalloproteinase and three serine proteinases were detected in the conditioned media obtained after incubating amoebae with the host cells. The addition of free alpha-Man and GlcNAc to the co-culture media inhibited the secretion of the metalloproteinase and serine proteinases, respectively. In summary, we have shown that the lectin-mediated adhesion of the Acanthamoeba to host cells is a prerequisite for the amoeba-induced cytolysis of target cells and have implicated a contact-dependent metalloproteinase in the cytopathogenic mechanisms of Acanthamoeba.

  14. Calpain-Mediated Tau Cleavage: A Mechanism Leading to Neurodegeneration Shared by Multiple Tauopathies

    PubMed Central

    Ferreira, Adriana; Bigio, Eileen H

    2011-01-01

    Tau dysfunction has been associated with a host of neurodegenerative diseases called tauopathies. These diseases share, as a common pathological hallmark, the presence of intracellular aggregates of hyperphosphorylated tau in affected brain areas. Aside from tau hyperphosphorylation, little is known about the role of other posttranslational modifications in tauopathies. Recently, we obtained data suggesting that calpain-mediated tau cleavage leading to the generation of a neurotoxic tau fragment might play an important role in Alzheimer’s disease. In the current study, we assessed the presence of this tau fragment in several tauopathies. Our results show high levels of the 17-kDa tau fragment and enhanced calpain activity in the temporal cortex of AD patients and in brain samples obtained from patients with other tauopathies. In addition, our data suggest that this fragment could partially inhibit tau aggregation. Conversely, tau aggregation might prevent calpain-mediated cleavage, establishing a feedback circuit that might lead to the accumulation of this toxic tau fragment. Collectively, these data suggest that the mechanism underlying the generation of the 17-kDa neurotoxic tau fragment might be part of a conserved pathologic process shared by multiple tauopathies. PMID:21442128

  15. Synchronization of evoked secretion of quanta of mediator as a mechanism facilitating the action of sympathomimetics.

    PubMed

    Bukharaeva, E A; Kim, K K; Nikol'skii, E E; Vyskochil, F

    2000-01-01

    Experiments on frog neuromuscular junction preparations with extracellular recording of nerve terminal action potentials and single-quantum end-plate currents (EPC) were used to assess the time course of evoked quantum secretion of mediator by analyzing histograms of the distribution of true synaptic delays. These studies showed that noradrenaline, isoproterenol, and dobutamine change the kinetics of secretion of quanta, leading to synchronization of the process of mediator release; substances blocking beta-adrenoceptors (atenolol, propranolol) blocked this effect. Clonidine and phenylephrine, which activate alpha-receptors, had no effect on the kinetics of secretion, while the alpha-blocker phentolamine had no effect on the synchronizing action of noradrenaline. Reconstruction of multiquantum EPC from changes in the level of synchronization in the release of individual quanta, showed that EPC amplitude increased in response to noradrenaline by 17%, and that this was due only to alterations in the time course of secretion. These data led to the conclusion that there is a special presynaptic mechanism which facilitates the action of sympathomimetics, acting via beta-adrenoceptors.

  16. Mechanisms of Nanoparticle Mediated siRNA Transfection by Melittin-Derived Peptides

    PubMed Central

    Hou, Kirk K.; Pan, Hua; Ratner, Lee; Schlesinger, Paul H.; Wickline, Samuel A.

    2014-01-01

    Traditional peptide-mediated siRNA transfection via peptide transduction domains exhibits limited cytoplasmic delivery of siRNA due to endosomal entrapment. This work overcomes these limitations with the use of membrane-destabilizing peptides derived from melittin for the knockdown of NFkB signaling in a model of adult T-Cell leukemia/lymphoma. While the mechanism of siRNA delivery into the cytoplasmic compartment by peptide transduction domains has not been well studied, our analysis of melittin derivatives indicates that concurrent nanocomplex disassembly and peptide-mediated endosomolysis are crucial to siRNA transfection. Importantly, in the case of the most active derivative, p5RHH, this process is initiated by acidic pH, indicating that endosomal acidification after macropinocytosis can trigger siRNA release into the cytoplasm. These data provide general principles regarding nanocomplex response to endocytosis which may guide the development of peptide/siRNA nanocomplex-based transfection. PMID:24053333

  17. From Nanowires to Biofilms: An Exploration of Novel Mechanisms of Uranium Transformation Mediated by Geobacter Bacteria

    SciTech Connect

    REGUERA, GEMMA

    2014-01-16

    One promising strategy for the in situ bioremediation of radioactive groundwater contaminants that has been identified by the SBR Program is to stimulate the activity of dissimilatory metal-reducing microorganisms to reductively precipitate uranium and other soluble toxic metals. The reduction of U(VI) and other soluble contaminants by Geobacteraceae is directly dependent on the reduction of Fe(III) oxides, their natural electron acceptor, a process that requires the expression of Geobacter’s conductive pili (pilus nanowires). Expression of conductive pili by Geobacter cells leads to biofilm development on surfaces and to the formation of suspended biogranules, which may be physiological closer to biofilms than to planktonic cells. Biofilm development is often assumed in the subsurface, particularly at the matrix-well screen interface, but evidence of biofilms in the bulk aquifer matrix is scarce. Our preliminary results suggest, however, that biofilms develop in the subsurface and contribute to uranium transformations via sorption and reductive mechanisms. In this project we elucidated the mechanism(s) for uranium immobilization mediated by Geobacter biofilms and identified molecular markers to investigate if biofilm development is happening in the contaminated subsurface. The results provided novel insights needed in order to understand the metabolic potential and physiology of microorganisms with a known role in contaminant transformation in situ, thus having a significant positive impact in the SBR Program and providing novel concept to monitor, model, and predict biological behavior during in situ treatments.

  18. Mechanisms Mediating the Vesicant Actions of Sulfur Mustard after Cutaneous Exposure

    PubMed Central

    Shakarjian, Michael P.; Heck, Diane E.; Gray, Joshua P.; Sinko, Patrick J.; Gordon, Marion K.; Casillas, Robert P.; Heindel, Ned D.; Gerecke, Donald R.; Laskin, Debra L.; Laskin, Jeffrey D.

    2010-01-01

    Sulfur mustard (SM), a chemical weapon first employed during World War I, targets the skin, eyes, and lung. It remains a significant military and civilian threat. The characteristic response of human skin to SM involves erythema of delayed onset, followed by edema with inflammatory cell infiltration, the appearance of large blisters in the affected area, and a prolonged healing period. Several in vivo and in vitro models have been established to understand the pathology and investigate the mechanism of action of this vesicating agent in the skin. SM is a bifunctional alkylating agent which reacts with many targets including lipids, proteins, and DNA, forming both intra- and intermolecular cross-links. Despite the relatively nonselective chemical reactivity of this agent, basal keratinocytes are more sensitive, and blistering involves detachment of these cells from their basement membrane adherence zones. The sequence and manner in which these cells die and detach is still unresolved. Much has been discovered over the past two decades with respect to the mechanisms of SM-induced cytotoxicity and the intracellular and extracellular targets of this vesicant. In this review, the effects of SM exposure on the skin are described, as well as potential mechanisms mediating its actions. Successful therapy for SM poisoning will depend on following new mechanistic leads to develop drugs that target one or more of its sites of action. PMID:19833738

  19. The cGMP/PKG pathway as a common mediator of cardioprotection: translatability and mechanism

    PubMed Central

    Inserte, Javier; Garcia-Dorado, David

    2015-01-01

    Cardiomyocyte cell death occurring during myocardial reperfusion (reperfusion injury) contributes to final infarct size after transient coronary occlusion. Different interrelated mechanisms of reperfusion injury have been identified, including alterations in cytosolic Ca2+ handling, sarcoplasmic reticulum-mediated Ca2+ oscillations and hypercontracture, proteolysis secondary to calpain activation and mitochondrial permeability transition. All these mechanisms occur during the initial minutes of reperfusion and are inhibited by intracellular acidosis. The cGMP/PKG pathway modulates the rate of recovery of intracellular pH, but has also direct effect on Ca2+ oscillations and mitochondrial permeability transition. The cGMP/PKG pathway is depressed in cardiomyocytes by ischaemia/reperfusion and preserved by ischaemic postconditioning, which importantly contributes to postconditioning protection. The present article reviews the mechanisms and consequences of the effect of ischaemic postconditioning on the cGMP/PKG pathway, the different pharmacological strategies aimed to stimulate it during myocardial reperfusion and the evidence, limitations and promise of translation of these strategies to the clinical practice. Overall, the preclinical and clinical evidence suggests that modulation of the cGMP/PKG pathway may be a therapeutic target in the context of myocardial infarction. PMID:25297462

  20. Nitric oxide-mediated mitochondrial damage in the brain: mechanisms and implications for neurodegenerative diseases.

    PubMed

    Bolaños, J P; Almeida, A; Stewart, V; Peuchen, S; Land, J M; Clark, J B; Heales, S J

    1997-06-01

    Within the CNS and under normal conditions, nitric oxide (.NO) appears to be an important physiological signalling molecule. Its ability to increase cyclic GMP concentration suggests that .NO is implicated in the regulation of important metabolic pathways in the brain. Under certain circumstances .NO synthesis may be excessive and .NO may become neurotoxic. Excessive glutamate-receptor stimulation may lead to neuronal death through a mechanism implicating synthesis of both .NO and superoxide (O2.-) and hence peroxynitrite (ONOO-) formation. In response to lipopolysaccharide and cytokines, glial cells may also be induced to synthesize large amounts of .NO, which may be deleterious to the neighbouring neurones and oligodendrocytes. The precise mechanism of .NO neurotoxicity is not fully understood. One possibility is that it may involve neuronal energy deficiency. This may occur by ONOO- interfering with key enzymes of the tricarboxylic acid cycle, the mitochondrial respiratory chain, mitochondrial calcium metabolism, or DNA damage with subsequent activation of the energy-consuming pathway involving poly(ADP-ribose) synthetase. Possible mechanisms whereby ONOO- impairs the mitochondrial respiratory chain and the relevance for neurotoxicity are discussed. The intracellular content of reduced glutathione also appears important in determining the sensitivity of cells to ONOO- production. It is concluded that neurotoxicity elicited by excessive .NO production may be mediated by mitochondrial dysfunction leading to an energy deficiency state.

  1. Metabotropic glutamate receptor-mediated long-term depression: molecular mechanisms.

    PubMed

    Gladding, Clare M; Fitzjohn, Stephen M; Molnár, Elek

    2009-12-01

    The ability to modify synaptic transmission between neurons is a fundamental process of the nervous system that is involved in development, learning, and disease. Thus, synaptic plasticity is the ability to bidirectionally modify transmission, where long-term potentiation and long-term depression (LTD) represent the best characterized forms of plasticity. In the hippocampus, two main forms of LTD coexist that are mediated by activation of either N-methyl-d-aspartic acid receptors (NMDARs) or metabotropic glutamate receptors (mGluRs). Compared with NMDAR-LTD, mGluR-LTD is less well understood, but recent advances have started to delineate the underlying mechanisms. mGluR-LTD at CA3:CA1 synapses in the hippocampus can be induced either by synaptic stimulation or by bath application of the group I selective agonist (R,S)-3,5-dihydroxyphenylglycine. Multiple signaling mechanisms have been implicated in mGluR-LTD, illustrating the complexity of this form of plasticity. This review provides an overview of recent studies investigating the molecular mechanisms underlying hippocampal mGluR-LTD. It highlights the role of key molecular components and signaling pathways that are involved in the induction and expression of mGluR-LTD and considers how the different signaling pathways may work together to elicit a persistent reduction in synaptic transmission.

  2. Mechanisms of N-acetyl cysteine-mediated protection from 2-hydroxyethyl methacrylate-induced apoptosis.

    PubMed

    Paranjpe, Avina; Cacalano, Nicholas A; Hume, Wyatt R; Jewett, Anahid

    2008-10-01

    Resin-based materials are now commonly used in dentistry in restorative materials as well as in endodontic sealers. These materials have been shown to be cytotoxic. The mechanisms by which resin-based materials mediate their adverse effects have not been completely elucidated. Here we show that 2-hydroxyethyl methacrylate (HEMA) induces apoptotic cell death in oral keratinocytes and immune cells through the intrinsic cell death pathway. Functional loss and cell death induced by HEMA was significantly inhibited in the presence of N-acetyl cysteine (NAC) treatment. In addition, HEMA induced a decrease in mitochondrial membrane potential, and an increase in cleaved caspases was potently inhibited in the presence of NAC treatment. Overall, the results reported in this article indicate that NAC is an effective chemoprotectant that can safely be used to protect the pulp and the surrounding tissues from adverse effects of dental restorative and endodontic materials.

  3. Molecular and cellular mechanisms of dopamine-mediated behavioral plasticity in the striatum.

    PubMed

    Cerovic, Milica; d'Isa, Raffaele; Tonini, Raffaella; Brambilla, Riccardo

    2013-10-01

    The striatum is the input structure of the basal ganglia system. By integrating glutamatergic signals from cortical and subcortical regions and dopaminergic signals from mesolimbic nuclei the striatum functions as an important neural substrate for procedural and motor learning as well as for reward-guided behaviors. In addition, striatal activity is significantly altered in pathological conditions in which either a loss of dopamine innervation (Parkinson's disease) or aberrant dopamine-mediated signaling (drug addiction and L-DOPA induced dyskinesia) occurs. Here we discuss cellular mechanisms of striatal synaptic plasticity and aspects of cell signaling underlying striatum-dependent behavior, with a major focus on the neuromodulatory action of the endocannabinoid system and on the role of the Ras-ERK cascade.

  4. "Epithelial Cell TRPV1-Mediated Airway Sensitivity as a Mechanism for Respiratory Symptoms Associated with Gulf War Illness?

    DTIC Science & Technology

    2010-06-01

    TITLE: “Epithelial Cell TRPV1 -Mediated Airway Sensitivity as a Mechanism for Respiratory Symptoms Associated with Gulf War Illness” PRINCIPAL...66,),&$7,212) E7(/(3+21(180%(5 ,QFOXGHDUHDFRGH 01-06-2010 Annual Report 1 JUN 2009 - 31 MAY 2010 Epithelial Cell TRPV1 -Mediated Airway...express functional TRPV1 . More recently we found that these cells also express another important irritant receptor, namely TRPA1. Activation of

  5. One night of total sleep deprivation promotes a state of generalized hyperalgesia: a surrogate pain model to study the relationship of insomnia and pain.

    PubMed

    Schuh-Hofer, Sigrid; Wodarski, Rachel; Pfau, Doreen B; Caspani, Ombretta; Magerl, Walter; Kennedy, Jeffrey D; Treede, Rolf-Detlef

    2013-09-01

    Sleep disturbances are highly prevalent in chronic pain patients. Understanding their relationship has become an important research topic since poor sleep and pain are assumed to closely interact. To date, human experimental studies exploring the impact of sleep disruption/deprivation on pain perception have yielded conflicting results. This inconsistency may be due to the large heterogeneity of study populations and study protocols previously used. In addition, none of the previous studies investigated the entire spectrum of nociceptive modalities. To address these shortcomings, a standardized comprehensive quantitative sensory protocol was used in order to compare the somatosensory profile of 14 healthy subjects (6 female, 8 male, 23.5 ± 4.1 year; mean ± SD) after a night of total sleep deprivation (TSD) and a night of habitual sleep in a cross-over design. One night of TSD significantly increased the level of sleepiness (P<0.001) and resulted in higher scores of the State Anxiety Inventory (P<0.01). In addition to previously reported hyperalgesia to heat (P<0.05) and blunt pressure (P<0.05), study participants developed hyperalgesia to cold (P<0.01) and increased mechanical pain sensitivity to pinprick stimuli (P<0.05) but no changes in temporal summation. Paradoxical heat sensations or dynamic mechanical allodynia were absent. TSD selectively modulated nociception, since detection thresholds of non-nociceptive modalities remained unchanged. Our findings show that a single night of TSD is able to induce generalized hyperalgesia and to increase State Anxiety scores. In the future, TSD may serve as a translational pain model to elucidate the pathomechanisms underlying the hyperalgesic effect of sleep disturbances.

  6. A reappraisal of humoral immunity based on mechanisms of antibody-mediated protection against intracellular pathogens.

    PubMed

    Casadevall, Arturo; Pirofski, Liise-anne

    2006-01-01

    Sometime in the mid to late twentieth century the study of antibody-mediated immunity (AMI) entered the doldrums, as many immunologists believed that the function of AMI was well understood, and was no longer deserving of intensive investigation. However, beginning in the 1990s studies using monoclonal antibodies (mAbs) revealed new functions for antibodies, including direct antimicrobial effects and their ability to modify host inflammatory and cellular responses. Furthermore, the demonstration that mAbs to several intracellular bacterial and fungal pathogens were protective issued a serious challenge to the paradigm that host defense against such microbes was strictly governed by cell-mediated immunity (CMI). Hence, a new view of AMI is emerging. This view is based on the concept that a major function of antibody (Ab) is to amplify or subdue the inflammatory response to a microbe. In this regard, the "damage-response framework" of microbial pathogenesis provides a new conceptual viewpoint for understanding mechanisms of AMI. According to this view, the ability of an Ab to affect the outcome of a host-microbe interaction is a function of its capacity to modify the damage ensuing from such an interaction. In fact, it is increasingly apparent that the efficacy of an Ab cannot be defined either by immunoglobulin or epitope characteristics alone, but rather by a complex function of Ab variables, such as specificity, isotype, and amount, host variables, such as genetic background and immune status, and microbial variables, such as inoculum, mechanisms of avoiding host immune surveillance and pathogenic strategy. Consequently, far from being understood, recent findings in AMI imply a system with unfathomable complexity and the field is poised for a long overdue renaissance.

  7. Mechanisms mediating enhanced neutralization efficacy of Staphylococcal enterotoxin B by combinations of monoclonal antibodies

    DOE PAGES

    Dutta, Kaushik; Varshney, Avanish K.; Franklin, Matthew C.; ...

    2015-01-08

    Staphylococcal enterotoxin B (SEB) is a superantigen that cross-links the major histocompatibility complex class II and specific V-β chains of the T-cell receptor, thus forming a ternary complex. Developing neutralizing mAb to disrupt the ternary complex and abrogate the resulting toxicity is a major therapeutic challenge because SEB is effective at very low concentrations. We show that combining two SEB-specific mAbs enhances their efficacy, even though one of the two mAbs by itself has no effect on neutralization. Crystallography was employed for fine-mapping conformational epitopes in binary and ternary complexes between SEB and Fab fragments. NMR spectroscopy was used tomore » validate and identify subtle allosteric changes induced by mAbs binding to SEB. The mapping of epitopes established that a combination of different mAbs can enhance efficacy of mAb-mediated protection from SEB induced lethal shock by two different mechanisms: one mAb mixture promoted clearance of the toxin both in vitro and in vivo by FcR-mediated cross-linking and clearance, whereas the other mAb mixture induced subtle allosteric conformational changes in SEB that perturbed formation of the SEB·T-cell receptor·major histocompatibility complex class II trimer. Lastly structural information accurately predicted mAb binding to other superantigens that share conformational epitopes with SEB. Fine mapping of conformational epitopes is a powerful tool to establish the mechanism and optimize the action of synergistic mAb combinations.« less

  8. Mechanisms mediating enhanced neutralization efficacy of Staphylococcal enterotoxin B by combinations of monoclonal antibodies

    SciTech Connect

    Dutta, Kaushik; Varshney, Avanish K.; Franklin, Matthew C.; Goger, Michael; Wang, Xiaobo; Fries, Bettina C.

    2015-01-08

    Staphylococcal enterotoxin B (SEB) is a superantigen that cross-links the major histocompatibility complex class II and specific V-β chains of the T-cell receptor, thus forming a ternary complex. Developing neutralizing mAb to disrupt the ternary complex and abrogate the resulting toxicity is a major therapeutic challenge because SEB is effective at very low concentrations. We show that combining two SEB-specific mAbs enhances their efficacy, even though one of the two mAbs by itself has no effect on neutralization. Crystallography was employed for fine-mapping conformational epitopes in binary and ternary complexes between SEB and Fab fragments. NMR spectroscopy was used to validate and identify subtle allosteric changes induced by mAbs binding to SEB. The mapping of epitopes established that a combination of different mAbs can enhance efficacy of mAb-mediated protection from SEB induced lethal shock by two different mechanisms: one mAb mixture promoted clearance of the toxin both in vitro and in vivo by FcR-mediated cross-linking and clearance, whereas the other mAb mixture induced subtle allosteric conformational changes in SEB that perturbed formation of the SEB·T-cell receptor·major histocompatibility complex class II trimer. Lastly structural information accurately predicted mAb binding to other superantigens that share conformational epitopes with SEB. Fine mapping of conformational epitopes is a powerful tool to establish the mechanism and optimize the action of synergistic mAb combinations.

  9. Androgen and estrogen receptor mediated mechanisms of testosterone action in male rat pelvic autonomic ganglia

    PubMed Central

    Purves-Tyson, T.D.; Arshi, M.S.; Handelsman, D. J.; Cheng, Y.; Keast, J. R.

    2007-01-01

    Although male reproductive function is primarily androgen dependent, many studies suggest that estrogens have direct actions on the male reproductive organs. Pelvic autonomic neurons provide the motor control of the internal reproductive organs and the penis and various properties of these neurons are affected by endogenous androgens. However, the possible role of estrogens at this site has not been examined. Here we have investigated the significance of estrogens produced by aromatisation of testosterone in the physiological actions of androgens on adult male rat pelvic ganglion neurons. RT-PCR studies showed that aromatase and both estrogen receptors (ERα and ERβ) are expressed in these ganglia. Western blotting also showed that aromatase is expressed in male pelvic ganglia. Using immunohistochemical visualisation, ERα was predominantly expressed by nitric oxide synthase (NOS)-positive parasympathetic pelvic ganglion neurons. In vivo studies showed that the decrease in pelvic ganglion soma size caused by gonadectomy could be prevented by administration of testosterone (T) or dihydrotestosterone (DHT), but not 17β-estradiol (E2), showing that this maintenance action of testosterone is mediated entirely by androgenic mechanisms. However, in vitro studies of cultured pelvic ganglion neurons revealed that T, DHT and E each stimulated the growth of longer and more complex neurites in both noradrenergic and cholinergic NOS-expressing neurons. The effects of T were attenuated by either androgen or estrogen receptor antagonists, or by inhibition of aromatase. Together these studies demonstrate that estrogens are likely to be synthesised in the male pelvic ganglia, produced from testosterone by local aromatase. The effects of androgens on axonal growth are likely to be at least partly mediated by estrogenic mechanisms, which may be important for understanding disease-, aging- and injury-induced plasticity in this part of the nervous system. PMID:17629410

  10. Mechanisms mediating enhanced neutralization efficacy of staphylococcal enterotoxin B by combinations of monoclonal antibodies.

    PubMed

    Dutta, Kaushik; Varshney, Avanish K; Franklin, Matthew C; Goger, Michael; Wang, Xiaobo; Fries, Bettina C

    2015-03-13

    Staphylococcal enterotoxin B (SEB) is a superantigen that cross-links the major histocompatibility complex class II and specific V-β chains of the T-cell receptor, thus forming a ternary complex. Developing neutralizing mAb to disrupt the ternary complex and abrogate the resulting toxicity is a major therapeutic challenge because SEB is effective at very low concentrations. We show that combining two SEB-specific mAbs enhances their efficacy, even though one of the two mAbs by itself has no effect on neutralization. Crystallography was employed for fine-mapping conformational epitopes in binary and ternary complexes between SEB and Fab fragments. NMR spectroscopy was used to validate and identify subtle allosteric changes induced by mAbs binding to SEB. The mapping of epitopes established that a combination of different mAbs can enhance efficacy of mAb-mediated protection from SEB induced lethal shock by two different mechanisms: one mAb mixture promoted clearance of the toxin both in vitro and in vivo by FcR-mediated cross-linking and clearance, whereas the other mAb mixture induced subtle allosteric conformational changes in SEB that perturbed formation of the SEB·T-cell receptor·major histocompatibility complex class II trimer. Finally structural information accurately predicted mAb binding to other superantigens that share conformational epitopes with SEB. Fine mapping of conformational epitopes is a powerful tool to establish the mechanism and optimize the action of synergistic mAb combinations.

  11. Basophil activation test for investigation of IgE-mediated mechanisms in drug hypersensitivity.

    PubMed

    Steiner, Markus; Harrer, Andrea; Lang, Roland; Schneider, Michael; Ferreira, Tima; Hawranek, Thomas; Himly, Martin

    2011-09-16

    used for the determination of IgE-mediated mechanisms in drug hypersensitivity. Here, we determine PP hypersensitivity as IgE-mediated and DF hypersensitivity as non-IgE-mediated by BAT.

  12. Expression and action of cyclic GMP-dependent protein kinase Ialpha in inflammatory hyperalgesia in rat spinal cord.

    PubMed

    Tao, Y X; Hassan, A; Haddad, E; Johns, R A

    2000-01-01

    important role in the central mechanism of formalin-induced inflammatory hyperalgesia in the spinal cord.

  13. Extracellular matrix hyaluronan signals via its CD44 receptor in the increased responsiveness to mechanical stimulation.

    PubMed

    Ferrari, L F; Araldi, D; Bogen, O; Levine, J D

    2016-06-02

    We propose that the extracellular matrix (ECM) signals CD44, a hyaluronan receptor, to increase the responsiveness to mechanical stimulation in the rat hind paw. We report that intradermal injection of hyaluronidase induces mechanical hyperalgesia, that is inhibited by co-administration of a CD44 receptor antagonist, A5G27. The intradermal injection of low (LMWH) but not high (HMWH) molecular weight hyaluronan also induces mechanical hyperalgesia, an effect that was attenuated by pretreatment with HMWH or A5G27. Pretreatment with HMWH also attenuated the hyperalgesia induced by hyaluronidase. Similarly, intradermal injection of A6, a CD44 receptor agonist, produced hyperalgesia that was inhibited by HMWH and A5G27. Inhibitors of protein kinase A (PKA) and Src, but not protein kinase C (PKC), significantly attenuated the hyperalgesia induced by both A6 and LMWH. Finally, to determine if CD44 receptor signaling is involved in a preclinical model of inflammatory pain, we evaluated the effect of A5G27 and HMWH on the mechanical hyperalgesia associated with the inflammation induced by carrageenan. Both A5G27 and HMWH attenuated carrageenan-induced mechanical hyperalgesia. Thus, while LMWH acts at its cognate receptor, CD44, to induce mechanical hyperalgesia, HMWH acts at the same receptor as an antagonist. That the local administration of HMWH or A5G27 inhibits carrageenan-induced hyperalgesia supports the suggestion that carrageenan produces changes in the ECM that contributes to inflammatory pain. These studies define a clinically relevant role for signaling by the hyaluronan receptor, CD44, in increased responsiveness to mechanical stimulation.

  14. Ca{sup 2+} influx and ATP release mediated by mechanical stretch in human lung fibroblasts

    SciTech Connect

    Murata, Naohiko; Ito, Satoru; Furuya, Kishio; Takahara, Norihiro; Naruse, Keiji; Aso, Hiromichi; Kondo, Masashi; Sokabe, Masahiro; Hasegawa, Yoshinori

    2014-10-10

    Highlights: • Uniaxial stretching activates Ca{sup 2+} signaling in human lung fibroblasts. • Stretch-induced intracellular Ca{sup 2+} elevation is mainly via Ca{sup 2+} influx. • Mechanical strain enhances ATP release from fibroblasts. • Stretch-induced Ca{sup 2+} influx is not mediated by released ATP or actin cytoskeleton. - Abstract: One cause of progressive pulmonary fibrosis is dysregulated wound healing after lung inflammation or damage in patients with idiopathic pulmonary fibrosis and severe acute respiratory distress syndrome. The mechanical forces are considered to regulate pulmonary fibrosis via activation of lung fibroblasts. In this study, the effects of mechanical stretch on the intracellular Ca{sup 2+} concentration ([Ca{sup 2+}]{sub i}) and ATP release were investigated in primary human lung fibroblasts. Uniaxial stretch (10–30% in strain) was applied to fibroblasts cultured in a silicone chamber coated with type I collagen using a stretching apparatus. Following stretching and subsequent unloading, [Ca{sup 2+}]{sub i} transiently increased in a strain-dependent manner. Hypotonic stress, which causes plasma membrane stretching, also transiently increased the [Ca{sup 2+}]{sub i}. The stretch-induced [Ca{sup 2+}]{sub i} elevation was attenuated in Ca{sup 2+}-free solution. In contrast, the increase of [Ca{sup 2+}]{sub i} by a 20% stretch was not inhibited by the inhibitor of stretch-activated channels GsMTx-4, Gd{sup 3+}, ruthenium red, or cytochalasin D. Cyclic stretching induced significant ATP releases from fibroblasts. However, the stretch-induced [Ca{sup 2+}]{sub i} elevation was not inhibited by ATP diphosphohydrolase apyrase or a purinergic receptor antagonist suramin. Taken together, mechanical stretch induces Ca{sup 2+} influx independently of conventional stretch-sensitive ion channels, the actin cytoskeleton, and released ATP.

  15. Biomechanical, biochemical, and morphological mechanisms of heat shock-mediated germination in Carica papaya seed

    PubMed Central

    Webster, Rachel E.; Waterworth, Wanda M.; Stuppy, Wolfgang; West, Christopher E.; Ennos, Roland; Bray, Clifford M.; Pritchard, Hugh W.

    2016-01-01

    Carica papaya (papaya) seed germinate readily fresh from the fruit, but desiccation induces a dormant state. Dormancy can be released by exposure of the hydrated seed to a pulse of elevated temperature, typical of that encountered in its tropical habitat. Carica papaya is one of only a few species known to germinate in response to heat shock (HS) and we know little of the mechanisms that control germination in tropical ecosystems. Here we investigate the mechanisms that mediate HS-induced stimulation of germination in pre-dried and re-imbibed papaya seed. Exogenous gibberellic acid (GA3 ≥250 µM) overcame the requirement for HS to initiate germination. However, HS did not sensitise seeds to GA3, indicative that it may act independently of GA biosynthesis. Seed coat removal also overcame desiccation-imposed dormancy, indicative that resistance to radicle emergence is coat-imposed. Morphological and biomechanical studies identified that neither desiccation nor HS alter the physical structure or the mechanical strength of the seed coat. However, cycloheximide prevented both seed coat weakening and germination, implicating a requirement for de novo protein synthesis in both processes. The germination antagonist abscisic acid prevented radicle emergence but had no effect on papaya seed coat weakening. Desiccation therefore appears to reduce embryo growth potential, which is reversed by HS, without physically altering the mechanical properties of the seed coat. The ability to germinate in response to a HS may confer a competitive advantage to C. papaya, an opportunistic pioneer species, through detection of canopy removal in tropical forests. PMID:27811004

  16. Biomechanical, biochemical, and morphological mechanisms of heat shock-mediated germination in Carica papaya seed.

    PubMed

    Webster, Rachel E; Waterworth, Wanda M; Stuppy, Wolfgang; West, Christopher E; Ennos, Roland; Bray, Clifford M; Pritchard, Hugh W

    2016-12-01

    Carica papaya (papaya) seed germinate readily fresh from the fruit, but desiccation induces a dormant state. Dormancy can be released by exposure of the hydrated seed to a pulse of elevated temperature, typical of that encountered in its tropical habitat. Carica papaya is one of only a few species known to germinate in response to heat shock (HS) and we know little of the mechanisms that control germination in tropical ecosystems. Here we investigate the mechanisms that mediate HS-induced stimulation of germination in pre-dried and re-imbibed papaya seed. Exogenous gibberellic acid (GA3 ≥250 µM) overcame the requirement for HS to initiate germination. However, HS did not sensitise seeds to GA3, indicative that it may act independently of GA biosynthesis. Seed coat removal also overcame desiccation-imposed dormancy, indicative that resistance to radicle emergence is coat-imposed. Morphological and biomechanical studies identified that neither desiccation nor HS alter the physical structure or the mechanical strength of the seed coat. However, cycloheximide prevented both seed coat weakening and germination, implicating a requirement for de novo protein synthesis in both processes. The germination antagonist abscisic acid prevented radicle emergence but had no effect on papaya seed coat weakening. Desiccation therefore appears to reduce embryo growth potential, which is reversed by HS, without physically altering the mechanical properties of the seed coat. The ability to germinate in response to a HS may confer a competitive advantage to C. papaya, an opportunistic pioneer species, through detection of canopy removal in tropical forests.

  17. Cholangiocytes express the aquaporin CHIP and transport water via a channel-mediated mechanism.

    PubMed Central

    Roberts, S K; Yano, M; Ueno, Y; Pham, L; Alpini, G; Agre, P; LaRusso, N F

    1994-01-01

    Cholangiocytes line the intrahepatic bile ducts and regulate salt and water secretion during bile formation, but the mechanism(s) regulating ductal water movement remains obscure. A water-selective channel, the aquaporin CHIP, was recently described in several epithelia, so we tested the hypothesis that osmotic water movement by cholangiocytes is mediated by CHIP. Isolated rodent cholangiocytes showed a rapid increase in volume in the presence of hypotonic extracellular buffers; the ratio of osmotic to diffusional permeability coefficients was > 10. The osmotically induced increase in cholangiocyte volume was inversely proportional to buffer osmolality, independent of temperature, and reversibly blocked by HgCl2. Also, the luminal area of isolated, enclosed bile duct units increased after exposure to hypotonic buffer and was reversibly inhibited by HgCl2. RNase protection assays, anti-CHIP immunoblots, and immunocytochemistry confirmed that CHIP transcript and protein were present in isolated cholangiocytes but not in hepatocytes. These results demonstrate that (i) isolated cholangiocytes and intact, polarized bile duct units manifest rapid, mercury-sensitive increases in cell size and luminal area, respectively, in response to osmotic gradients and (ii) isolated cholangiocytes express aquaporin CHIP at both the mRNA and the protein level. The data implicate aquaporin water channels in the transcellular movement of water across cholangiocytes lining intrahepatic bile ducts and provide a plausible molecular explanation for ductal water secretion. Images Fig. 1 Fig. 4 Fig. 5 PMID:7528928

  18. Mechanism of arctigenin-mediated specific cytotoxicity against human lung adenocarcinoma cell lines.

    PubMed

    Susanti, Siti; Iwasaki, Hironori; Inafuku, Masashi; Taira, Naoyuki; Oku, Hirosuke

    2013-12-15

    The lignan arctigenin (ARG) from the herb Arctium lappa L. possesses anti-cancer activity, however the mechanism of action of ARG has been found to vary among tissues and types of cancer cells. The current study aims to gain insight into the ARG mediated mechanism of action involved in inhibiting proliferation and inducing apoptosis in lung adenocarcinoma cells. This study also delineates the cancer cell specificity of ARG by comparison with its effects on various normal cell lines. ARG selectively arrested the proliferation of cancer cells at the G0/G1 phase through the down-regulation of NPAT protein expression. This down-regulation occurred via the suppression of either cyclin E/CDK2 or cyclin H/CDK7, while apoptosis was induced through the modulation of the Akt-1-related signaling pathway. Furthermore, a GSH synthase inhibitor specifically enhanced the cytotoxicity of ARG against cancer cells, suggesting that the intracellular GSH content was another factor influencing the susceptibility of cancer cells to ARG. These findings suggest that specific cytotoxicity of ARG against lung cancer cells was explained by its selective modulation of the expression of NPAT, which is involved in histone biosynthesis. The cytotoxicity of ARG appeared to be dependent on the intracellular GSH level.

  19. Actin polymerization does not provide direct mechanical forces for vesicle fission during clathrin-mediated endocytosis.

    PubMed

    Yao, Li-Hua; Rao, Yan; Bang, Chi; Kurilova, Svetlana; Varga, Kelly; Wang, Chun-Yang; Weller, Brandon D; Cho, Wonhwa; Cheng, Jun; Gong, Liang-Wei

    2013-10-02

    Actin polymerization is important for vesicle fission during clathrin-mediated endocytosis (CME), and it has been proposed that actin polymerization may promote vesicle fission during CME by providing direct mechanical forces. However, there is no direct evidence in support of this hypothesis. In the present study, the role of actin polymerization in vesicle fission was tested by analyzing the kinetics of the endocytic tubular membrane neck (the fission-pore) with cell-attached capacitance measurements to detect CME of single vesicles in a millisecond time resolution in mouse chromaffin cells. Inhibition in dynamin GTPase activity increased the fission-pore conductance (Gp), supporting the mechanical role of dynamin GTPase in vesicle fission. However, disruptions in actin polymerization did not alter the fission-pore conductance Gp, thus arguing against the force-generating role of actin polymerization in vesicle fission during CME. Similar to disruptions of actin polymerization, cholesterol depletion results in an increase in the fission-pore duration, indicating a role for cholesterol-dependent membrane reorganization in vesicle fission. Further experiments suggested that actin polymerization and cholesterol might function in vesicle fission during CME in the same pathway. Our results thus support a model in which actin polymerization promotes vesicle fission during CME by inducing cholesterol-dependent membrane reorganization.

  20. Bottom-up and top-down mechanisms indirectly mediate interactions between benthic biotic ecosystem components

    NASA Astrophysics Data System (ADS)

    Van Colen, Carl; Thrush, Simon F.; Parkes, Samantha; Harris, Rachel; Woodin, Sally A.; Wethey, David S.; Pilditch, Conrad A.; Hewitt, Judi E.; Lohrer, Andrew M.; Vincx, Magda

    2015-04-01

    The loss or decline in population size of key species can instigate a cascade of effects that have implications for interacting species, therewith impacting biodiversity and ecosystem functioning. We examined how top-down and bottom-up interactions may mediate knock-on effects of a coastal deposit-feeding clam, Macomona liliana (hereafter Macomona), on sandflat meiobenthos densities. Therefore we manipulated densities of Macomona in combination with predator exclusion and experimental shading that was expected to alter microphytobenthos biomass. We show that Macomona regulated densities of meiobenthic (38-500 μm) nematodes, copepods, polychaetes, turbellarians, and ostracodes during the three months of incubation via indirect mechanisms. Predator pressure on Macomona by eagle rays (Myliobatis tenuicaudatus) was found to have a negative effect on densities of some meiobenthic taxa. Furthermore, experimental shading resulted in the loss of a positive relation between Macomona and microphytobenthos biomass, while concurrently increasing the density of some meiobenthic taxa. We suggest that this observation can be explained by the release from bioturbation interference effects of the cockle Austrovenus stutchburyi that was found to thrive in the presence of Macomona under non-shaded conditions. Our results highlight the importance of interactions between macrofaunal bioturbation, microphyte biomass, sediment stability, and predation pressure for the structuring of benthic communities. This experiment illustrates that manipulative field experiments may be particularly suitable to study such multiple indirect mechanisms that regulate ecosystem diversity and related functioning because such approaches may best capture the complex feedbacks and processes that determine ecosystem dynamics.

  1. Endothelial Cells in Antibody-Mediated Rejection of Kidney Transplantation: Pathogenesis Mechanisms and Therapeutic Implications

    PubMed Central

    Wang, Shuo; Wang, Jina; Yang, Cheng; Xu, Ming

    2017-01-01

    Antibody-mediated rejection (AMR) has been identified as a main obstacle for stable immune tolerance and long survival of kidney allografts. In spite of new insights into the underlying mechanisms of AMR, accurate diagnosis and efficient treatment are still challenges in clinical practice. Endothelium is the first barrier between recipients' immune systems and grafts in vascularized organ transplants. Considering that endothelial cells express a number of antigens that can be attacked by various allo- and autoantibodies, endothelial cells act as main targets for the recipients' humoral immune responses. Importantly, emerging evidence has shown that endothelial cells in transplants could also initiate protective mechanisms in response to immune injuries. A better understanding of the role of endothelial cells during the pathogenesis of AMR might provide novel therapeutic targets. In the present review, we summarize the antigens expressed by endothelial cells and also discuss the activation and accommodation of endothelial cells as well as their clinical implications. Collectively, the progress discussed in this review indicates endothelial cells as promising targets to improve current diagnosis and therapeutic regimens for AMR. PMID:28255564

  2. Endothelial Cells in Antibody-Mediated Rejection of Kidney Transplantation: Pathogenesis Mechanisms and Therapeutic Implications.

    PubMed

    Wang, Shuo; Zhang, Chao; Wang, Jina; Yang, Cheng; Xu, Ming; Rong, Ruiming; Zhu, Tongyu; Zhu, Dong

    2017-01-01

    Antibody-mediated rejection (AMR) has been identified as a main obstacle for stable immune tolerance and long survival of kidney allografts. In spite of new insights into the underlying mechanisms of AMR, accurate diagnosis and efficient treatment are still challenges in clinical practice. Endothelium is the first barrier between recipients' immune systems and grafts in vascularized organ transplants. Considering that endothelial cells express a number of antigens that can be attacked by various allo- and autoantibodies, endothelial cells act as main targets for the recipients' humoral immune responses. Importantly, emerging evidence has shown that endothelial cells in transplants could also initiate protective mechanisms in response to immune injuries. A better understanding of the role of endothelial cells during the pathogenesis of AMR might provide novel therapeutic targets. In the present review, we summarize the antigens expressed by endothelial cells and also discuss the activation and accommodation of endothelial cells as well as their clinical implications. Collectively, the progress discussed in this review indicates endothelial cells as promising targets to improve current diagnosis and therapeutic regimens for AMR.

  3. Dexmedetomidine blocks thermal hyperalgesia and spinal glial activation in rat model of monoarthritis

    PubMed Central

    Xu, Bo; Zhang, Wei-shi; Yang, Jia-le; Xu, Hua; Deng, Xiao-ming; Zhang, Yu-qiu

    2010-01-01

    Aim: To investigate the effect of systemic administration dexmedetomidine, a selective alpha 2 adrenergic receptor (α2AR) agonist, on thermal hyperalgesia and spinal glial activation evoked by monoarthritis (MA). Methods: MA was induced by an intra-articular injection of complete Freund's adjuvant (CFA). Thermal hyperalgesia was measured by Hargreaves' test. The spinal glial activation status was analyzed by GFAP (an astrocytic marker) and Iba-1 (a microglial marker) immunohistochemistry or immunoblotting. Results: Unilateral intra-articular injection of CFA produced a robust glial activation of astrocytes and microglia in the spinal cord, which was associated with the development and maintenance of thermal hyperalgesia. Intraperitoneal (ip) injection of dexmedetomidine (2.5 and 10 μg/kg) was repeatedly given once daily for 5 days with the first injection 60 min before intra-articular CFA. At the dose of 10 μg/kg, dexmedetomidine significantly attenuated MA-induced ipsilateral hyperalgesia from day 2 to day 5. MA-induced up-regulation of GFAP expression on both sides of the spinal dorsal horn was significantly suppressed by day 5 post-MA following dexmedetomidine application, whereas MA-induced Iba-1 up-regulation was only partially suppressed. Conclusion: Systemic dexmedetomidine inhibits the activation of spinal glia, which is possibly associated with its antihyperalgesia in monoarthritic rats. PMID:20364156

  4. Intracellular mechanisms mediating tocotrienol-induced apoptosis in neoplastic mammary epithelial cells.

    PubMed

    Sylvester, Paul W; Shah, Sumit

    2005-01-01

    Tocotrienols and tocopherols represent the two subgroups that make up the vitamin E family of compounds. However, tocotrienols display significantly more potent apoptotic activity in neoplastic mammary epithelial cells than tocopherols. Studies were conducted to determine the intracellular mechanism(s) mediating tocotrienol-induced apoptosis in neoplastic +SA mouse mammary epithelial cells in vitro. An initial step in apoptosis is the activation of 'initiator' caspases (caspase-8 or -9) that subsequently activate 'effector' caspases (caspase-3, -6 and -7) and induce apoptosis. Treatment with cytotoxic doses of alpha-tocotrienol (20 microM) resulted in a time-dependent increase in caspase-8 and caspase-3 activity. Combined treatment with specific caspase-8 or caspase-3 inhibitors completely blocked alpha-tocotrienol-induced apoptosis and caspase-8 or caspase-3 activity, respectively. In contrast, alpha-tocotrienol treatment had no effect on caspase-9 activation, and combined treatment with a specific caspase-9 inhibitor did not block alpha-tocotrienol-induced apoptosis in (+)SA cells. Since caspase-8 activation is associated with the activation of death receptors, such as Fas, tumor necrosis factor (TNF), or TNF-related apoptosis-inducing ligand (TRAIL) receptors, studies were conducted to determine the exact death receptor(s) and ligand(s) involved in mediating tocotrienol-induced caspase-8 activation and apoptosis. Treatment with Fas-ligand (FasL), Fas-activating antibody, or TRAIL failed to induce cell death in (+)SA neoplastic mammary epithelial cells, suggesting that these cells are resistant to death receptor-induced apoptosis. Moreover, treatment with cytotoxic doses of alpha-tocotrienol did not alter the intracellular levels of Fas, FasL, or Fas-associated death domain (FADD) in these cells. Western blot analysis also showed that alpha-tocotrienol did not induce FasL or FADD translocation from the cytosolic to membrane fraction in these cells. Finally

  5. Evaluation of the analgesic efficacy and psychoactive effects of AZD1940, a novel peripherally acting cannabinoid agonist, in human capsaicin-induced pain and hyperalgesia.

    PubMed

    Kalliomäki, Jarkko; Annas, Peter; Huizar, Karin; Clarke, Cyril; Zettergren, Annika; Karlsten, Rolf; Segerdahl, Märta

    2013-03-01

    The aim of the present study was to investigate the effects of AZD1940, a novel peripherally acting cannabinoid CB(1) /CB(2) receptor agonist, on capsaicin-induced pain and hyperalgesia, as well as on biomarkers of cannabinoid central nervous system (CNS) effects. The present study was a randomized, double-blind, placebo-controlled, four-sequence, two-period, cross-over study in 44 male healthy volunteers aged 20-45 years. The effects of two single oral doses of AZD1940 (400 and 800 μg) were compared with placebo. Pain intensity after intradermal capsaicin injections in the forearm was assessed on a continuous visual analogue scale (VAS; 0-100 mm). Primary and secondary hyperalgesia induced by application of capsaicin cream on the calf were assessed by measuring heat pain thresholds and the area of mechanical allodynia, respectively. The CNS effects were assessed at baseline and up to 24 h after dosing using a visual analogue mood scales (VAMS) for feeling 'stimulated', 'high', 'anxious', 'sedated' or 'down'. AZD1940 did not significantly attenuate ongoing pain or primary or secondary hyperalgesia compared with placebo. Mild CNS effects for AZD1940were observed on the VAMS for 'high' and 'sedated'. Dose-dependent mild-to-moderate CNS-related and gastrointestinal adverse events were reported following treatment with AZD1940. No evidence of analgesic efficacy was found for a peripherally acting CB(1)/CB(2) receptor agonist in the human capsaicin pain model. The emergence of mild dose-dependent CNS effects suggests that the dose range predicted from preclinical data had been attained.

  6. Generation of ROS mediated by mechanical waves (ultrasound) and its possible applications.

    PubMed

    Duco, Walter; Grosso, Viviana; Zaccari, Daniel; Soltermann, Arnaldo T

    2016-10-15

    The thermal decomposition of 9,10 diphenylanthracene peroxide (DPAO2) generates DPA and a mix of triplet and singlet molecular oxygen. For DPAO2 the efficiency to produce singlet molecular oxygen is 0.35. On the other hand, it has shown that many thermal reactions can be carried out through the interaction of molecules with ultrasound. Ultrasound irradiation can create hydrodynamic stress (sonomechanical process), inertial cavitation (pyrolitic process) and long range effects mediated by radicals or ROS. Sonochemical reactions can be originated by pyrolytic like process, shock mechanical waves, thermal reactions and radical and ROS mediated reactions. Sonolysis of pure water can yield hydrogen or hydroxyl radicals and hydrogen peroxide (ROS). When DPAO2 in 1,4 dioxane solution is treated with 20 or 24kHz and different power intensity the production of molecular singlet oxygen is observed. Specific scavengers like tetracyclone (TC) are used to demonstrate it. The efficiency now is 0.85 showing that the sonochemical process is much more efficient that the thermal one. Another endoperoxide, artemisinin was also studied. Unlike the concept of photosensitizer of photodynamic therapy, in spite of large amount of reported results in literature, the term sonosensitizer and the sonosensitization process are not well defined. We define sonosensitized reaction as one in which a chemical species decompose as consequence of cavitation phenomena producing ROS or other radicals and some other target species does undergo a chemical reaction. The concept could be reach rapidly other peroxides which are now under experimental studies. For artemisinin, an important antimalarian and anticancer drug, was established that ultrasound irradiation increases the effectiveness of the treatment but without any explanation. We show that artemisinin is an endoperoxide and behaves as a sonosensitizer in the sense of our definition.

  7. MAPK signaling cascades mediate distinct glucocorticoid resistance mechanisms in pediatric leukemia

    PubMed Central

    Jones, Courtney L.; Gearheart, Christy M.; Fosmire, Susan; Delgado-Martin, Cristina; Evensen, Nikki A.; Bride, Karen; Waanders, Angela J.; Pais, Faye; Wang, Jinhua; Bhatla, Teena; Bitterman, Danielle S.; de Rijk, Simone R.; Bourgeois, Wallace; Dandekar, Smita; Park, Eugene; Burleson, Tamara M.; Madhusoodhan, Pillai Pallavi; Teachey, David T.; Raetz, Elizabeth A.; Hermiston, Michelle L.; Müschen, Markus; Loh, Mignon L.; Hunger, Stephen P.; Zhang, Jinghui; Garabedian, Michael J.; Porter, Christopher C.

    2015-01-01

    The outcome for pediatric acute lymphoblastic leukemia (ALL) patients who relapse is dismal. A hallmark of relapsed disease is acquired resistance to multiple chemotherapeutic agents, particularly glucocorticoids. In this study, we performed a genome-scale short hairpin RNA screen to identify mediators of prednisolone sensitivity in ALL cell lines. The incorporation of these data with an integrated analysis of relapse-specific genetic and epigenetic changes allowed us to identify the mitogen-activated protein kinase (MAPK) pathway as a mediator of prednisolone resistance in pediatric ALL. We show that knockdown of the specific MAPK pathway members MEK2 and MEK4 increased sensitivity to prednisolone through distinct mechanisms. MEK4 knockdown increased sensitivity specifically to prednisolone by increasing the levels of the glucocorticoid receptor. MEK2 knockdown increased sensitivity to all chemotherapy agents tested by increasing the levels of p53. Furthermore, we demonstrate that inhibition of MEK1/2 with trametinib increased sensitivity of ALL cells and primary samples to chemotherapy in vitro and in vivo. To confirm a role for MAPK signaling in patients with relapsed ALL, we measured the activation of the MEK1/2 target ERK in matched diagnosis-relapse primary samples and observed increased phosphorylated ERK levels at relapse. Furthermore, relapse samples have an enhanced response to MEK inhibition compared to matched diagnosis samples in xenograft models. Together, our data indicate that inhibition of the MAPK pathway increases chemosensitivity to glucocorticoids and possibly other agents and that the MAPK pathway is an attractive target for prevention and/or treatment of relapsed disease. PMID:26324703

  8. A mechanism of hypoxia-mediated escape from adaptive immunity in cancer cells.

    PubMed

    Barsoum, Ivraym B; Smallwood, Chelsea A; Siemens, D Robert; Graham, Charles H

    2014-02-01

    Immune escape is a fundamental trait of cancer in which mechanistic knowledge is incomplete. Here, we describe a novel mechanism by which hypoxia contributes to tumoral immune escape from cytotoxic T lymphocytes (CTL). Exposure of human or murine cancer cells to hypoxia for 24 hours led to upregulation of the immune inhibitory molecule programmed cell death ligand-1 (PD-L1; also known as B7-H1), in a manner dependent on the transcription factor hypoxia-inducible factor-1α (HIF-1α). In vivo studies also demonstrated cellular colocalization of HIF-1α and PD-L1 in tumors. Hypoxia-induced expression of PD-L1 in cancer cells increased their resistance to CTL-mediated lysis. Using glyceryl trinitrate (GTN), an agonist of nitric oxide (NO) signaling known to block HIF-1α accumulation in hypoxic cells, we prevented hypoxia-induced PD-L1 expression and diminished resistance to CTL-mediated lysis. Moreover, transdermal administration of GTN attenuated tumor growth in mice. We found that higher expression of PD-L1 induced in tumor cells by exposure to hypoxia led to increased apoptosis of cocultured CTLs and Jurkat leukemia T cells. This increase in apoptosis was prevented by blocking the interaction of PD-L1 with PD-1, the PD-L1 receptor on T cells, or by addition of GTN. Our findings point to a role for hypoxia/HIF-1 in driving immune escape from CTL, and they suggest a novel cancer immunotherapy to block PD-L1 expression in hypoxic-tumor cells by administering NO mimetics.

  9. Probing the Role of HDACs and Mechanisms of Chromatin-Mediated Neuroplasticity

    PubMed Central

    Haggarty, Stephen J.; Tsai, Li-Huei

    2011-01-01

    Advancing our understanding of neuroplasticity and the development of novel therapeutics based upon this knowledge is critical in order to improve the treatment and prevention of a myriad of nervous system disorders. Epigenetic mechanisms of neuroplasticity involve the post-translational modification of chromatin and the recruitment or loss of macromolecular complexes that control neuronal activity-dependent gene expression. While over a century after Ramón y Cajal first described nuclear subcompartments and foci that we now know correspond to sites of active transcription with acetylated histones that are under epigenetic control, the rate and extent to which epigenetic processes act in a dynamic and combinatorial fashion to shape experience-dependent phenotypic and behavioral plasticity in response to various types of neuronal stimuli over a range of time scales is only now coming into focus. With growing recognition that a subset of human diseases involving cognitive dysfunction can be classified as ‘chromatinopathies’, in which aberrant chromatin-mediated neuroplasticity plays a causal role in the underlying disease pathophysiology, understanding the molecular nature of epigenetic mechanisms in the nervous system may provide important new avenues for the development of novel therapeutics. In this review, we discuss the chemistry and neurobiology of the histone deacetylase (HDAC) family of chromatin-modifying enzymes, outline the role of HDACs in the epigenetic control of neuronal function, and discuss the potential relevance of these epigenetic mechanisms to the development of therapeutics aiming to enhance memory and neuroplasticity. Finally, open questions, challenges, and critical needs for the field of ‘neuroepigenetics’ in the years to come will be summarized. PMID:21545841

  10. Effect of a 5-lipoxygenase inhibitor on nerve growth factor-induced thermal hyperalgesia in the rat.

    PubMed

    Amann, R; Schuligoi, R; Lanz, I; Peskar, B A

    1996-06-13

    Intraplantar injection of mouse beta (2.5S) nerve growth factor (NGF) caused thermal hyperalgesia and stimulated release of immunoreactive leukotriene B4 from the rat paw skin. Both effects of NGF were prevented by the 5-lipoxygenase inhibitor, (R)-2-[4-quinolin-2-yl-methoxy)phenyl]-2-cyclopentyl acetic acid (BAY X1005). BAY X1005 did not affect bradykinin-induced thermal hyperalgesia. These results suggest the participation of 5-lipoxygenase products of arachidonate in NGF-induced local thermal hyperalgesia.

  11. β-arrestin-1 mediates the TCR-triggered re-routing of distal receptors to the immunological synapse by a PKC-mediated mechanism

    PubMed Central

    Fernández-Arenas, Elena; Calleja, Enrique; Martínez-Martín, Nadia; Gharbi, Severine I; Navajas, Rosana; García-Medel, Noel; Penela, Petronila; Alcamí, Antonio; Mayor, Federico; Albar, Juan P; Alarcón, Balbino

    2014-01-01

    T-cell receptors (TCR) recognize their antigen ligand at the interface between T cells and antigen-presenting cells, known as the immunological synapse (IS). The IS provides a means of sustaining the TCR signal which requires the continual supply of new TCRs. These are endocytosed and redirected from distal membrane locations to the IS. In our search for novel cytoplasmic effectors, we have identified β-arrestin-1 as a ligand of non-phosphorylated resting TCRs. Using dominant-negative and knockdown approaches we demonstrate that β-arrestin-1 is required for the internalization and downregulation of non-engaged bystander TCRs. Furthermore, TCR triggering provokes the β-arrestin-1-mediated downregulation of the G-protein coupled chemokine receptor CXCR4, but not of other control receptors. We demonstrate that β-arrestin-1 recruitment to the TCR, and bystander TCR and CXCR4 downregulation, are mechanistically mediated by the TCR-triggered PKC-mediated phosphorylation of β-arrestin-1 at Ser163. This mechanism allows the first triggered TCRs to deliver a stop migration signal, and to promote the internalization of distal TCRs and CXCR4 and their translocation to the IS. This receptor crosstalk mechanism is critical to sustain the TCR signal. PMID:24502978

  12. Amine oxidation mediated by lysine-specific demethylase 1: quantum mechanics/molecular mechanics insights into mechanism and role of lysine 661.

    PubMed

    Karasulu, Bora; Patil, Mahendra; Thiel, Walter

    2013-09-11

    We report classical molecular dynamics (MD) simulations and combined quantum mechanics/molecular mechanics (QM/MM) calculations to elucidate the catalytic mechanism of the rate-determining amine oxidation step in the lysine-specific demethylase 1 (LSD1)-catalyzed demethylation of the histone tail lysine (H3K4), with flavin adenine dinucleotide (FAD) acting as cofactor. The oxidation of substrate lysine (sLys) involves the cleavage of an α-CH bond accompanied by the transfer of a hydride ion equivalent to FAD, leading to an imine intermediate. This hydride transfer pathway is shown to be clearly favored for sLys oxidation over other proposed mechanisms, including the radical (or single-electron transfer) route as well as carbanion and polar-nucleophilic mechanisms. MD simulations on six NVT ensembles (covering different protonation states of sLys and K661 as well as the K661M mutant) identify two possible orientations of the reacting sLys and FAD subunits (called "downward" and "upward"). Calculations at the QM(B3LYP-D/6-31G*)/CHARMM22 level provide molecular-level insights into the mechanism, helping to understand how LSD1 achieves the activation of the rather inert methyl-CH bond in a metal-free environment. Factors such as proper alignment of sLys (downward orientation), transition-state stabilization (due to the protein environment and favorable orbital interactions), and product stabilization via adduct formation are found to be crucial for facilitating the oxidative α-CH bond cleavage. The current study also sheds light on the role of important active-site residues (Y761, K661, and W695) and of the conserved water-bridge motif. The steric influence of Y761 helps to position the reaction partners properly, K661 is predicted to get deprotonated prior to substrate binding and to act as an active-site base that accepts a proton from sLys to enable the subsequent amine oxidation, and the water bridge that is stabilized by K661 and W695 mediates this proton

  13. Exploring How School Intra-Organizational Mechanisms Mediate the Effects of External Interventions on Improving Teaching and Learning

    ERIC Educational Resources Information Center

    Sun, Min

    2011-01-01

    This dissertation collects three independent but interrelated studies exploring how school intra-organizational mechanism may mediate the impact of external interventions on improving teaching and learning. This first study examines how high-quality professional development (PD) can promote the diffusion of effective teaching strategies among…

  14. GR-127935-sensitive mechanism mediating hypotension in anesthetized rats: are 5-HT5B receptors involved?

    PubMed

    Sánchez-Maldonado, Carolina; López-Sánchez, Pedro; Anguiano-Robledo, Liliana; Leopoldo, Marcello; Lacivita, Enza; Terrón, José A

    2015-04-01

    The 5-HT1B/1D receptor antagonist, GR-127935, inhibits hypotensive responses produced by the 5-HT1A, 5-HT1B/1D and 5-HT7 receptor agonist, and 5-HT5A/5B receptor ligand, 5-carboxamidotryptamine (5-CT), in rats. This work further characterized the above mechanism using more selective 5-HT1B and 5-HT1D receptor antagonists. Also, expression of 5-HT5A and 5-HT5B receptor mRNAs in blood vessels was searched by reverse transcription polymerase chain reaction. Decreases in diastolic blood pressure induced by 5-CT (0.001-10 μg/kg, intravenously) were analyzed in anesthetized rats that had received intravenous vehicle (1 mL/kg), SB-224289 (5-HT1B antagonist; 0.3 and 1.0 mg/kg), BRL15572 (5-HT1D antagonist; 0.3 and 1.0 mg/kg), SB-224289 + BRL15572 (0.3 mg/kg, each), or SB-224289 + BRL15572 (0.3 mg/kg, each) + GR-127935 (1 mg/kg). Because only the latter treatment inhibited 5-CT-induced hypotension, suggestive of a mechanism unrelated to 5-HT1B/1D receptors, the effects of antagonists/ligands at 5-HT5A (SB-699551, 1 mg/kg), 5-HT6 (SB-399885, 1 mg/kg), and 5-HT1B/1D/5A/5B/7 receptors (ergotamine, 0.1 mg/kg) on 5-CT-induced hypotension were tested. Interestingly, only ergotamine blocked 5-CT-induced responses; this effect closely paralleled that of SB-224289 + BRL-15572 + GR-127935. Neither did ergotamine nor GR-127935 inhibit hypotensive responses induced by the 5-HT7 receptor agonist, LP-44. Faint but clear bands corresponding to 5-HT5A and 5-HT5B receptor mRNAs in aorta and mesenteric arteries were detected. Results suggest that the GR-127935-sensitive mechanism mediating hypotension in rats is unrelated to 5-HT1B, 5-HT1D, 5-HT5A, 5-HT6, and 5-HT7 receptors. This mechanism, however, resembles putative 5-HT5B receptors.

  15. β-Arrestin mediates the Frank-Starling mechanism of cardiac contractility.

    PubMed

    Abraham, Dennis M; Davis, Robert T; Warren, Chad M; Mao, Lan; Wolska, Beata M; Solaro, R John; Rockman, Howard A

    2016-12-13

    The Frank-Starling law of the heart is a physiological phenomenon that describes an intrinsic property of heart muscle in which increased cardiac filling leads to enhanced cardiac contractility. Identified more than a century ago, the Frank-Starling relationship is currently known to involve length-dependent enhancement of cardiac myofilament Ca(2+) sensitivity. However, the upstream molecular events that link cellular stretch to the length-dependent myofilament Ca(2+) sensitivity are poorly understood. Because the angiotensin II type 1 receptor (AT1R) and the multifunctional transducer protein β-arrestin have been shown to mediate mechanosensitive cellular signaling, we tested the hypothesis that these two proteins are involved in the Frank-Starling mechanism of the heart. Using invasive hemodynamics, we found that mice lacking β-arrestin 1, β-arrestin 2, or AT1R were unable to generate a Frank-Starling force in response to changes in cardiac volume. Although wild-type mice pretreated with the conventional AT1R blocker losartan were unable to enhance cardiac contractility with volume loading, treatment with a β-arrestin-biased AT1R ligand to selectively activate β-arrestin signaling preserved the Frank-Starling relationship. Importantly, in skinned muscle fiber preparations, we found markedly impaired length-dependent myofilament Ca(2+) sensitivity in β-arrestin 1, β-arrestin 2, and AT1R knockout mice. Our data reveal β-arrestin 1, β-arrestin 2, and AT1R as key regulatory molecules in the Frank-Starling mechanism, which potentially can be targeted therapeutically with β-arrestin-biased AT1R ligands.

  16. Plasmid-Mediated OqxAB Is an Important Mechanism for Nitrofurantoin Resistance in Escherichia coli

    PubMed Central

    Ng, Ka-Ying; Lo, Wai-U; Law, Pierra Y.; Lai, Eileen Ling-Yi; Wang, Ya; Chow, Kin-Hung

    2015-01-01

    Increasing consumption of nitrofurantoin (NIT) for treatment of acute uncomplicated urinary tract infections (UTI) highlights the need to monitor emerging NIT resistance mechanisms. This study investigated the molecular epidemiology of the multidrug-resistant efflux gene oqxAB and its contribution to nitrofurantoin resistance by using Escherichia coli isolates originating from patients with UTI (n = 205; collected in 2004 to 2013) and food-producing animals (n = 136; collected in 2012 to 2013) in Hong Kong. The oqxAB gene was highly prevalent among NIT-intermediate (11.5% to 45.5%) and -resistant (39.2% to 65.5%) isolates but rare (0% to 1.7%) among NIT-susceptible (NIT-S) isolates. In our isolates, the oqxAB gene was associated with IS26 and was carried by plasmids of diverse replicon types. Multilocus sequence typing revealed that the clones of oqxAB-positive E. coli were diverse. The combination of oqxAB and nfsA mutations was found to be sufficient for high-level NIT resistance. Curing of oqxAB-carrying plasmids from 20 NIT-intermediate/resistant UTI isolates markedly reduced the geometric mean MIC of NIT from 168.9 μg/ml to 34.3 μg/ml. In the plasmid-cured variants, 20% (1/5) of isolates with nfsA mutations were NIT-S, while 80% (12/15) of isolates without nfsA mutations were NIT-S (P = 0.015). The presence of plasmid-based oqxAB increased the mutation prevention concentration of NIT from 128 μg/ml to 256 μg/ml and facilitated the development of clinically important levels of nitrofurantoin resistance. In conclusion, plasmid-mediated oqxAB is an important nitrofurantoin resistance mechanism. There is a great need to monitor the dissemination of this transferable multidrug-resistant efflux pump. PMID:26552976

  17. A peripheral endocannabinoid mechanism contributes to glucocorticoid-mediated metabolic syndrome.

    PubMed

    Bowles, Nicole P; Karatsoreos, Ilia N; Li, Xiaosong; Vemuri, V Kiran; Wood, Jodi-Anne; Li, Zhiying; Tamashiro, Kellie L K; Schwartz, Gary J; Makriyannis, Alexandros M; Kunos, George; Hillard, Cecilia J; McEwen, Bruce S; Hill, Matthew N

    2015-01-06

    Glucocorticoids are known to promote the development of metabolic syndrome through the modulation of both feeding pathways and metabolic processes; however, the precise mechanisms of these effects are not well-understood. Recent evidence shows that glucocorticoids possess the ability to increase endocannabinoid signaling, which is known to regulate appetite, energy balance, and metabolic processes through both central and peripheral pathways. The aim of this study was to determine the role of endocannabinoid signaling in glucocorticoid-mediated obesity and metabolic syndrome. Using a mouse model of excess corticosterone exposure, we found that the ability of glucocorticoids to increase adiposity, weight gain, hormonal dysregulation, hepatic steatosis, and dyslipidemia was reduced or reversed in mice lacking the cannabinoid CB1 receptor as well as mice treated with the global CB1 receptor antagonist AM251. Similarly, a neutral, peripherally restricted CB1 receptor antagonist (AM6545) was able to attenuate the metabolic phenotype caused by chronic corticosterone, suggesting a peripheral mechanism for these effects. Biochemical analyses showed that chronic excess glucocorticoid exposure produced a significant increase in hepatic and circulating levels of the endocannabinoid anandamide, whereas no effect was observed in the hypothalamus. To test the role of the liver, specific and exclusive deletion of hepatic CB1 receptor resulted in a rescue of the dyslipidemic effects of glucocorticoid exposure, while not affecting the obesity phenotype or the elevations in insulin and leptin. Together, these data indicate that glucocorticoids recruit peripheral endocannabinoid signaling to promote metabolic dysregulation, with hepatic endocannabinoid signaling being especially important for changes in lipid metabolism.

  18. Copper mediated decyano decarboxylative coupling of cyanoacetate ligands: Pesci versus Lewis acid mechanism.

    PubMed

    Li, Jiawei; Khairallah, George N; Steinmetz, Vincent; Maitre, Philippe; O'Hair, Richard A J

    2015-05-21

    A combination of gas-phase ion trap multistage mass spectrometry (MS(n)) experiments and density functional theory (DFT) calculations have been used to examine the mechanisms of the sequential decomposition reactions of copper cyanoacetate anions, [(NCCH2CO2)2Cu](-), introduced into the gas-phase via electrospray ionization. Gas phase IR spectroscopy, used to probe the coordination mode of the cyanoacetate ligands, revealed that the initial precursor ions are bound to the Cu via the carboxylate, [NCCH2CO2CuO2CCH2CN], 1. Multistage collision-induced dissociation (CID) of 1 gave sequential losses of CO2 and ethene. DFT calculations suggest that the lowest energy pathways for sequential decarboxylation involve Lewis acid mechanisms in which the binding of the cyanoacetate ligand sequentially rearranges from O to N: [NCCH2CO2CuO2CCH2CN](-) → [NCCH2CO2CuNCCH2CO2](-) → [NCCH2CO2CuNCCH2](-) + CO2 and [NCCH2CO2CuNCCH2](-) → [O2CCH2CNCuNCCH2](-) → [CH2CNCuNCCH2](-) + CO2. Loss of ethene involves sequential rearrangement of the binding of the cyanomethyl carbanion ligands from N to C: [CH2CNCuNCCH2](-) → [NCCH2CuNCCH2](-) → [NCCH2CuCH2CN](-). CH2=CH2 loss then proceeds via a 1,2-dyotropic rearrangement to form [NCCuCH2CH2CN](-) followed by β-cyanide transfer. This study highlights the rich mechanistic possibilities for metal mediated decarboxylation reactions involving ambidentate carboxylate ligands.

  19. Molecular mechanism of trichloroethylene-induced hepatotoxicity mediated by CYP2E1

    SciTech Connect

    Ramdhan, Doni Hikmat; Kamijima, Michihiro; Yamada, Naoyasu; Ito, Yuki; Yanagiba, Yukie; Nakamura, Daichi; Okamura, Ai; Ichihara, Gaku; Aoyama, Toshifumi; Gonzalez, Frank J.; Nakajima, Tamie

    2008-09-15

    Cytochrome P450 (CYP) 2E1 was suggested to be the major enzyme involved in trichloroethylene (TRI) metabolism and TRI-induced hepatotoxicity, although the latter molecular mechanism is not fully understood. The involvement of CYP2E1 in TRI-induced hepatotoxicity and its underlying molecular mechanism were studied by comparing hepatotoxicity in cyp2e1{sup +/+} and cyp2e1{sup -/-} mice. The mice were exposed by inhalation to 0 (control), 1000, or 2000 ppm of TRI for 8 h a day, for 7 days, and TRI-hepatotoxicity was assessed by measuring plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities and histopathology. Urinary metabolites of trichloroethanol and trichloroacetic acid (TCA) were considerably greater in cyp2e1{sup +/+} compared to cyp2e1{sup -/-} mice, suggesting that CYP2E1 is the major P450 involved in the formation of these metabolites. Consistent with elevated plasma ALT and AST activities, cyp2e1{sup +/+} mice in the 2000 ppm group showed histopathological inflammation. TRI significantly upregulated PPAR{alpha}, which might function to inhibit NF{kappa}B p50 and p65 signalling. In addition, TRI-induced NF{kappa}B p52 mRNA, and significantly positive correlation between NF{kappa}B p52 mRNA expression and plasma ALT activity levels were observed, suggesting the involvement of p52 in liver inflammation. Taken together, the current study directly demonstrates that CYP2E1 was the major P450 involved in the first step of the TRI metabolism, and the metabolites produced may have two opposing roles: one inducing hepatotoxicity and the other protecting against the toxicity. Intermediate metabolite(s) from TRI to chloral hydrate produced by CYP2E1-mediated oxidation may be involved in the former, and TCA in the latter.

  20. Therapeutic targeting of myeloid-derived suppressor cells involves a novel mechanism mediated by clusterin

    PubMed Central

    Zhou, Junmin; Donatelli, Sarah S.; Gilvary, Danielle L.; Tejera, Melba M.; Eksioglu, Erika A.; Chen, Xianghong; Coppola, Domenico; Wei, Sheng; Djeu, Julie Y.

    2016-01-01

    Myeloid-derived suppressor cells (MDSCs) constitute a key checkpoint that impedes tumor immunity against cancer. Chemotherapeutic intervention of MDSCs has gained ground as a strategy for cancer therapy but its mechanism remains obscure.We report here a unique mechanism by which monocytic (M)-MDSCs are spared, allowing them to polarize towards M1 macrophages for reactivation of immunity against breast cancer. We first demonstrated that curcumin, like docetaxel (DTX), can selectively target CD11b+Ly6G+Ly6Clow granulocytic (G)-MDSCs, sparing CD11b+Ly6G−Ly6Chigh M-MDSCs, with reduced tumor burden in 4T1-Neu tumor-bearing mice. Curcumin treatment polarized surviving M-MDSCs toward CCR7+ Dectin-1−M1 cells, accompanied by IFN-γ production and cytolytic function in T cells. Selective M-MDSC chemoresistence to curcumin and DTX was mediated by secretory/cytoplasmic clusterin (sCLU). sCLU functions by trapping Bax from mitochondrial translocation, preventing the apoptotic cascade. Importantly, sCLU was only found in M-MDSCs but not in G-MDSCs. Knockdown of sCLU in M-MDSCs and RAW264.7 macrophages was found to reverse their natural chemoresistance. Clinically, breast cancer patients possess sCLU expression only in mature CD68+ macrophages but not in immature CD33+ immunosuppressive myeloid cells infiltrating the tumors. We thus made the seminal discovery that sCLU expression in M-MDSCs accounts for positive immunomodulation by chemotherapeutic agents. PMID:27405665

  1. The Molecular Mechanism of Ethylene-Mediated Root Hair Development Induced by Phosphate Starvation

    PubMed Central

    Song, Li; Yu, Haopeng; Dong, Jinsong; Liu, Dong

    2016-01-01

    Enhanced root hair production, which increases the root surface area for nutrient uptake, is a typical adaptive response of plants to phosphate (Pi) starvation. Although previous studies have shown that ethylene plays an important role in root hair development induced by Pi starvation, the underlying molecular mechanism is not understood. In this work, we characterized an Arabidopsis mutant, hps5, that displays constitutive ethylene responses and increased sensitivity to Pi starvation due to a mutation in the ethylene receptor ERS1. hps5 accumulates high levels of EIN3 protein, a key transcription factor involved in the ethylene signaling pathway, under both Pi sufficiency and deficiency. Pi starvation also increases the accumulation of EIN3 protein. Combined molecular, genetic, and genomic analyses identified a group of genes that affect root hair development by regulating cell wall modifications. The expression of these genes is induced by Pi starvation and is enhanced in the EIN3-overexpressing line. In contrast, the induction of these genes by Pi starvation is suppressed in ein3 and ein3eil1 mutants. EIN3 protein can directly bind to the promoter of these genes, some of which are also the immediate targets of RSL4, a key transcription factor that regulates root hair development. Based on these results, we propose that under normal growth conditions, the level of ethylene is low in root cells; a group of key transcription factors, including RSL4 and its homologs, trigger the transcription of their target genes to promote root hair development; Pi starvation increases the levels of the protein EIN3, which directly binds to the promoters of the genes targeted by RSL4 and its homologs and further increase their transcription, resulting in the enhanced production of root hairs. This model not only explains how ethylene mediates root hair responses to Pi starvation, but may provide a general mechanism for how ethylene regulates root hair development under both stress

  2. Using hierarchical linear growth models to evaluate protective mechanisms that mediate science achievement

    NASA Astrophysics Data System (ADS)

    von Secker, Clare Elaine

    The study of students at risk is a major topic of science education policy and discussion. Much research has focused on describing conditions and problems associated with the statistical risk of low science achievement among individuals who are members of groups characterized by problems such as poverty and social disadvantage. But outcomes attributed to these factors do not explain the nature and extent of mechanisms that account for differences in performance among individuals at risk. There is ample theoretical and empirical evidence that demographic differences should be conceptualized as social contexts, or collections of variables, that alter the psychological significance and social demands of life events, and affect subsequent relationships between risk and resilience. The hierarchical linear growth models used in this dissertation provide greater specification of the role of social context and the protective effects of attitude, expectations, parenting practices, peer influences, and learning opportunities on science achievement. While the individual influences of these protective factors on science achievement were small, their cumulative effect was substantial. Meta-analysis conducted on the effects associated with psychological and environmental processes that mediate risk mechanisms in sixteen social contexts revealed twenty-two significant differences between groups of students. Positive attitudes, high expectations, and more intense science course-taking had positive effects on achievement of all students, although these factors were not equally protective in all social contexts. In general, effects associated with authoritative parenting and peer influences were negative, regardless of social context. An evaluation comparing the performance and stability of hierarchical linear growth models with traditional repeated measures models is included as well.

  3. Microfluidic Emulation of Mechanical Circulatory Support Device Shear-Mediated Platelet Activation

    PubMed Central

    Dimasi, A.; Rasponi, M.; Sheriff, J.; Chiu, W.-C.; Bluestein, D.; Tran, P.L.; Slepian, M. J.; Redaelli, A.

    2016-01-01

    Thrombosis of ventricular assist devices (VADs) compromises their performance, with associated risks of systemic embolization, stroke, pump stop and possible death. Anti-thrombotic (AT) drugs, utilized to limit thrombosis, are largely dosed empirically, with limited testing of their efficacy. Further, such testing, if performed, typically examines efficacy under static conditions, which is not reflective of actual shear-mediated flow. Here we adopted our previously developed Device Thrombogenicity Emulation methodology to design microfluidic platforms able to emulate representative shear stress profiles of mechanical circulatory support (MCS) devices. Our long-term goal is to utilize these systems for point-of-care (POC) personalized testing of AT efficacy under specific, individual shear profiles. First, we designed different types of microfluidic channels able to replicate sample shear stress patterns observed in MCS devices. Second, we explored the flexibility of microfluidic technology in generating dynamic shear stress profiles by modulating the geometrical features of the channels. Finally, we designed microfluidic channel systems able to emulate the shear stress profiles of two commercial VADs. From CFD analyses, the VAD-emulating microfluidic systems were able to replicate the main characteristics of the shear stress waveforms of the macroscale VADs (i.e. shear stress peaks and duration). Our results establish the basis for development of a lab-on-chip POC system able to perform device-specific and patient-specific platelet activation state assays. PMID:26578003

  4. Photoperiodic regulation of testis function in rats: mediation by a circadian mechanism.

    PubMed

    Nelson, R J; Bamat, M K; Zucker, I

    1982-03-01

    Laboratory rats traditionally are classified as nonphotoperiodic because variations in daylength have little or no effect on their gonadal function. After olfactory bulbectomy, however, rats show clear evidence of photoperiodic regulation of the gonads. The present study demonstrates, by means of resonance experiments, that the testicular response to daylength in rats is mediated by a circadian photoperiodic time measurement system similar to that of photoperiodic rodents. Olfactory-bulbectomized rats were maintained in fixed photoperiods in which a 6 h light period was coupled with dark periods of 18, 30, 42, or 54 h (6L:18D, 6L:30D, etc.); a fifth group was maintained in a 45L:10D photoperiod. Rats from the 6L:30D, 6L:54D and 14L:10D photoperiods had testes and seminal vesicle weights, plasma testosterone titers and spermatogenesis indices indicative of functional reproductive status. Rats exposed to the 6L:18D and 6L:42D photoperiods had reduced testicular and seminal vesicle weights, lower testosterone levels and reduced spermatogenesis. We hypothesize that photo- and nonphotoperiodic rodent species use similar mechanisms for distinguishing long from short photoperiods, but differ in the extent to which discrimination of short daylengths is transduced into altered gonadal activity.

  5. Overcoming NS1-Mediated Immune Antagonism Involves Both Interferon-Dependent and Independent Mechanisms

    PubMed Central

    Thakar, Juilee; Schmid, Sonja; Duke, Jamie L.; García-Sastre, Adolfo

    2013-01-01

    To ensure survival, our immune system must overcome the action of pathogen-encoded immune antagonists, such as influenza A nonstructural protein-1 (NS1). NS1 subverts the host interferon (IFN) response at multiple levels and blocks the induction of IFN-β, a critical antiviral cytokine. This immune antagonism can be overcome in some cases. It has been shown that IFN-β is upregulated by 48 h in the lungs of wild-type C57BL/6 mice infected with influenza A. In contrast, it is shown here that IFNB1 continues to be repressed in IFNAR1−/− IL28Rα−/− mice, which are deficient in Type-I and III IFN signaling, implying induction of IFNB1 depends on effective IFN signaling. Despite the complete lack of IFN signaling in this system, some IFN stimulated genes (ISGs) were induced following infection with a Flu strain lacking NS1. While the expression of these viral stress-inducible genes (VSIGs) was initially repressed following infection with wild-type Flu, many of these genes became upregulated by 48 h postinfection. These results demonstrate the existence of IFN-independent mechanisms that can overcome NS1-mediated immune antagonism of VSIGs. PMID:23772952

  6. Photoperiodic regulation of testis function in rats: mediation by a circadian mechanism

    SciTech Connect

    Nelson, R.J.; Bamat, M.K.; Zucker, I.

    1982-05-01

    Laboratory rats traditionally are classified as nonphotoperiodic because variations in daylength have little or no effect on their gonadal function. After olfactory bulbectomy, however, rats show clear evidence of photoperiodic regulation of the gonads. The present study demonstrates, by means of resonance experiments, that the testicular response to daylength in rats is mediated by a circadian photoperiodic time measurement system similar to that of photoperiodic rodents. Olfactory-bulbectomized rats were maintained in fixed photoperiods in which a 6 h light period was coupled with dark periods of 18, 30, 42, or 54 h; a fifth group was maintained in a 45L:10D photoperiod. Rats from the 6L:30D, 6L:54D and 14L:10D photoperiods had testes and seminal vesicle weights, plasma testosterone titers and spermatogenesis indices indicative of functional reproductive status. Rats exposed to the 6L:18D and 6L:42D photoperiods had reduced testicular and seminal vesicle weights, lower testosterone levels and reduced spermatogenesis. Researchers hypothesize that photo- and nonphotoperiodic rodent species use similar mechanisms for distinguishing long from short photoperiods, but differ in the extent to which discrimination of short daylengths is transduced into altered gonadal activity.

  7. Structurally novel steroidal spirooxindole by241 potently inhibits tumor growth mainly through ROS-mediated mechanisms

    PubMed Central

    Shi, Xiao-Jing; Yu, Bin; Wang, Jun-Wei; Qi, Ping-Ping; Tang, Kai; Huang, Xin; Liu, Hong-Min

    2016-01-01

    Cancer cells always have increased ROS levels, thus making them more vulnerable to persistent endogenous oxidative stress. The biochemical difference between cancer and normal cells could be exploited to achieve selective cancer cell killing by exogenous ROS-producing agents. Herein we described a structurally novel steroidal spirooxindole by241 and its anticancer efficacy. By241 exhibited potent inhibition against human cancer cells and less toxic to normal cells. By241 concentration-dependently induced apoptosis of MGC-803 and EC9706 cells, accompanied with the mitochondrial dysfunction and increased ROS levels. NAC can completely restore the decreased cell viability of MGC-803 cells caused by by241, suggesting ROS-mediated mechanisms. The expression levels of proteins involved in the mitochondrion-related pathways were detected, showing increased expression of proapoptotic proteins and decreased expression of anti-apoptotic proteins, and activation of caspases-9/-3, but without activating caspase-8 expression. Pretreatment with Z-VAD-FMK partially rescued by241-induced apoptosis of MGC-803 cells. Additionally, by241 inhibited mTOR, activated p53 and its downstream proteins, cleaved MDM2 and PI3K/AKT as well as NF-κB signaling pathway. In vivo experiments showed that by241 did not have significant acute oral toxicity and exerted good anticancer efficacy against MGC-803 bearing mice models. Therefore, by241 may serve as a lead for further development for cancer therapy. PMID:27527552

  8. MutS mediates heteroduplex loop formation by a translocation mechanism.

    PubMed Central

    Allen, D J; Makhov, A; Grilley, M; Taylor, J; Thresher, R; Modrich, P; Griffith, J D

    1997-01-01

    Interaction of Escherichia coli MutS and MutL with heteroduplex DNA has been visualized by electron microscopy. In a reaction dependent on ATP hydrolysis, complexes between a MutS dimer and a DNA heteroduplex are converted to protein-stabilized, alpha-shaped loop structures with the mismatch in most cases located within the DNA loop. Loop formation depends on ATP hydrolysis and loop size increases linearly with time at a rate of 370 base pairs/min in phosphate buffer and about 10,000 base pairs/min in the HEPES buffer used for repair assay. These observations suggest a translocation mechanism in which a MutS dimer bound to a mismatch subsequently leaves this site by ATP-dependent tracking or unidimensional movement that is in most cases bidirectional from the mispair. In view of the bidirectional capability of the methyl-directed pathway, this reaction may play a role in determination of heteroduplex orientation. The rate of MutS-mediated DNA loop growth is enhanced by MutL, and when both proteins are present, both are found at the base of alpha-loop structures, and both can remain associated with excision intermediates produced in later stages of the reaction. PMID:9250691

  9. Th17 Pathway–Mediated Immunopathogenesis of Schizophrenia: Mechanisms and Implications

    PubMed Central

    Debnath, Monojit; Berk, Michael

    2014-01-01

    Schizophrenia is a highly complex and severe neuropsychiatric disorder with an unknown etiopathology. Evidence for a dysregulated immune system in both the risk for and progression of schizophrenia has recently been overwhelming. Importantly, chronic low-grade inflammation both in the periphery and central nervous system has been shown to contribute predominantly to the pathogenesis of schizophrenia in a subset of individuals. Inflammation in the central nervous system is mediated by a range of proinflammatory cytokines, resident immune cells such as microglia, and brain infiltrating peripheral immunocompetent cells, such as T lymphocytes. Recently, Th17 cells, a subset of T helper cells have emerged as crucial players in mucosal defense against infections. It is linked to atopic, inflammatory, and autoimmune disorders. The risk factors/mechanisms leading to low-grade inflammation in schizophrenia are diverse and include infectious agents, stress, trauma, environmental toxins, genetic vulnerability, physical inactivity, obesity, poor diet, and sleep disruption. Herein, we propose that fetal programming of cellular immune components driven by intrauterine adversity can lead to the generation of long-lasting effector/memory Th17 cells. Th17 cells can disrupt the blood-brain barrier, infiltrate the central nervous system, and, along with other cytokines and microglia, lead to neuroprogression through neuroinflammation in schizophrenia. PMID:24711545

  10. Mechanism of allopurinol-mediated increase in enzyme activity in man

    PubMed Central

    Beardmore, Thomas D.; Cashman, Jay S.; Kelley, William N.

    1972-01-01

    Allopurinol therapy in man interferes with pyrimidine biosynthesis de novo by inhibition of one or both of the two enzymes, orotate phosphoribosyltransferase (OPRT) and orotidylic decarboxylase (ODC), responsible for the conversion of orotic acid to uridine-5′-monophosphate. Inhibition of this pathway in vivo is followed in 1-3 wk by an increase in the activity of both of these enzymes in erythrocytes and of ODC in circulating leukocytes. This drug-mediated increase in enzyme activity in erythrocytes could not be attributed to enzyme stabilization or induction in vivo but appeared to be due to enzyme “activation.” “Activation” of the OPRT enzyme was directly demonstrated in erythrocytes studied in vitro after incubation with oxipurinol, and to a lesser extent, with allopurinol. No evidence for “activation” of the ODC enzyme was demonstrated in vitro. This response to allopurinol therapy provides an excellent model for examining the mechanism of increased enzyme activity in response to drug administration. PMID:5032526

  11. Cellular mechanisms of the 5-HT7 receptor-mediated signaling

    PubMed Central

    Guseva, Daria; Wirth, Alexander; Ponimaskin, Evgeni

    2014-01-01

    Serotonin (5-hydroxytryptamine or 5-HT) is an important neurotransmitter regulating a wide range of physiological and pathological functions via activation of heterogeneously expressed 5-HT receptors. The 5-HT7 receptor is one of the most recently described members of the 5-HT receptor family. Functionally, 5-HT7 receptor is associated with a number of physiological and pathological responses, including serotonin-induced phase shifting of the circadian rhythm, control of memory as well as locomotor and exploratory activity. A large body of evidence indicates involvement of the 5-HT7 receptor in anxiety and depression, and recent studies suggest that 5-HT7 receptor can be highly relevant for the treatment of major depressive disorders. The 5-HT7 receptor is coupled to the stimulatory Gs-protein, and receptor stimulation results in activation of adenylyl cyclase (AC) leading to a rise of cAMP concentration. In addition, this receptor is coupled to the G12-protein to activate small GTPases of the Rho family. This review focuses on molecular mechanisms responsible for the 5-HT7 receptor-mediated signaling. We provide detailed overview of signaling cascades controlled and regulated by the 5-HT7 receptor and discuss the functional impact of 5-HT7 receptor for the regulation of different cellular and subcellular processes. PMID:25324743

  12. Development of a tailorable and tunable mechanism for cell-responsive substrate-mediated gene delivery

    NASA Astrophysics Data System (ADS)

    Blocker, Kory M.

    Due to the spatial and temporal control as well as the cell-type specificity necessary to extend gene delivery to therapeutic applications, there exists a need to create systems capable of gene transfer that are well-understood and easily manipulated. Furthermore, the creation of such materials will enable further exploration of the correlation between biochemical cues and the resulting cellular responses. In response to this as yet unmet need, a method to promote cell-responsive substrate-mediated gene delivery was developed for this dissertation. Through the use of non-viral gene delivery, flexibility of the vehicle design was incorporated into the system. Using PNA technology, pDNA was able to be specifically tethered to a self-assembled monolayer via an enzymatically-labile peptide tether. This construct was shown to promote cell-responsive delivery while retaining flexibility over the chemical and physical properties of the vehicle and substrate. By alteration of some design parameters including tether number, pDNA surface coverage, and complexation agent, temporal control over the release profile was demonstrated. Furthermore, the ability to extend the applicability of the system was detailed by transitioning to a poly-D-lysine coated substrate upon which the pDNA is immobilized. This dissertation details proof-of-principle work in the formation of a controlled release gene delivery mechanism that may be used to promote understanding of cellular responses to biochemical signaling as well as be extended to use in tissue engineering applications.

  13. Mechanisms Mediating Environmental Chemical-Induced Endocrine Disruption in the Adrenal Gland

    PubMed Central

    Martinez-Arguelles, Daniel B.; Papadopoulos, Vassilios

    2015-01-01

    Humans are continuously exposed to hundreds of man-made chemicals that pollute the environment in addition to multiple therapeutic drug treatments administered throughout life. Some of these chemicals, known as endocrine disruptors (EDs), mimic endogenous signals, thereby altering gene expression, influencing development, and promoting disease. Although EDs are eventually removed from the market or replaced with safer alternatives, new evidence suggests that early-life exposure leaves a fingerprint on the epigenome, which may increase the risk of disease later in life. Epigenetic changes occurring in early life in response to environmental toxicants have been shown to affect behavior, increase cancer risk, and modify the physiology of the cardiovascular system. Thus, exposure to an ED or combination of EDs may represent a first hit to the epigenome. Only limited information is available regarding the effect of ED exposure on adrenal function. The adrenal gland controls the stress response, blood pressure, and electrolyte homeostasis. This endocrine organ therefore has an important role in physiology and is a sensitive target of EDs. We review herein the effect of ED exposure on the adrenal gland with particular focus on in utero exposure to the plasticizer di(2-ethylehyl) phthalate. We discuss the challenges associated with identifying the mechanism mediating the epigenetic origins of disease and availability of biomarkers that may identify individual or population risks. PMID:25788893

  14. A novel mechanism for host-mediated photoprotection in endosymbiotic foraminifera.

    PubMed

    Petrou, Katherina; Ralph, Peter J; Nielsen, Daniel A

    2017-02-01

    Light underpins the health and function of coral reef ecosystems, where symbiotic partnerships with photosynthetic algae constitute the life support system of the reef. Decades of research have given us detailed knowledge of the photoprotective capacity of phototrophic organisms, yet little is known about the role of the host in providing photoprotection in symbiotic systems. Here we show that the intracellular symbionts within the large photosymbiotic foraminifera Marginopora vertebralis exhibit phototactic behaviour, and that the phototactic movement of the symbionts is accomplished by the host, through rapid actin-mediated relocation of the symbionts deeper into the cavities within the calcium carbonate test. Using a photosynthetic inhibitor, we identified that the infochemical signalling for host regulation is photosynthetically derived, highlighting the presence of an intimate communication between the symbiont and the host. Our results emphasise the central importance of the host in photosymbiotic photoprotection via a new mechanism in foraminifera that can serve as a platform for exploring host-symbiont communication in other photosymbiotic organisms.

  15. Cellular mechanisms of the 5-HT7 receptor-mediated signaling.

    PubMed

    Guseva, Daria; Wirth, Alexander; Ponimaskin, Evgeni

    2014-01-01

    Serotonin (5-hydroxytryptamine or 5-HT) is an important neurotransmitter regulating a wide range of physiological and pathological functions via activation of heterogeneously expressed 5-HT receptors. The 5-HT7 receptor is one of the most recently described members of the 5-HT receptor family. Functionally, 5-HT7 receptor is associated with a number of physiological and pathological responses, including serotonin-induced phase shifting of the circadian rhythm, control of memory as well as locomotor and exploratory activity. A large body of evidence indicates involvement of the 5-HT7 receptor in anxiety and depression, and recent studies suggest that 5-HT7 receptor can be highly relevant for the treatment of major depressive disorders. The 5-HT7 receptor is coupled to the stimulatory Gs-protein, and receptor stimulation results in activation of adenylyl cyclase (AC) leading to a rise of cAMP concentration. In addition, this receptor is coupled to the G12-protein to activate small GTPases of the Rho family. This review focuses on molecular mechanisms responsible for the 5-HT7 receptor-mediated signaling. We provide detailed overview of signaling cascades controlled and regulated by the 5-HT7 receptor and discuss the functional impact of 5-HT7 receptor for the regulation of different cellular and subcellular processes.

  16. Neural correlates of hyperalgesia in the monosodium iodoacetate model of osteoarthritis pain

    PubMed Central

    Abaei, Maryam; Sagar, Devi R; Stockley, Elizabeth G; Spicer, Clare H; Prior, Malcolm; Auer, Dorothee P

    2016-01-01

    Background The mechanisms driving osteoarthritic pain remain poorly understood, but there is increasing evidence for a role of the central nervous system in the chronification of pain. We used functional magnetic resonance imaging to investigate the influence of a model of unilateral knee osteoarthritis on nociceptive processing. Results Four to five weeks post intra-articular injection of monosodium iodoacetate (MIA, 1 mg) into the left knee, Sprague Dawley rats were anesthetized for functional magnetic resonance imaging studies to characterize the neural response to a noxious stimulus (intra-articular capsaicin injection). In a two-arm cross-over design, 5 µM/50 µl capsaicin was injected into either the left knee (n = 8, CAPS-MIA) or right control knee (n = 8, CAPS-CON), preceded by contralateral vehicle (SAL) injection. To assess neural correlates of mechanical hyperalgesia, hindpaws were stimulated with von Frey hairs (8 g: MIA; 15 g: control knee, based on behavioral withdrawal responses). The CAPS-MIA group exhibited significant activation of the periaqueductal gray, unilateral thalamus and bilateral mensencephalon, superior-colliculus, and hippocampus, with no significant activation in the other groups/conditions. Capsaicin injection increased functional connectivity in the mid-brain network and mediodorsal thalamic nucleus, hippocampus, and globus pallidus, which was significantly stronger in CAPS-MIA compared to CAPS-CON groups. Mechanical stimulation of the hyperalgesic (ipsilateral to MIA knee) and normalgesic (contralateral) hindpaws evoked qualitatively different brain activation with more widespread brainstem and anterior cingulate (ACC) activation when stimulating the hyperalgesic paw, and clearer frontal sensory activation from the normalgesic paw. Conclusions We provide evidence for modulation of nociceptive processing in a chronic knee osteoarthritis pain model with stronger brain activation and alteration of brain networks

  17. Simultaneous Inhibition of PGE2 and PGI2 Signals Is Necessary to Suppress Hyperalgesia in Rat Inflammatory Pain Models

    PubMed Central

    Kuwabara, Harumi

    2016-01-01

    Prostaglandin E2 (PGE2) is well known as a mediator of inflammatory symptoms such as fever, arthritis, and inflammatory pain. In the present study, we evaluated the analgesic effect of our selective PGE2 synthesis inhibitor, compound I, 2-methyl-2-[cis-4-([1-(6-methyl-3-phenylquinolin-2-yl)piperidin-4-yl]carbonyl amino)cyclohexyl] propanoic acid, in rat yeast-induced acute and adjuvant-induced chronic inflammatory pain models. Although this compound suppressed the synthesis of PGE2 selectively, no analgesic effect was shown in both inflammatory pain models. Prostacyclin (PGI2) also plays crucial roles in inflammatory pain, so we evaluated the involvement of PGI2 signaling in rat inflammatory pain models using prostacyclin receptor (IP) antagonist, RO3244019. RO3244019 showed no analgesic effect in inflammatory pain models, but concomitant administration of compound I and RO3244019 showed analgesic effects comparable to celecoxib, a specific cyclooxygenase- (COX-) 2 inhibitor. Furthermore, coadministration of PGE2 receptor 4 (EP4) antagonist, CJ-023423, and RO3244019 also showed an analgesic effect. These findings suggest that both PGE2 signaling, especially through the EP4 receptor, and PGI2 signaling play critical roles in inflammatory pain and concurrent inhibition of both signals is important for suppression of inflammatory hyperalgesia. PMID:27478311

  18. The Causal Mediation Formula - A Guide to the Assessment of Pathways and Mechanisms

    DTIC Science & Technology

    2011-10-01

    few decades the analysis of mediation has been dominated by linear regression paradigms, most notably the one advanced by Baron and Kenny (1986...mediation (often associated with Baron and Kenny ) has two major drawbacks. One (mentioned above) is its reliance on the untested assumption of...of controlling for mediator-output con- founders, the point was not mentioned in the influential paper of ( Baron & Kenny , 1986) and, as a result, it

  19. trans-10,cis-12 CLA promotes osteoblastogenesis via SMAD mediated mechanism in bone marrow mesenchymal stem cells.

    PubMed

    Kim, Jonggun; Park, Yooheon; Park, Yeonhwa

    2014-05-01

    The inverse relationship between osteoblast and adipocyte differentiation in bone marrow mesenchymal stem cells has been linked to overall bone mass. It has previously been reported that conjugated linoleic acid (CLA) inhibits adipogenesis via a peroxisome-proliferator activated receptor-γ (PPARγ) mediated mechanism, while it increases osteoblastogenesis via a PPARγ-independent mechanism in mesenchymal stem cells. This suggests potential implication of CLA on improving bone mass. Thus the purpose of this study was to determine involvement of CLA on regulation of osteoblastogenesis in murine mesenchymal stem cells by focusing on the Mothers against decapentaplegic (MAD)-related family of molecules 8 (SMAD8), one of key regulators of osteoblastogenesis. The trans-10,cis-12 CLA, but not the cis-9,trans-11, significantly increased osteoblastogenesis via SMAD8, and inhibited adipogenesis independent of SMAD8, while inhibiting factors regulating osteoclastogenesis in this model. These suggest that CLA may help improve osteoblastogenesis via a SMAD8 mediated mechanism.

  20. trans-10,cis-12 CLA promotes osteoblastogenesis via SMAD mediated mechanism in bone marrow mesenchymal stem cells

    PubMed Central

    Kim, Jonggun; Park, Yooheon; Park, Yeonhwa

    2014-01-01

    The inverse relationship between osteoblast and adipocyte differentiation in bone marrow mesenchymal stem cells has been linked to overall bone mass. It has previously been reported that conjugated linoleic acid (CLA) inhibits adipogenesis via a peroxisome-proliferator activated receptor-γ (PPARγ) mediated mechanism, while it increases osteoblastogenesis via a PPARγ-independent mechanism in mesenchymal stem cells. This suggests potential implication of CLA on improving bone mass. Thus the purpose of this study was to determine involvement of CLA on regulation of osteoblastogenesis in murine mesenchymal stem cells by focusing on the Mothers against decapentaplegic (MAD)-related family of molecules 8 (SMAD8), one of key regulators of osteoblastogenesis. The trans-10,cis-12 CLA, but not the cis-9,trans-11, significantly increased osteoblastogenesis via SMAD8, and inhibited adipogenesis independent of SMAD8, while inhibiting factors regulating osteoclastogenesis in this model. These suggest that CLA may help improve osteoblastogenesis via a SMAD8 mediated mechanism. PMID:25035711

  1. Feline immunodeficiency virus envelope glycoprotein mediates apoptosis in activated PBMC by a mechanism dependent on gp41 function

    SciTech Connect

    Garg, Himanshu; Joshi, Anjali; Tompkins, Wayne A. . E-mail: Wayne_Tompkins@ncsu.edu

    2004-12-20

    Feline Immunodeficiency Virus (FIV) is a lentivirus that causes immunodeficiency in cats, which parallels HIV-1-induced immunodeficiency in humans. It has been established that HIV envelope (Env) glycoprotein mediates T cell loss via a mechanism that requires CXCR4 binding. The Env glycoprotein of FIV, similar to HIV, requires CXCR4 binding for viral entry, as well as inducing membrane fusion leading to syncytia formation. However, the role of FIV Env in T cell loss and the molecular mechanisms governing this process have not been elucidated. We studied the role of Env glycoprotein in FIV-mediated T cell apoptosis in an in vitro model. Our studies demonstrate that membrane-expressed FIV Env induces apoptosis in activated feline peripheral blood mononuclear cells (PBMC) by a mechanism that requires CXCR4 binding, as the process was inhibited by CXCR4 antagonist AMD3100 in a dose-dependent manner. Interestingly, studies regarding the role of CD134, the recently identified primary receptor of FIV, suggest that binding to CD134 may not be important for induction of apoptosis in PBMC. However, inhibiting Env-mediated fusion post CXCR4 binding by FIV gp41-specific fusion inhibitor also inhibited apoptosis. Under similar conditions, a fusion-defective gp41 mutant was unable to induce apoptosis in activated PBMC. Our findings are the first report suggesting the potential of FIV Env to mediate apoptosis in bystander cells by a process that is dependent on gp41 function.

  2. Analgesic action of suspended moxibustion in rats with chronic visceral hyperalgesia correlates with enkephalins in the spinal cord☆

    PubMed Central

    Yi, Tao; Qi, Li; Wu, Huangan; Ma, Xiaopeng; Liu, Huirong; Wang, Xiaomei

    2012-01-01

    Rats that modeled chronic visceral hyperalgesia received suspended moxibustion at bilateral Tianshu (ST25) and Shangjuxu (ST37) once daily over a period of 7 days. Results show that suspended moxibustion significantly depressed abdominal withdrawal reflex scores and increased enkephalin concentration in the spinal cord. The experimental findings suggest that spinal enkephalins contributed to the analgesic effect of suspended moxibustion in rats with chronic visceral hyperalgesia. PMID:25767503

  3. Activation of GRs-Akt-nNOs-NR2B signaling pathway by second dose GR agonist contributes to exacerbated hyperalgesia in a rat model of radicular pain.

    PubMed

    Zhang, Jing; Zhang, Wei; Sun, Yu'e; Liu, Yue; Song, Lihua; Ma, Zhengliang; Gu, Xiaoping

    2014-06-01

    Central Akt, neuronal nitric oxide synthase (nNOS) and N-methyl-D-aspartate receptor subunit 2B (NR2B) play key roles in the development of neuropathic pain. Here we investigate the effects of glucocorticoid receptors (GRs) on the expression and activation of spinal Akt, nNOS and NR2B after chronic compression of dorsal root ganglia (CCD). Thermal hyperalgesia test and mechanical allodynia test were used to measure rats after intrathecal injection of GR antagonist mifepristone or GR agonist dexamethasone for 21 days postoperatively. Expression of spinal Akt, nNOS, NR2B and their phosphorylation state after CCD was examined by western blot. The effects of intrathecal treatment with dexamethasone or mifepristone on nociceptive behaviors and the corresponding expression of Akt, nNOS and NR2B in spinal cord were also investigated. Intrathecal injection of mifepristone or dexamethasone inhibited PWMT and PWTL in CCD rats. However, hyperalgesia was induced by intrathecal injection of dexamethasone on days 12 to 14 after surgery. Treatment of dexamethasone increased the expression and phosphorylation levels of spinal Akt, nNOS, GR and NR2B time dependently, whereas administration of mifepristone downregulated the expression of these proteins significantly. GRs activated spinal Akt-nNOS/NR2B pathway play important roles in the development of neuropathic pain in a time-dependent manner.

  4. Mediator mechanisms involved in TRPV1, TRPA1 and P2X receptor-mediated sensory transduction of pulmonary ROS by vagal lung C-fibers in rats.

    PubMed

    Lin, Yu-Jung; Hsu, Hsao-Hsun; Ruan, Ting; Kou, Yu Ru

    2013-10-01

    We investigated the mediator mechanisms involved in the sensory transduction of pulmonary reactive oxygen species (ROS) by vagal lung C-fibers in anesthetized rats. Airway challenge of aerosolized H₂O₂ (0.4%) stimulated these afferent fibers. The H₂O₂-induced responses were reduced by a cyclooxygenase inhibitor or ATP scavengers and also attenuated by an antagonist of TRPV1, TRPA1 or P2X receptors. The suppressive effect of the cyclooxygenase inhibitor was not affected by a combined treatment with the TRPV1 or TRPA1 antagonist, but was amplified by a combined treatment with the P2X antagonists. The suppressive effect of ATP scavengers was not affected by a combined treatment with the P2X antagonist, but was amplified by a combined treatment with the TRPV1 or TRPA1 antagonist. Thus, the actions of cyclooxygenase metabolites are mediated through the functioning of the TRPV1 and TRPA1 receptors, whereas the action of ATP is mediated through the functioning of P2X receptors.

  5. CDI Systems Are Stably Maintained by a Cell-Contact Mediated Surveillance Mechanism

    PubMed Central

    Chen, Annette J.; Leung, Nicole Y.; Hayes, Christopher S.; Low, David A.

    2016-01-01

    Contact-dependent growth inhibition (CDI) systems are widespread amongst Gram-negative bacteria where they play important roles in inter-cellular competition and biofilm formation. CDI+ bacteria use cell-surface CdiA proteins to bind neighboring bacteria and deliver C-terminal toxin domains. CDI+ cells also express CdiI immunity proteins that specifically neutralize toxins delivered from adjacent siblings. Genomic analyses indicate that cdi loci are commonly found on plasmids and genomic islands, suggesting that these Type 5 secretion systems are spread through horizontal gene transfer. Here, we examine whether CDI toxin and immunity activities serve to stabilize mobile genetic elements using a minimal F plasmid that fails to partition properly during cell division. This F plasmid is lost from Escherichia coli populations within 50 cell generations, but is maintained in ~60% of the cells after 100 generations when the plasmid carries the cdi gene cluster from E. coli strain EC93. By contrast, the ccdAB "plasmid addiction" module normally found on F exerts only a modest stabilizing effect. cdi-dependent plasmid stabilization requires the BamA receptor for CdiA, suggesting that plasmid-free daughter cells are inhibited by siblings that retain the CDI+ plasmid. In support of this model, the CDI+ F plasmid is lost rapidly from cells that carry an additional cdiI immunity gene on a separate plasmid. These results indicate that plasmid stabilization occurs through elimination of non-immune cells arising in the population via plasmid loss. Thus, genetic stabilization reflects a strong selection for immunity to CDI. After long-term passage for more than 300 generations, CDI+ plasmids acquire mutations that increase copy number and result in 100% carriage in the population. Together, these results show that CDI stabilizes genetic elements through a toxin-mediated surveillance mechanism in which cells that lose the CDI system are detected and eliminated by their siblings

  6. Mechanisms underlying the postexercise baroreceptor‐mediated suppression of heat loss

    PubMed Central

    McGinn, Ryan; Paull, Gabrielle; Meade, Robert D.; Fujii, Naoto; Kenny, Glen P.

    2014-01-01

    Abstract Reports indicate that postexercise heat loss is modulated by baroreceptor input; however, the mechanisms remain unknown. We examined the time‐dependent involvement of adenosine receptors, noradrenergic transmitters, and nitric oxide (NO) in modulating baroreceptor‐mediated changes in postexercise heat loss. Eight males performed two 15‐min cycling bouts (85% VO2max) each followed by a 45‐min recovery in the heat (35°C). Lower body positive (LBPP), negative (LBNP), or no (Control) pressure were applied in three separate sessions during the final 30‐min of each recovery. Four microdialysis fibres in the forearm skin were perfused with: (1) lactated Ringer's (Ringer's); (2) 4 mmol·L−1 Theophylline (inhibits adenosine receptors); (3) 10 mmol·L−1 Bretylium (inhibits noradrenergic transmitter release); or (4) 10 mmol·L−1 l‐NAME (inhibits NO synthase). We measured cutaneous vascular conductance (CVC; percentage of maximum) calculated as perfusion units divided by mean arterial pressure, and local sweat rate. Compared to Control, LBPP did not influence CVC at l‐NAME, Theophylline or Bretylium during either recovery (P >0.07); however, CVC at Ringer's was increased by ~5‐8% throughout 30 min of LBPP during Recovery 1 (all P <0.02). In fact, CVC at Ringer's was similar to Theophylline and Bretylium during LBPP. Conversely, LBNP reduced CVC at all microdialysis sites by ~7–10% in the last 15 min of Recovery 2 (all P <0.05). Local sweat rate was similar at all treatment sites as a function of pressure condition (P >0.10). We show that baroreceptor input modulates postexercise CVC to some extent via adenosine receptors, noradrenergic vasoconstriction, and NO whereas no influence was observed for postexercise sweating. PMID:25293599

  7. A mechanism underlying NOTCH-induced and ubiquitin-mediated JAK3 degradation.

    PubMed

    Wu, Wei; Sun, Xiao-Hong

    2011-12-02

    Although NOTCH signaling is well known to regulate lymphopoiesis, Janus kinase 3 (JAK3) also plays a critical role in promoting lymphocyte development. We have previously found that NOTCH signaling leads to the degradation of JAK3 in B lineage cells, suggesting that NOTCH signaling exerts its biological effect on lymphopoiesis through modulating JAK3 levels. Here, we delineate the biochemical mechanisms involved in NOTCH-induced JAK3 ubiquitination and degradation. NOTCH signaling is known to transcriptionally activate the genes encoding ASB2 (ankyrin-repeat SOCS box containing protein 2) and SKP2 (S-phase kinase-associated protein 2). We show that not only NOTCH but also ASB2 and SKP2 can promote the ubiquitination and degradation of JAK3. Both ASB2 and SKP2 can interact with JAK3 through different domains; the FERM and pseudo-kinase domains each had high affinities for ASB2, whereas the kinase domain primarily associated with SKP2. ASB2 and SKP2 previously have been shown to associate with each other to bridge the formation of a non-canonical Cullin1 and Cullin5-containing dimeric E3 ligase complex. Interestingly, the R980W mutant of JAK3 exhibited diminished interaction with SKP2 and resistance to NOTCH or ASB2-induced degradation. Furthermore, dominant-negative mutants of either Cullin1 or Cullin5, which lack the C terminus responsible for recruiting the E2 enzymes, were able to prevent JAK3 degradation induced by both ASB2/SKP2 and NOTCH signaling. Together, these results suggest that JAK3 ubiquitination involves the non-canonical dimeric E3 ligase complex, and the R980W mutant will serve as an excellent tool for investigating the biological significance of NOTCH-mediated JAK3 turnover.

  8. Persistent polar depletion of stratospheric ozone and emergent mechanisms of ultraviolet radiation-mediated health dysregulation.

    PubMed

    Dugo, Mark A; Han, Fengxiang; Tchounwou, Paul B

    2012-01-01

    Year 2011 noted the first definable ozone "hole" in the Arctic region, serving as an indicator to the continued threat of dangerous ultraviolet radiation (UVR) exposure caused by the deterioration of stratospheric ozone in the northern hemisphere. Despite mandates of the Montreal Protocol to phase out the production of ozone-depleting chemicals (ODCs), the relative stability of ODCs validates popular notions of persistent stratospheric ozone for several decades. Moreover, increased UVR exposure through stratospheric ozone depletion is occurring within a larger context of physiologic stress and climate change across the biosphere. In this review, we provide commentaries on stratospheric ozone depletion with relative comparisons between the well-known Antarctic ozone hole and the newly defined ozone hole in the Arctic. Compared with the Antarctic region, the increased UVR exposure in the Northern Hemisphere poses a threat to denser human populations across North America, Europe, and Asia. In this context, we discuss emerging targets of UVR exposure that can potentially offset normal biologic rhythms in terms of taxonomically conserved photoperiod-dependent seasonal signaling and entrainment of circadian clocks. Consequences of seasonal shifts during critical life history stages can alter fitness and condition, whereas circadian disruption is increasingly becoming associated as a causal link to increased carcinogenesis. We further review the significance of genomic alterations via UVR-induced modulations of phase I and II transcription factors located in skin cells, the aryl hydrocarbon receptor (AhR), and the nuclear factor (erythroid-derived 2)-related factor 2 (Nrf2), with emphasis on mechanism that can lead to metabolic shifts and cancer. Although concern for adverse health consequences due to increased UVR exposure are longstanding, recent advances in biochemical research suggest that AhR and Nrf2 transcriptional regulators are likely targets for UVR-mediated

  9. Functionalized fullerenes mediate photodynamic killing of cancer cells: Type I versus Type II photochemical mechanism

    PubMed Central

    Mroz, Pawel; Pawlak, Anna; Satti, Minahil; Lee, Haeryeon; Wharton, Tim; Gali, Hariprasad; Sarna, Tadeusz; Hamblin, Michael R.

    2007-01-01

    Photodynamic therapy (PDT) employs the combination of non-toxic photosensitizers (PS) and harmless visible light to generate reactive oxygen species (ROS) and kill cells. Most clinically studied PS are based on the tetrapyrrole structure of porphyrins, chlorins and related molecules, but new non-tetrapyrrole PS are being sought. Fullerenes are soccer-ball shaped molecules composed of sixty or seventy carbon atoms and have attracted interest in connection with the search for biomedical applications of nanotechnology. Fullerenes are biologically inert unless derivatized with functional groups, whereupon they become soluble and can act as PS. We have compared the photodynamic activity of six functionalized fullerenes with 1, 2, or 3 hydrophilic or 1, 2, or 3 cationic groups. The octanol-water partition coefficients were determined and the relative contributions of Type I photochemistry (photogeneration of superoxide in the presence of NADH) and Type II photochemistry (photogeneration of singlet oxygen) were studied by measurement of oxygen consumption, 1270-nm luminescence and EPR spin-trapping of the superoxide product. We studied three mouse cancer cell lines: (J774, LLC and CT26) incubated for 24 h with fullerenes and illuminated with white light. The order of effectiveness as PS was inversely proportional to the degree of substitution of the fullerene nucleus for both the neutral and cationic series. The mono-pyrrolidinium fullerene was the most active PS against all cell lines and induced apoptosis 4–6 hours after illumination. It produced diffuse intracellular fluorescence when dichlorodihydrofluorescein was added as an ROS probe suggesting a Type I mechanism for phototoxicity. We conclude that certain functionalized fullerenes have potential as novel PDT agents and phototoxicity may be mediated both by superoxide and by singlet oxygen. PMID:17664135

  10. Ligand-specific transcriptional mechanisms underlie aryl hydrocarbon receptor-mediated developmental toxicity of oxygenated PAHs

    DOE PAGES

    Goodale, B. C.; Geisel School of Medicine at Dartmouth, Hanover, NH; La Du, J.; ...

    2015-07-03

    Polycyclic aromatic hydrocarbons (PAHs) are priority environmental contaminants that exhibit mutagenic, carcinogenic, proinflammatory, and teratogenic properties. Oxygen-substituted PAHs (OPAHs) are formed during combustion processes and via phototoxidation and biological degradation of parent (unsubstituted) PAHs. Despite their prevalence both in contaminated industrial sites and in urban air, OPAH mechanisms of action in biological systems are relatively understudied. Like parent PAHs, OPAHs exert structure-dependent mutagenic activities and activation of the aryl hydrocarbon receptor (AHR) and cytochrome p450 metabolic pathway. Four-ring OPAHs 1,9-benz-10-anthrone (BEZO) and benz(a)anthracene-7,12-dione (7,12-B[a]AQ) cause morphological aberrations and induce markers of oxidative stress in developing zebrafish with similar potency, butmore » only 7,12-B[a]AQ induces robust Cyp1a protein expression. We investigated the role of the AHR in mediating the toxicity of BEZO and 7,12-B[a]AQ, and found that knockdown of AHR2 rescued developmental effects caused by both compounds. Using RNA-seq and molecular docking, we identified transcriptional responses that precede developmental toxicity induced via differential interaction with AHR2. Redox-homeostasis genes were affected similarly by these OPAHs, while 7,12-B[a]AQ preferentially activated phase 1 metabolism and BEZO uniquely decreased visual system genes. Analysis of biological functions and upstream regulators suggests that BEZO is a weak AHR agonist, but interacts with other transcriptional regulators to cause developmental toxicity in an AHR-dependent manner. Furthermore, identifying ligand-dependent AHR interactions and signaling pathways is essential for understanding toxicity of this class of environmentally relevant compounds.« less

  11. Ligand-specific transcriptional mechanisms underlie aryl hydrocarbon receptor-mediated developmental toxicity of oxygenated PAHs

    SciTech Connect

    Goodale, B. C.; La Du, J.; Tilton, S. C.; Sullivan, C. M.; Bisson, W. H.; Waters, K. M.; Tanguay, R. L.

    2015-07-03

    Polycyclic aromatic hydrocarbons (PAHs) are priority environmental contaminants that exhibit mutagenic, carcinogenic, proinflammatory, and teratogenic properties. Oxygen-substituted PAHs (OPAHs) are formed during combustion processes and via phototoxidation and biological degradation of parent (unsubstituted) PAHs. Despite their prevalence both in contaminated industrial sites and in urban air, OPAH mechanisms of action in biological systems are relatively understudied. Like parent PAHs, OPAHs exert structure-dependent mutagenic activities and activation of the aryl hydrocarbon receptor (AHR) and cytochrome p450 metabolic pathway. Four-ring OPAHs 1,9-benz-10-anthrone (BEZO) and benz(a)anthracene-7,12-dione (7,12-B[a]AQ) cause morphological aberrations and induce markers of oxidative stress in developing zebrafish with similar potency, but only 7,12-B[a]AQ induces robust Cyp1a protein expression. We investigated the role of the AHR in mediating the toxicity of BEZO and 7,12-B[a]AQ, and found that knockdown of AHR2 rescued developmental effects caused by both compounds. Using RNA-seq and molecular docking, we identified transcriptional responses that precede developmental toxicity induced via differential interaction with AHR2. Redox-homeostasis genes were affected similarly by these OPAHs, while 7,12-B[a]AQ preferentially activated phase 1 metabolism and BEZO uniquely decreased visual system genes. Analysis of biological functions and upstream regulators suggests that BEZO is a weak AHR agonist, but interacts with other transcriptional regulators to cause developmental toxicity in an AHR-dependent manner. Furthermore, identifying ligand-dependent AHR interactions and signaling pathways is essential for understanding toxicity of this class of environmentally relevant compounds.

  12. Ciprofloxacin mediates cancer stem cell phenotypes in lung cancer cells through caveolin-1-dependent mechanism.

    PubMed

    Phiboonchaiyanan, Preeyaporn Plaimee; Kiratipaiboon, Chayanin; Chanvorachote, Pithi

    2016-04-25

    Cancer stem cells (CSCs), a subpopulation of cancer cells with high aggressive behaviors, have been identified in many types of cancer including lung cancer as one of the key mediators driving cancer progression and metastasis. Here, we have reported for the first time that ciprofloxacin (CIP), a widely used anti-microbial drug, has a potentiating effect on CSC-like features in human non-small cell lung cancer (NSCLC) cells. CIP treatment promoted CSC-like phenotypes, including enhanced anchorage-independent growth and spheroid formation. The known lung CSC markers: CD133, CD44, ABCG2 and ALDH1A1 were found to be significantly increased, while the factors involving in epithelial to mesenchymal transition (EMT): Slug and Snail, were depleted. Also, self-renewal transcription factors Oct-4 and Nanog were found to be up-regulated in CIP-treated cells. The treatment of CIP on CSC-rich populations obtained from secondary spheroids resulted in the further increase of CSC markers. In addition, we have proven that the mechanistic insight of the CIP induced stemness is through Caveolin-1 (Cav-1)-dependent mechanism. The specific suppression of Cav-1 by stably transfected Cav-1 shRNA plasmid dramatically reduced the effect of CIP on CSC markers as well as the CIP-induced spheroid formation ability. Cav-1 was shown to activate protein kinase B (Akt) and extracellular signal-regulated kinase (ERK) pathways in CSC-rich population; however, such an effect was rarely found in the main lung cancer cells population. These findings reveal a novel effect of CIP in positively regulating CSCs in lung cancer cells via the activation of Cav-1, Akt and ERK, and may provoke the awareness of appropriate therapeutic strategy in cancer patients.

  13. Chronic elevation of IL-1β induces diuresis via a cyclooxygenase 2-mediated mechanism.

    PubMed

    Boesen, E I

    2013-07-15

    Chronic renal inflammation is an increasingly recognized phenomenon in multiple disease states, but the impact of specific cytokines on renal function is unclear. Previously, we found that 14-day interleukin-1β (IL-1β) infusion increased urine flow in mice. To determine the mechanism by which this occurs, the current study tested the possible involvement of three classical prodiuretic pathways. Chronic IL-1β infusion significantly increased urine flow (6.5 ± 1 ml/day at day 14 vs. 2.3 ± 0.3 ml/day in vehicle group; P < 0.05) and expression of cyclooxygenase (COX)-2, all three nitric oxide synthase (NOS) isoforms, and endothelin (ET)-1 in the kidney (P < 0.05 in all cases). Urinary prostaglandin E metabolite (PGEM) excretion was also significantly increased at day 14 of IL-1β infusion (1.21 ± 0.26 vs. 0.29 ± 0.06 ng/day in vehicle-infused mice; P = 0.001). The selective COX-2 inhibitor celecoxib markedly attenuated urinary PGEM excretion and abolished the diuretic response to chronic IL-1β infusion. In contrast, deletion of NOS3, or inhibition of NOS1 with L-VNIO, did not blunt the diuretic effect of IL-1β, nor did pharmacological blockade of endothelin ETA and ETB receptors with A-182086. Consistent with a primary effect on water transport, IL-1β infusion markedly reduced inner medullary aquaporin-2 expression (P < 0.05) and did not alter urinary Na⁺ or K⁺ excretion. These data indicate a critical role for COX-2 in mediating the effects of chronic IL-1β elevation on the kidney.

  14. Ligand-Specific Transcriptional Mechanisms Underlie Aryl Hydrocarbon Receptor-Mediated Developmental Toxicity of Oxygenated PAHs

    PubMed Central

    Goodale, B. C.; La Du, J.; Tilton, S. C.; Sullivan, C. M.; Bisson, W. H.; Waters, K. M.; Tanguay, R. L.

    2015-01-01

    Polycyclic aromatic hydrocarbons (PAHs) are priority environmental contaminants that exhibit mutagenic, carcinogenic, proinflammatory, and teratogenic properties. Oxygen-substituted PAHs (OPAHs) are formed during combustion processes and via phototoxidation and biological degradation of parent (unsubstituted) PAHs. Despite their prevalence both in contaminated industrial sites and in urban air, OPAH mechanisms of action in biological systems are relatively understudied. Like parent PAHs, OPAHs exert structure-dependent mutagenic activities and activation of the aryl hydrocarbon receptor (AHR) and cytochrome p450 metabolic pathway. Four-ring OPAHs 1,9-benz-10-anthrone (BEZO) and benz(a)anthracene-7,12-dione (7,12-B[a]AQ) cause morphological aberrations and induce markers of oxidative stress in developing zebrafish with similar potency, but only 7,12-B[a]AQ induces robust Cyp1a protein expression. We investigated the role of the AHR in mediating the toxicity of BEZO and 7,12-B[a]AQ, and found that knockdown of AHR2 rescued developmental effects caused by both compounds. Using RNA-seq and molecular docking, we identified transcriptional responses that precede developmental toxicity induced via differential interaction with AHR2. Redox-homeostasis genes were affected similarly by these OPAHs, while 7,12-B[a]AQ preferentially activated phase 1 metabolism and BEZO uniquely decreased visual system genes. Analysis of biological functions and upstream regulators suggests that BEZO is a weak AHR agonist, but interacts with other transcriptional regulators to cause developmental toxicity in an AHR-dependent manner. Identifying ligand-dependent AHR interactions and signaling pathways is essential for understanding toxicity of this class of environmentally relevant compounds. PMID:26141390

  15. Molecular mechanisms of AGE/RAGE-mediated fibrosis in the diabetic heart

    PubMed Central

    Zhao, Jia; Randive, Rushil; Stewart, James A

    2014-01-01

    Chronic hyperglycemia is one of the main characteristics of diabetes. Persistent exposure to elevated glucose levels has been recognized as one of the major causal factors of diabetic complications. In pathologies, like type 2 diabetes mellitus (T2DM), mechanical and biochemical stimuli activate profibrotic signaling cascades resulting in myocardial fibrosis and subsequent impaired cardiac performance due to ventricular stiffness. High levels of glucose nonenzymatically react with long-lived proteins, such as collagen, to form advanced glycation end products (AGEs). AGE-modified collagen increase matrix stiffness making it resistant to hydrolytic turnover, resulting in an accumulation of extracellular matrix (ECM) proteins. AGEs account for many of the diabetic cardiovascular complications through their engagement of the receptor for AGE (RAGE). AGE/RAGE activation stimulates the secretion of numerous profibrotic growth factors, promotes increased collagen deposition leading to tissue fibrosis, as well as increased RAGE expression. To date, the AGE/RAGE cascade is not fully understood. In this review, we will discuss one of the major fibrotic signaling pathways, the AGE/RAGE signaling cascade, as well as propose an alternate pathway via Rap1a that may offer insight into cardiovascular ECM remodeling in T2DM. In a series of studies, we demonstrate a role for Rap1a in the regulation of fibrosis and myofibroblast differentiation in isolated diabetic and non-diabetic fibroblasts. While these studies are still in a preliminary stage, inhibiting Rap1a protein expression appears to down-regulate the molecular switch used to activate the ζ isotype of protein kinase C thereby promote AGE/RAGE-mediated fibrosis. PMID:25512788

  16. Nucleation and growth mechanisms of ion etching and surfactant-mediated growth

    NASA Astrophysics Data System (ADS)

    Chan, Lana Hiului

    2001-07-01

    Nucleation and growth mechanisms of ion etching of Cu(100) and surfactant (Sb) mediated Ag growth on Ge(100) were studied in ultra-high vacuum (UHV) using scanning tunneling microscopy (STM), low energy electron diffraction (LEED), temperature programmed desorption (TPD), and low energy ion scattering spectroscopy (LEISS). The formation of vacancy clusters from ion etching was characterized and could be modeled and understood as the reverse of film growth. The results suggest that a barrier exists for vacancies to ascend steps. This additional barrier also contributes to pit ordering. The interaction of the Sb with Ge was investigated. Antimony was found to form intermixed surface with Ge. The intermixing, however, could be controlled by deposition and annealing temperatures and Sb coverage. The growth of Ag with and without Sb on Ge(100) was studied and compared. Without Sb, Ag formed elongated 2D islands and 3D clusters. A new metastable Ag-Ge surface alloy phase was found when Ag was deposited at 470 K. This alloy phase can only be obtained by deposition at 470 K. Annealing to 470 K gave a surface with 3D clusters only. When Sb was used as a surfactant for Ag growth, no surface alloy was observed and the cluster shape was isotropic instead of rectangular. On the Sb covered surface, nucleation appeared to be dominated by defects at low temperatures. At higher temperatures, Sb increased the diffusion barrier for Ag. Other effects of using Sb on Ag growth included increase in cluster density and decrease in cluster size. Antimony did not work as a conventional surfactant since it did not segregate to the top of the film at elevated temperatures.

  17. Ionic mechanisms mediating oscillatory membrane potentials in wide-field retinal amacrine cells.

    PubMed

    Vigh, Jozsef; Solessio, Eduardo; Morgans, Catherine W; Lasater, Eric M

    2003-07-01

    Particular types of amacrine cells of the vertebrate retina show oscillatory membrane potentials (OMPs) in response to light stimulation. Historically it has been thought the oscillations arose as a result of circuit properties. In a previous study we found that in some amacrine cells, the ability to oscillate was an intrinsic property of the cell. Here we characterized the ionic mechanisms responsible for the oscillations in wide-field amacrine cells (WFACs) in an effort to better understand the functional properties of the cell. The OMPs were found to be calcium (Ca2+) dependent; blocking voltage-gated Ca2+ channels eliminated the oscillations, whereas elevating extracellular Ca2+ enhanced them. Strong intracellular Ca2+ buffering (10 mM EGTA or bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid) eliminated any attenuation in the OMPs as well as a Ca2+-dependent inactivation of the voltage-gated Ca2+ channels. Pharmacological and immunohistochemical characterization revealed that WFACs express L- and N-type voltage-sensitive Ca2+ channels. Block of the L-type channels eliminated the OMPs, but omega-conotoxin GVIA did not, suggesting a different function for the N-type channels. The L-type channels in WFACs are functionally coupled to a set of calcium-dependent potassium (K(Ca)) channels to mediate OMPs. The initiation of OMPs depended on penitrem-A-sensitive (BK) K(Ca) channels, whereas their duration is under apamin-sensitive (SK) K(Ca) channel control. The Ca2+ current is essential to evoke the OMPs and triggering the K(Ca) currents, which here act as resonant currents, enhances the resonance as an amplifying current, influences the filtering characteristics of the cell membrane, and attenuates the OMPs via CDI of the L-type Ca2+ channel.

  18. Mechanisms of a novel anticancer therapeutic strategy involving atmospheric pressure plasma-mediated apoptosis and DNA strand break formation.

    PubMed

    Chung, Woo-Hyun

    2016-01-01

    Atmospheric pressure plasma has been developed for a variety of biomedical applications due to its chemically reactive components. Recently, the plasma has emerged as a promising novel cancer therapy based on its ability to selectively ablate cancer cells while leaving normal cells essentially unaffected. The therapeutic effect of plasma is attributed to intracellular generation of reactive oxygen/nitrogen species (ROS/RNS) leading to mitochondria-mediated apoptosis and to activation of the DNA damage checkpoint signaling pathway via severe DNA strand break formation. However, the biochemical mechanisms responsible for appropriate activation of these physiological events and which pathway is more crucial for plasma-mediated cytotoxicity have not been clarified. Understanding the molecular link between ROS/RNS-mediated apoptosis and DNA damage-involved chromosome instability is critical for the development of more efficacious therapeutic strategies for selective killing of diverse cancer cells.

  19. New Insights into the Mechanism and an Expanded Scope of the Fe(III)-mediated Vinblastine Coupling Reaction

    PubMed Central

    Gotoh, Hiroaki; Sears, Justin E.; Eschenmoser, Albert; Boger, Dale L.

    2012-01-01

    A definition of the scope of aromatic substrates that participate with catharanthine in an Fe(III)-mediated coupling reaction, an examination of the key structural features of catharanthine required for participation in the reaction, and the development of a generalized indole functionalization reaction that bears little structural relationship to catharanthine itself are detailed. In addition to providing insights into the mechanism of the Fe(III)-mediated coupling reaction of catharanthine with vindoline suggesting the reaction conducted in acidic aqueous buffer may be radical mediated, the studies provide new opportunities for the preparation of previously inaccessible vinblastine analogs and define powerful new methodology for the synthesis of indole-containing natural and unnatural products. PMID:22856867

  20. Mechanism of PP2A-mediated IKKβ dephosphorylation: a systems biological approach

    PubMed Central

    Witt, Johannes; Barisic, Sandra; Schumann, Eva; Allgöwer, Frank; Sawodny, Oliver; Sauter, Thomas; Kulms, Dagmar

    2009-01-01

    Background Biological effects of nuclear factor-κB (NFκB) can differ tremendously depending on the cellular context. For example, NFκB induced by interleukin-1 (IL-1) is converted from an inhibitor of death receptor induced apoptosis into a promoter of ultraviolet-B radiation (UVB)-induced apoptosis. This conversion requires prolonged NFκB activation and is facilitated by IL-1 + UVB-induced abrogation of the negative feedback loop for NFκB, involving a lack of inhibitor of κB (IκBα) protein reappearance. Permanent activation of the upstream kinase IKKβ results from UVB-induced inhibition of the catalytic subunit of Ser-Thr phosphatase PP2A (PP2Ac), leading to immediate phosphorylation and degradation of newly synthesized IκBα. Results To investigate the mechanism underlying the general PP2A-mediated tuning of IKKβ phosphorylation upon IL-1 stimulation, we have developed a strictly reduced mathematical model based on ordinary differential equations which includes the essential processes concerning the IL-1 receptor, IKKβ and PP2A. Combining experimental and modelling approaches we demonstrate that constitutively active, but not post-stimulation activated PP2A, tunes out IKKβ phosphorylation thus allowing for IκBα resynthesis in response to IL-1. Identifiability analysis and determination of confidence intervals reveal that the model allows reliable predictions regarding the dynamics of PP2A deactivation and IKKβ phosphorylation. Additionally, scenario analysis is used to scrutinize several hypotheses regarding the mode of UVB-induced PP2Ac inhibition. The model suggests that down regulation of PP2Ac activity, which results in prevention of IκBα reappearance, is not a direct UVB action but requires instrumentality. Conclusion The model developed here can be used as a reliable building block of larger NFκB models and offers comprehensive simplification potential for future modeling of NFκB signaling. It gives more insight into the newly discovered

  1. Prevention of Remifentanil Induced Postoperative Hyperalgesia by Dexmedetomidine via Regulating the Trafficking and Function of Spinal NMDA Receptors as well as PKC and CaMKII Level In Vivo and In Vitro

    PubMed Central

    Chen, Yi; Zheng, Yuxin; Xie, Ke-liang; He, Ying; Wang, Zhifen; Wang, Guo-lin; Yu, Yong-hao

    2017-01-01

    Remifentanil-induced secondary hyperalgesia has been demonstrated in both animal experiments and clinical trials. Enhancement of N-methyl-D-aspartate (NMDA) receptor trafficking as well as protein kinase C (PKC) and calmodulin-dependent protein kinase II (CaMKII) have been reported to be involved in the induction and maintenance of central sensitization. In the current study, it was demonstrated that dexmedetomidine could prevent remifentanil-induced hyperalgesia (RIH) via regulating spinal NMDAR-PKC-Ca2+/ CaMKII pathway in vivo and in vitro. We firstly investigated the effect of dexmedetomidine, a highly selective α2-adrenergic receptor agonist, on mechanical and thermal hyperalgesia using a rat model of RIH. NMDA receptor subunits (NR1, NR2A and NR2B) expression and membrane trafficking as well as PKC and CaMKII expression in spinal cord L4–L5 segments were measured by Western blot analysis. The expression of NMDA receptor subunits (NR1, NR2A and NR2B) were also detected by immunohistochemistry. Further more, the effect of dexmedetomidine on NMDA receptor current amplitude and frequency in spinal cord slices were investigated by whole-cell patch-clamp recording. We found that remifentail infusion at 1.2 μg.kg−1.min−1 for 90 min caused mechanical and thermal hyperalgesia, up-regulated NMDA receptor subunits NR1 and NR2B expression in both membrane fraction and total lysate as well as increased PKC and CaMKII expression in spinal cord dorsal horn. Subcutaneously injection of dexmedetomidine at the dose of 50 μg/kg at 30 min before plantar incision significantly attenuated remifentanil-induced mechanical and thermal hyperalgesia from 2 h to 48 h after infusion, and this was associated with reversal of up-regulated NR1 and NR2B subunits in both membrane fraction and total lysate as well as increased PKC and CaMKII expression in spinal cord dorsal horn. Furthermore, remifentanil incubation increased amplitude and frequency of NMDA receptor-induced current in

  2. Opioid-Induced Hyperalgesia - Worsening Pain in Opioid-Dependent Patients

    DTIC Science & Technology

    2013-02-01

    shown to reduce pain . Amantadine is an NMDA receptor antagonist that may mitigate central sensitization. Adjuvant analgesics may lessen nociceptive ...FEB 2013 2. REPORT TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Opioid-induced hyperalgesia--worsening pain in opioid-dependent...Report Opioid-induced hyperalgesia—worsening pain in opioid-dependent patients☆ Abstract Patients with chronic opioid use are commonly treated in the

  3. A phenomenological model for mechanically mediated growth, remodeling, damage, and plasticity of gel-derived tissue engineered blood vessels.

    PubMed

    Raykin, Julia; Rachev, Alexander I; Gleason, Rudolph L

    2009-10-01

    Mechanical stimulation has been shown to dramatically improve mechanical and functional properties of gel-derived tissue engineered blood vessels (TEBVs). Adjusting factors such as cell source, type of extracellular matrix, cross-linking, magnitude, frequency, and time course of mechanical stimuli (among many other factors) make interpretation of experimental results challenging. Interpretation of data from such multifactor experiments requires modeling. We present a modeling framework and simulations for mechanically mediated growth, remodeling, plasticity, and damage of gel-derived TEBVs that merge ideas from classical plasticity, volumetric growth, and continuum damage mechanics. Our results are compared with published data and suggest that this model framework can predict the evolution of geometry and material behavior under common experimental loading scenarios.

  4. EAR motif-mediated transcriptional repression in plants: an underlying mechanism for epigenetic regulation of gene expression.

    PubMed

    Kagale, Sateesh; Rozwadowski, Kevin

    2011-02-01

    Ethylene-responsive element binding factor-associated Amphiphilic Repression (EAR) motif-mediated transcriptional repression is emerging as one of the principal mechanisms of plant gene regulation. The EAR motif, defined by the consensus sequence patterns of either LxLxL or DLNxxP, is the most predominant form of transcriptional repression motif so far identified in plants. Additionally, this active repression motif is highly conserved in transcriptional regulators known to function as negative regulators in a broad range of developmental and physiological processes across evolutionarily diverse plant species. Recent discoveries of co-repressors interacting with EAR motifs, such as TOPLESS (TPL) and AtSAP18, have begun to unravel the mechanisms of EAR motif-mediated repression. The demonstration of genetic interaction between mutants of TPL and AtHDA19, co-complex formation between TPL-related 1 (TPR1) and AtHDA19, as well as direct physical interaction between AtSAP18 and AtHDA19 support a model where EAR repressors, via recruitment of chromatin remodeling factors, facilitate epigenetic regulation of gene expression. Here, we discuss the biological significance of EAR-mediated gene regulation in the broader context of plant biology and present literature evidence in support of a model for EAR motif-mediated repression via the recruitment and action of chromatin modifiers. Additionally, we discuss the possible influences of phosphorylation and ubiquitination on the function and turnover of EAR repressors.

  5. Molecular mechanisms of CD8+ T cell-mediated delayed hypersensitivity: implications for allergies, asthma, and autoimmunity.

    PubMed

    Kalish, R S; Askenase, P W

    1999-02-01

    Delayed-type hypersensitivity (DTH) is defined as the recruitment of T cells into tissues to be activated by antigen-presenting cells to produce cytokines that mediate local inflammation. CD8+ T cells are now known to mediate DTH responses in allergic contact dermatitis, drug eruptions, asthma, and autoimmune diseases. This inflammatory effector capability of CD8+ cytotoxic T cells was previously poorly recognized, but there is now considerable evidence that these diseases may be mediated by CD8+ DTH. The difference between CD8+ T cells and CD4+ T cells mediating DTH relates to the molecular mechanisms by which antigens are processed and presented to the T cells. Antigens external to the cell are phagocytosed and processed for presentation on MHC class II molecules (eg, HLA-DR) to CD4+ T cells. In contrast, internal cytoplasmic antigens are processed by the endogenous pathway for presentation on MHC class I molecules (eg, HLA-A, -B, and -C) to CD8+ T cells. External allergens can also enter the endogenous pathway to be presented to CD8+ T cells. These include many contact sensitizers, chemical and protein respiratory allergens, viral antigens, metabolic products of drugs, and autoantigens. The resulting CD8+ T-cell response explains the role of CD8+ T-cell DTH mechanisms in allergic contact dermatitis, asthma, drug eruptions, and autoimmune diseases.

  6. Reduced GABAergic transmission in the ventrobasal thalamus contributes to thermal hyperalgesia in chronic inflammatory pain

    PubMed Central

    Zhang, Chan; Chen, Rong-Xiang; Zhang, Yu; Wang, Jie; Liu, Feng-Yu; Cai, Jie; Liao, Fei-Fei; Xu, Fu-Qiang; Yi, Ming; Wan, You

    2017-01-01

    The ventrobasal (VB) thalamus is innervated by GABAergic afferents from the thalamic reticular nucleus (TRN) and participates in nociception. But how the TRN-VB pathway regulates pain is not fully understood. In the present study, we reported decreased extracellular GABA levels in the VB of rats with CFA-induced chronic inflammatory pain, measured by microdialysis with HPLC analysis. In vitro whole-cell patch-clamp recording showed decreased amplitudes of tonic currents, increased frequencies of mIPSCs, and increased paired-pulse ratios in thalamic slices from chronic inflammatory rats (7 days). Microinjection of the GABAAR agonist muscimol and optogenetic activation of the TRN-VB pathway relieved thermal hyperalgesia in chronic inflammatory pain. By contrast, microinjecting the extrasynaptic GABAAR agonist THIP or selective knockout of synaptic GABAAR γ2 subunits aggravated thermal hyperalgesia in the chronic stage of inflammatory pain. Our findings indicate that reduced GABAergic transmission in the VB contributes to thermal hyperalgesia in chronic inflammatory pain, which could be a synaptic target for pharmacotherapy. PMID:28150719

  7. Model of methadone-induced hyperalgesia in rats and effect of memantine.

    PubMed

    Hay, Justin L; Kaboutari, Jahangir; White, Jason M; Salem, Abdallah; Irvine, Rod

    2010-01-25

    Methadone used for opioid dependence therapy is associated with increased pain sensitivity. This study aimed to investigate whether methadone administration lowers nociceptive threshold in adult male Sprague-Dawley (SD) rats, and if this threshold could be altered by the NMDA receptor antagonist memantine. Rats were implanted with osmotic pumps delivering 1mg/kg/day methadone (n=6), or saline placebo (n=6) (0.51 microl/h). A separate cohort of rats received either methadone 1mg/kg/day (n=8) or methadone 1mg/kg/day with 20mg/kg/day memantine (n=8). Nociception was measured by the Hargreave's paw withdrawal test. Baseline nociception was measured on day 0 prior to osmotic pump implantation and was measured daily for the following 21 days. Osmotic pumps were removed following nociceptive testing on day 14. Methadone only treated rats had a mean paw withdrawal latency significantly lower than the corresponding values for saline on days 8, 9, 10, 11, 12, 14, and 17 (P<0.05). At all other time points the mean paw withdrawal latency was not significantly different from saline (P>0.05). Paw withdrawal latency of rats treated with methadone co-administered with memantine did not differ significantly compared to methadone only (P>0.05). This demonstrates that methadone induces hyperalgesia in the SD rat yet this hyperalgesia resolves following discontinuation of methadone administration. Furthermore, memantine does not alter the development of methadone-induced hyperalgesia.

  8. Reduction of spinal PGE2 concentrations prevents swim stress-induced thermal hyperalgesia.

    PubMed

    Guevara, Coram; Fernandez, Ana Cristina; Cardenas, Ricardo; Suarez-Roca, Heberto

    2015-03-30

    We evaluated the association between spinal PGE2 and thermal hyperalgesia following repeated stress. Thermal nociception was determined in male Sprague-Dawley rats using the hot-plate test, before and after forced-swimming; non-conditioned rats served as controls. Animals were pretreated with ketoprofen or meloxicam, preferential COX-1 and COX-2 inhibitors, respectively. After the second hot-plate test, we measured serum corticosterone (stress marker), and lumbar spinal PGE2 (neuroinflammation marker) under peripheral inflammation (1% formalin plantar injection). Stressed rats displayed response latencies 40% shorter and inflammatory spinal PGE2 levels 95% higher than controls. Pretreatment with ketoprofen or meloxicam prevented hyperalgesia and elevation of spinal PGE2, increasing the escape behavior time during forced swimming 95% respect to saline-treated rats. Corticosterone levels in stressed rats were 97% higher than controls; COX inhibitors reduced them by 84%. PGE2 could participate in stress-induced hyperalgesia, learned helplessness, and corticosterone production, supporting the use of non-steroidal anti-inflammatory drugs (NSAIDs) for persistent pain associated with chronic stress and depression.

  9. Discrimination between platelet-mediated and coagulation-mediated mechanisms in a model of complex thrombus formation in vivo

    SciTech Connect

    Cadroy, Y.; Horbett, T.A.; Hanson, S.R.

    1989-04-01

    To study mechanisms of complex thrombus formation in vivo, and to compare the relative antithrombotic effects of anticoagulants and antiplatelet agents, a model was developed in baboons. Segments of collagen-coated tubing followed by two sequentially placed expansion chambers exhibiting disturbed flow patterns were exposed to native blood under laminar flow conditions. The device was incorporated for 1 hour into an exteriorized arteriovenous shunt in baboons under controlled blood flow (20 ml/min). Morphologic evaluation by scanning electron microscopy showed that thrombi associated with collagen were relatively rich in platelets but thrombi in the chambers were rich in fibrin and red cells. Deposition of indium 111-labeled platelets was continuously measured with a scintillation camera. Platelet deposition increased in a linear (collagen-coated segment) or exponential (chambers 1 and 2) fashion over time, with values after 40 minutes averaging 24.1 +/- 3.3 x 10(8) platelets (collagen segment), 16.7 +/- 3.4 x 10(8) platelets (chamber 1), and 8.4 +/- 2.4 x 10(8) platelets (chamber 2). Total fibrinogen deposition after 40 minutes was determined by using iodine 125-labeled baboon fibrinogen and averaged 0.58 +/- 0.14 mg in the collagen segment, 1.51 +/- 0.27 mg in chamber 1, and 0.95 +/- 0.25 mg in chamber 2. Plasma levels of beta-thromboglobulin (beta TG), platelet-factor 4 (PF4), and fibrinopeptide A (FPA) increased fourfold to fivefold after 60 minutes of blood exposure to the thrombotic device. Platelet deposition onto the collagen segment, chamber 1, and chamber 2 was linearly dependent on the circulating platelet count. Platelet accumulation in chamber 1 and chamber 2 was also dependent on the presence of the proximal collagen segment.

  10. Circadian clocks govern calorie restriction-mediated life span extension through BMAL1- and IGF-1-dependent mechanisms.

    PubMed

    Patel, Sonal A; Chaudhari, Amol; Gupta, Richa; Velingkaar, Nikkhil; Kondratov, Roman V

    2016-04-01

    Calorie restriction (CR) increases longevity in many species by unknown mechanisms. The circadian clock was proposed as a potential mediator of CR. Deficiency of the core component of the circadian clock-transcriptional factor BMAL1 (brain and muscle ARNT [aryl hydrocarbon receptor nuclear translocator]-like protein 1)-results in accelerated aging. Here we investigated the role of BMAL1 in mechanisms of CR. The 30% CR diet increased the life span of wild-type (WT) mice by 20% compared to mice on anad libitum(AL) diet but failed to increase life span ofBmal1(-/-)mice. BMAL1 deficiency impaired CR-mediated changes in the plasma levels of IGF-1 and insulin. We detected a statistically significantly reduction of IGF-1 in CRvs.AL by 50 to 70% in WT mice at several daily time points tested, while inBmal1(-/-)the reduction was not significant. Insulin levels in WT were reduced by 5 to 9%, whileBmal1(-/-)induced it by 10 to 35% at all time points tested. CR up-regulated the daily average expression ofBmal1(by 150%) and its downstream target genesPeriods(by 470% forPer1and by 130% forPer2). We propose that BMAL1 is an important mediator of CR, and activation of BMAL1 might link CR mechanisms with biologic clocks.-Patel, S. A., Chaudhari, A., Gupta, R., Velingkaar, N., Kondratov, R. V. Circadian clocks govern calorie restriction-mediated life span extension through BMAL1- and IGF-1-dependent mechanisms.

  11. Vimentin contributes to epithelial-mesenchymal transition cancer cell mechanics by mediating cytoskeletal organization and focal adhesion maturation

    PubMed Central

    Liu, Ching-Yi; Lin, Hsi-Hui; Tang, Ming-Jer; Wang, Yang-Kao

    2015-01-01

    Modulations of cytoskeletal organization and focal adhesion turnover correlate to tumorigenesis and epithelial-mesenchymal transition (EMT), the latter process accompanied by the loss of epithelial markers and the gain of mesenchymal markers (e.g., vimentin). Clinical microarray results demonstrated that increased levels of vimentin mRNA after chemotherapy correlated to a poor prognosis of breast cancer patients. We hypothesized that vimentin mediated the reorganization of cytoskeletons to maintain the mechanical integrity in EMT cancer cells. By using knockdown strategy, the results showed reduced cell proliferation, impaired wound healing, loss of directional migration, and increased large membrane extension in MDA-MB 231 cells. Vimentin depletion also induced reorganization of cytoskeletons and reduced focal adhesions, which resulted in impaired mechanical strength because of reduced cell stiffness and contractile force. In addition, overexpressing vimentin in MCF7 cells increased cell stiffness, elevated cell motility and directional migration, reoriented microtubule polarity, and increased EMT phenotypes due to the increased β1-integrin and the loss of junction protein E-cadherin. The EMT-related transcription factor slug was also mediated by vimentin. The current study demonstrated that vimentin serves as a regulator to maintain intracellular mechanical homeostasis by mediating cytoskeleton architecture and the balance of cell force generation in EMT cancer cells. PMID:25965826

  12. Polycystin-1 Mediates Mechanical Strain-Induced Osteoblastic Mechanoresponses via Potentiation of Intracellular Calcium and Akt/β-Catenin Pathway

    PubMed Central

    Wang, Hua; Sun, Wen; Ma, Junqing; Pan, Yongchu; Wang, Lin; Zhang, Weibing

    2014-01-01

    Mechanical regulation of bone formation involves a complex biophysical process, yet the underlying mechanisms remain poorly understood. Polycystin-1 (PC1) is postulated to function as a mechanosensory molecule mediating mechanical signal transduction in renal epithelial cells. To investigate the involvement of PC1 in mechanical strain-induced signaling cascades controlling osteogenesis, PKD1 gene was stably silenced in osteoblastic cell line MC3T3-E1 by using lentivirus-mediated shRNA technology. Here, our findings showed that mechanical tensile strain sufficiently enhanced osteogenic gene expressions and osteoblastic proliferation. However, PC1 deficiency resulted in the loss of the ability to sense external mechanical stimuli thereby promoting osteoblastic osteogenesis and proliferation. The signal pathways implicated in this process were intracellular calcium and Akt/β-catenin pathway. The basal levels of intracellular calcium, phospho-Akt, phospho-GSK-3β and nuclear accumulation of active β-catenin were significantly attenuated in PC1 deficient osteoblasts. In addition, PC1 deficiency impaired mechanical strain-induced potentiation of intracellular calcium, and activation of Akt-dependent and Wnt/β-catenin pathways, which was able to be partially reversed by calcium ionophore A23187 treatment. Furthermore, applications of LiCl or A23187 in PC1 deficient osteoblasts could promote osteoblastic differentiation and proliferation under mechanical strain conditions. Therefore, our results demonstrated that osteoblasts require mechanosensory molecule PC1 to adapt to external mechanical tensile strain thereby inducing osteoblastic mechanoresponse, partially through the potentiation of intracellular calcium and downstream Akt/β-catenin signaling pathway. PMID:24618832

  13. Irinotecan-Induced Gastrointestinal Dysfunction and Pain Are Mediated by Common TLR4-Dependent Mechanisms.

    PubMed

    Wardill, Hannah R; Gibson, Rachel J; Van Sebille, Ysabella Z A; Secombe, Kate R; Coller, Janet K; White, Imogen A; Manavis, Jim; Hutchinson, Mark R; Staikopoulos, Vasiliki; Logan, Richard M; Bowen, Joanne M

    2016-06-01

    Strong epidemiological data indicate that chemotherapy-induced gut toxicity and pain occur in parallel, indicating common underlying mechanisms. We have recently outlined evidence suggesting that TLR4 signaling may contribute to both side effects. We therefore aimed to determine if genetic deletion of TLR4 improves chemotherapy-induced gut toxicity and pain. Forty-two female wild-type (WT) and 42 Tlr4 null (-/-) BALB/c mice weighing between 18 and 25 g (10-13 weeks) received a single 270 mg/kg (i.p.) dose of irinotecan hydrochloride or vehicle control and were killed at 6, 24, 48, 72, and 96 hours. Bacterial sequencing was conducted on cecal samples of control animals to determine the gut microbiome profile. Gut toxicity was assessed using validated clinical and histopathologic markers, permeability assays, and inflammatory markers. Chemotherapy-induced pain was assessed using the validated rodent facial grimace criteria, as well as immunologic markers of glial activation in the lumbar spinal cord. TLR4 deletion attenuated irinotecan-induced gut toxicity, with improvements in weight loss (P = 0.0003) and diarrhea (P < 0.0001). Crypt apoptosis was significantly decreased in BALB/c-Tlr4(-/-billy) mice (P < 0.0001), correlating with lower mucosal injury scores (P < 0.005). Intestinal permeability to FITC-dextran (4 kDa) and LPS translocation was greater in WT mice than in BALB/c-Tlr4(-/-billy) (P = 0.01 and P < 0.0001, respectively). GFAP staining in the lumbar spinal cord, indicative of astrocytic activation, was increased at 6 and 72 hours in WT mice compared with BALB/c-Tlr4(-/-billy) mice (P = 0.008, P = 0.01). These data indicate that TLR4 is uniquely positioned to mediate irinotecan-induced gut toxicity and pain, highlighting the possibility of a targetable gut/CNS axis for improved toxicity outcomes. Mol Cancer Ther; 15(6); 1376-86. ©2016 AACR.

  14. Mechanisms of Subsurface Drip Irrigation-Mediated Suppression of Lettuce Drop Caused by Sclerotinia minor.

    PubMed

    Bell, A A; Liu, L; Reidy, B; Davis, R M; Subbarao, K V

    1998-03-01

    ABSTRACT Subsurface drip irrigation and associated mandatory minimum tillage practices significantly reduced the incidence of lettuce drop (Sclerotinia minor) and the severity of corky root on lettuce compared with furrow irrigation and conventional tillage. Three possible mechanisms for the drip irrigation-mediated disease suppression were examined in this study: qualitative and quantitative differences in the soil microflora under furrow and subsurface drip irrigation; their antagonism and potential bio-control effects on S. minor; and the physical distribution of soil moisture and temperature relative to the two irrigation methods. To determine if the suppressive effects under subsurface drip irrigation were related to changes in soil microflora, soils were assayed for actinomycetes, bacteria, and fungi during the spring and fall seasons. The effects of the irrigation methods on microbial populations were nearly identical during both seasons. In the spring season, the total number of fungal colonies recovered on potato dextrose agar amended with rose Bengal generally was greater in soils under drip irrigation than under furrow irrigation, but no such differences were observed during the fall. Numbers of actinomycetes and bacteria were not significantly different between irrigation methods during either season. No interaction between sampling time and irrigation methods was observed for any of the microbial populations during both seasons. Thus, the significant effect of sampling time observed for actinomycete and bacterial populations during the spring was most likely not caused by the irrigation treatments. There were also no qualitative differences in the three groups of soil microflora between the irrigation treatments. Even though some fungal, actinomycete, and bacterial isolates suppressed mycelial growth of S. minor in in vitro assays, the isolates came from both subsurface drip- and furrow-irrigated soils. In in planta assays, selected isolates failed to

  15. Mechanisms of inflammation-mediated airway smooth muscle plasticity and airways remodeling in asthma.

    PubMed

    Halayko, Andrew J; Amrani, Yassine

    2003-09-16

    Recent evidence points to progressive structural change in the airway wall, driven by chronic local inflammation, as a fundamental component for development of irreversible airway hyperresponsiveness. Acute and chronic inflammation is orchestrated by cytokines from recruited inflammatory cells, airway myofibroblasts and myocytes. Airway myocytes exhibit functional plasticity in their capacity for contraction, proliferation, and synthesis of matrix protein and cytokines. This confers a principal role in driving different components of the airway remodeling process, and mediating constrictor hyperresponsiveness. Functional plasticity of airway smooth muscle (ASM) is regulated by an array of environmental cues, including cytokines, which mediate their effects through receptors and a number of intracellular signaling pathways. Despite numerous studies of the cellular effects of cytokines on cultured airway myocytes, few have identified how intracellular signaling pathways modulate or induce these cellular responses. This review summarizes current understanding of these concepts and presents a model for the effects of inflammatory mediators on functional plasticity of ASM in asthma.

  16. Molecular mechanism for USP7-mediated DNMT1 stabilization by acetylation

    NASA Astrophysics Data System (ADS)

    Cheng, Jingdong; Yang, Huirong; Fang, Jian; Ma, Lixiang; Gong, Rui; Wang, Ping; Li, Ze; Xu, Yanhui

    2015-05-01

    DNMT1 is an important epigenetic regulator that plays a key role in the maintenance of DNA methylation. Here we determined the crystal structure of DNMT1 in complex with USP7 at 2.9 Å resolution. The interaction between the two proteins is primarily mediated by an acidic pocket in USP7 and Lysine residues within DNMT1's KG linker. This intermolecular interaction is required for USP7-mediated stabilization of DNMT1. Acetylation of the KG linker Lysine residues impair DNMT1-USP7 interaction and promote the degradation of DNMT1. Treatment with HDAC inhibitors results in an increase in acetylated DNMT1 and decreased total DNMT1 protein. This negative correlation is observed in differentiated neuronal cells and pancreatic cancer cells. Our studies reveal that USP7-mediated stabilization of DNMT1 is regulated by acetylation and provide a structural basis for the design of inhibitors, targeting the DNMT1-USP7 interaction surface for therapeutic applications.

  17. Electroacupuncture Inhibition of Hyperalgesia in Rats with Adjuvant Arthritis: Involvement of Cannabinoid Receptor 1 and Dopamine Receptor Subtypes in Striatum

    PubMed Central

    Shou, Yin; Yang, Yang; Xu, Ming-Shu; Zhao, Ying-Qian; Ge, Lin-Bao; Zhang, Bi-Meng

    2013-01-01

    Electroacupuncture (EA) has been regarded as an alternative treatment for inflammatory pain for several decades. However, the molecular mechanisms underlying the antinociceptive effect of EA have not been thoroughly clarified. Previous studies have shown that cannabinoid CB1 receptors are related to pain relief. Accumulating evidence has shown that the CB1 and dopamine systems sometimes interact and may operate synergistically in rat striatum. To our knowledge, dopamine D1/D2 receptors are involved in EA analgesia. In this study, we found that repeated EA at Zusanli (ST36) and Kunlun (BL60) acupoints resulted in marked improvements in thermal hyperalgesia. Both western blot assays and FQ-PCR analysis results showed that the levels of CB1 expression in the repeated-EA group were much higher than those in any other group (P = 0.001). The CB1-selective antagonist AM251 inhibited the effects of repeated EA by attenuating the increases in CB1 expression. The two kinds of dopamine receptors imparted different actions on the EA-induced CB1 upregulation in AA rat model. These results suggested that the strong activation of the CB1 receptor after repeated EA resulted in the concomitant phenomenon of the upregulation of D1 and D2 levels of gene expression. PMID:23762129

  18. N-acetyl-cysteine attenuates remifentanil-induced postoperative hyperalgesia via inhibiting matrix metalloproteinase-9 in dorsal root ganglia.

    PubMed

    Liu, Yue; Ni, Yuan; Zhang, Wei; Sun, Yu-E; Ma, Zhengliang; Gu, Xiaoping

    2017-02-09

    Treatment of remifentanil-induced postoperative hyperalgesia (RIH) remains a clinical challenge because the mechanisms are not fully understood. Matrix metalloproteinase-9 (MMP-9) is a key component in neuroinflammation because of its facilitation of pro-inflammatory cytokine maturation. Therefore, inhibition of MMP-9 may represent a novel therapeutic approach to the treatment of RIH. Sprague-Dawley rats were randomly divided into three groups: Control, Incision and Remifentanil. A right plantar surgical incision was performed in Group Incision, and intraoperative remifentanil (0.04 mg/kg, 0.4 ml) was infused subcutaneously for 30 min in Group Remifentanil. The results indicated that intraoperative remifentanil induced an up-regulation and activation of MMP-9 in DRGs but not spinal cords. MMP-9 was expressed primarily in DRG neurons co-expressing mu opioid receptors (MOR), and elicited interleukin-1β (IL-1β) cleavage in DRG neurons and satellite glial cells (SGCs). Intraperitoneal injection of N-acetyl-cysteine (NAC), a broadly used safe drug, significantly attenuated RIH via suppressing the activation of MMP-9 in DRGs. NAC inhibited the cleavage of IL-1β in DRGs, which is a critical substrate of MMP-9, and markedly suppressed glial activation and neuron excitability in spinal dorsal horn induced by remifentanil. These results demonstrated that NAC can effectively alleviate RIH via powerfully inhibiting MMP-9 activation in DRGs.

  19. Alcohol consumption enhances antiretroviral painful peripheral neuropathy by mitochondrial mechanisms.

    PubMed

    Ferrari, Luiz F; Levine, Jon D

    2010-09-01

    A major dose-limiting side effect of human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) chemotherapies, such as the nucleoside reverse transcriptase inhibitors (NRTIs), is a small-fiber painful peripheral neuropathy, mediated by its mitochondrial toxicity. Co-morbid conditions may also contribute to this dose-limiting effect of HIV/AIDS treatment. Alcohol abuse, which alone also produces painful neuropathy, is one of the most important co-morbid risk factors for peripheral neuropathy in patients with HIV/AIDS. Despite the prevalence of this problem and its serious impact on the quality of life and continued therapy in HIV/AIDS patients, the mechanisms by which alcohol abuse exacerbates highly active antiretroviral therapy (HAART)-induced neuropathic pain has not been demonstrated. In this study, performed in rats, we investigated the cellular mechanism by which consumed alcohol impacts antiretroviral-induced neuropathic pain. NRTI 2',3'-dideoxycytidine (ddC; 50 mg/kg) neuropathy was mitochondrial-dependent and PKCε-independent, and alcohol-induced painful neuropathy was PKCε-dependent and mitochondrial-independent. At low doses, ddC (5 mg/kg) and alcohol (6.5% ethanol diet for 1 week), which alone do not affect nociception, together produce profound mechanical hyperalgesia. This hyperalgesia is mitochondrial-dependent but PKCε-independent. These experiments, which provide the first model for studying the impact of co-morbidity in painful neuropathy, support the clinical impression that alcohol consumption enhances HIV/AIDS therapy neuropathy, and provide evidence for a role of mitochondrial mechanisms underlying this interaction.

  20. Molecular Mechanisms for cAMP-Mediated Immunoregulation in T cells – Role of Anchored Protein Kinase A Signaling Units

    PubMed Central

    Wehbi, Vanessa L.; Taskén, Kjetil

    2016-01-01

    The cyclic AMP/protein kinase A (cAMP/PKA) pathway is one of the most common and versatile signal pathways in eukaryotic cells. A-kinase anchoring proteins (AKAPs) target PKA to specific substrates and distinct subcellular compartments providing spatial and temporal specificity for mediation of biological effects channeled through the cAMP/PKA pathway. In the immune system, cAMP is a potent negative regulator of T cell receptor-mediated activation of effector T cells (Teff) acting through a proximal PKA/Csk/Lck pathway anchored via a scaffold consisting of the AKAP Ezrin holding PKA, the linker protein EBP50, and the anchoring protein phosphoprotein associated with glycosphingolipid-enriched microdomains holding Csk. As PKA activates Csk and Csk inhibits Lck, this pathway in response to cAMP shuts down proximal T cell activation. This immunomodulating pathway in Teff mediates clinically important responses to regulatory T cell (Treg) suppression and inflammatory mediators, such as prostaglandins (PGs), adrenergic stimuli, adenosine, and a number of other ligands. A major inducer of T cell cAMP levels is PG E2 (PGE2) acting through EP2 and EP4 prostanoid receptors. PGE2 plays a crucial role in the normal physiological control of immune homeostasis as well as in inflammation and cancer immune evasion. Peripherally induced Tregs express cyclooxygenase-2, secrete PGE2, and elicit the immunosuppressive cAMP pathway in Teff as one tumor immune evasion mechanism. Moreover, a cAMP increase can also be induced by indirect mechanisms, such as intercellular transfer between T cells. Indeed, Treg, known to have elevated levels of intracellular cAMP, may mediate their suppressive function by transferring cAMP to Teff through gap junctions, which we speculate could also be regulated by PKA/AKAP complexes. In this review, we present an updated overview on the influence of cAMP-mediated immunoregulatory mechanisms acting through localized cAMP signaling and the therapeutical

  1. A novel Netrin-1–sensitive mechanism promotes local SNARE-mediated exocytosis during axon branching

    PubMed Central

    Winkle, Cortney C.; McClain, Leslie M.; Valtschanoff, Juli G.; Park, Charles S.; Maglione, Christopher

    2014-01-01

    Developmental axon branching dramatically increases synaptic capacity and neuronal surface area. Netrin-1 promotes branching and synaptogenesis, but the mechanism by which Netrin-1 stimulates plasma membrane expansion is unknown. We demonstrate that SNARE-mediated exocytosis is a prerequisite for axon branching and identify the E3 ubiquitin ligase TRIM9 as a critical catalytic link between Netrin-1 and exocytic SNARE machinery in murine cortical neurons. TRIM9 ligase activity promotes SNARE-mediated vesicle fusion and axon branching in a Netrin-dependent manner. We identified a direct interaction between TRIM9 and the Netrin-1 receptor DCC as well as a Netrin-1–sensitive interaction between TRIM9 and the SNARE component SNAP25. The interaction with SNAP25 negatively regulates SNARE-mediated exocytosis and axon branching in the absence of Netrin-1. Deletion of TRIM9 elevated exocytosis in vitro and increased axon branching in vitro and in vivo. Our data provide a novel model for the spatial regulation of axon branching by Netrin-1, in which localized plasma membrane expansion occurs via TRIM9-dependent regulation of SNARE-mediated vesicle fusion. PMID:24778312

  2. PUVB-mediated prevention of luminal narrowing after arterial wall injury: modulation of mechanical arterial properties as a putative mechanism of action

    NASA Astrophysics Data System (ADS)

    Perree, Jop; Kerindongo, Raphaela P.; van Leeuwen, Ton G. J. M.

    2001-10-01

    In a previous study we have found that the photodynamic modality PUVB (8-methoxy-Psoralen + UVB) reduces luminal narrowing after arterial endovascular injury. We hypothesized that PUVB may modulate the arterial mechanical properties and tested this hypothesis by measuring the stress as a function of the strain in segments of carotid artery. Furthermore, we have investigated the potential for PUVB-induced cross-linking of extracellular matrix proteins by gel electrophoresis. It was found that both techniques were suitable for testing our hypotheses as evidenced by a statistically significant difference for the positive control. However, no differences between A) control, B) sensitizer only, C) light only and D) PUVB-treated samples could be found with respect to macro- and micro-mechanical properties. Therefore, the hypothesis that PUVB mediates its luminal narrowing reduction effect by directly changing the arterial mechanical properties should be rejected.

  3. A Nibbling Mechanism for Clathrin-mediated Retrieval of Secretory Granule Membrane after Exocytosis*

    PubMed Central

    Bittner, Mary A.; Aikman, Rachel L.; Holz, Ronald W.

    2013-01-01

    Clathrin-mediated endocytosis is the major pathway for recycling of granule membrane components after strong stimulation and high exocytotic rates. It resembles “classical” receptor-mediated endocytosis but has a trigger that is unique to secretion, the sudden appearance of the secretory granule membrane in the plasma membrane. The spatial localization, the relationship to individual fusion events, the nature of the cargo, and the timing and nature of the nucleation events are unknown. Furthermore, a size mismatch between chromaffin granules (∼300-nm diameter) and typical clathrin-coated vesicles (∼90 nm) makes it unlikely that clathrin-mediated endocytosis internalizes as a unit the entire fused granule membrane. We have used a combination of total internal reflection fluorescence microscopy of transiently expressed proteins and time-resolved quantitative confocal imaging of endogenous proteins along with a fluid-phase marker to address these issues. We demonstrate that the fused granule membrane remains a distinct entity and serves as a nucleation site for clathrin- and dynamin-mediated endocytosis that internalizes granule membrane components in small increments. PMID:23386611

  4. Perilipin Promotes HSL-Mediated Adipocyte Lipolysis via Phosphorylation-dependent and Independent Mechanisms

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Hormone-sensitive lipase (HSL) is the predominant lipase effector of catecholamine-stimulated lipolysis in adipocytes. HSL-dependent lipolysis, in response to catecholamines, is mediated by protein kinase A (PKA)-dependent phosphorylation of perilipin A (Peri A), an essential lipid droplet (LD)-ass...

  5. An epigenetic mechanism mediates developmental nicotine effects on neuronal structure and behavior

    PubMed Central

    Jung, Yonwoo; Hsieh, Lawrence S.; Lee, Angela M.; Zhou, Zhifeng; Coman, Daniel; Heath, Christopher J.; Hyder, Fahmeed; Mineur, Yann S.; Yuan, Qiaoping; Goldman, David; Bordey, Angelique; Picciotto, Marina R.

    2016-01-01

    Developmental nicotine exposure causes persistent changes in cortical neuron morphology and in behavior. We used microarray screening to identify master transcriptional or epigenetic regulators mediating these effects of nicotine and discovered increases in Ash2l, a component of a histone methyltransferase complex. We therefore examined genome-wide changes in H3K4 tri-methylation, a mark induced by the Ash2l complex associated with increased gene transcription. A significant number of regulated promoter sites were involved in synapse maintenance. We found that Mef2c interacts with Ash2l and mediates changes in H3K4 tri-methylation. Knockdown of Ash2l or Mef2c abolishes nicotine-mediated alterations of dendritic complexity in vitro and in vivo, and attenuates nicotine-dependent changes in passive avoidance behavior. In contrast, overexpression mimics nicotine-mediated alterations of neuronal structure and passive avoidance behavior. These studies identify Ash2l as a novel target induced by nicotinic stimulation that couples developmental nicotine exposure to changes in brain epigenetic marks, neuronal structure and behavior. PMID:27239938

  6. Mechanically induced c-fos expression is mediated by cAMP in MC3T3-E1 osteoblasts

    NASA Technical Reports Server (NTRS)

    Fitzgerald, J.; Hughes-Fulford, M.

    1999-01-01

    In serum-deprived MC3T3-E1 osteoblasts, mechanical stimulation caused by mild (287 x g) centrifugation induced a 10-fold increase in mRNA levels of the proto-oncogene, c-fos. Induction of c-fos was abolished by the cAMP-dependent protein kinase inhibitor H-89, suggesting that the transient c-fos mRNA increase is mediated by cAMP. Down-regulation of protein kinase C (PKC) activity by chronic TPA treatment failed to significantly reduce c-fos induction, suggesting that TPA-sensitive isoforms of PKC are not responsible for c-fos up-regulation. In addition, 287 x g centrifugation increased intracellular prostaglandin E2 (PGE2) levels 2.8-fold (P<0. 005). Since we have previously shown that prostaglandin E2 (PGE2) can induce c-fos expression via a cAMP-mediated mechanism, we asked whether the increase in c-fos mRNA was due to centrifugation-induced PGE2 release. Pretreatment with the cyclooxygenase inhibitors indomethacin and flurbiprofen did not hinder the early induction of c-fos by mechanical stimulation. We conclude that c-fos expression induced by mild mechanical loading is dependent primarily on cAMP, not PKC, and initial induction of c-fos is not necessarily dependent on the action of newly synthesized PGE2.

  7. A Novel Perspective and Approach to Intestinal Octreotide Absorption: Sinomenine-Mediated Reversible Tight Junction Opening and Its Molecular Mechanism

    PubMed Central

    Li, Yuling; Duan, Zhijun; Tian, Yan; Liu, Zhen; Wang, Qiuming

    2013-01-01

    In this work, we assessed the effects of sinomenine (SN) on intestinal octreotide (OCT) absorption both in Caco-2 cell monolayers and in rats. We also investigated the molecular mechanisms of tight junction (TJ) disruption and recovery by SN-mediated changes in the claudin-1 and protein kinase C (PKC) signaling pathway. The data showed that exposure to SN resulted in a significant decrease in the expression of claudin-1, which represented TJ weakening and paracellular permeability enhancement. Then, the recovery of TJ after SN removal required an increase in claudin-1, which demonstrated the transient and reversible opening for TJ. Meanwhile, the SN-mediated translocation of PKC-α from the cytosol to the membrane was found to prove PKC activation. Finally, SN significantly improved the absolute OCT bioavailability in rats and the transport rate in Caco-2 cell monolayers. We conclude that SN has the ability to enhance intestinal OCT absorption and that these mechanisms are related at least in part to the important role of claudin-1 in SN-mediated, reversible TJ opening via PKC activation. PMID:23787475

  8. An Fcγ receptor-dependent mechanism drives antibody-mediated target-receptor signaling in cancer cells.

    PubMed

    Wilson, Nicholas S; Yang, Becky; Yang, Annie; Loeser, Stefanie; Marsters, Scot; Lawrence, David; Li, Yun; Pitti, Robert; Totpal, Klara; Yee, Sharon; Ross, Sarajane; Vernes, Jean-Michel; Lu, Yanmei; Adams, Cam; Off