Sample records for pyridinealdoxime methiodide pam-type

  1. Blood-brain barrier penetration of novel pyridinealdoxime methiodide (PAM)-type oximes examined by brain microdialysis with LC-MS/MS

    SciTech Connect

    Okuno, Sou; Sakurada, Koichi Ohta, Hikoto; Ikegaya, Hiroshi; Kazui, Yuko; Akutsu, Tomoko; Takatori, Takehiko; Iwadate, Kimiharu


    To develop a new reactivator of inhibited acetylcholinesterase (AChE) that can easily penetrate the blood-brain barrier (BBB), BBB penetration of 6 known and novel pyridinealdoxime methiodide (PAM)-type oximes (alkylPAMs) with relatively high reactivation activities was examined by in vivo rat brain microdialysis with liquid chromatography-mass spectrometry (LC-MS/MS). The 50% lethal dose (LD{sub 50}) of alkylPAMs was intravenously determined for Wistar rats, then the limit of detection, quantification range and linearity of the calibration curve of the alkylPAMs in dialysate and blood were determined by LC-MS/MS. Following 10% LD{sub 50} intravenous administration of the alkylPAMs, 4-[(hydroxyimino) methyl]-1-(2-phenylethyl) pyridinium bromide (4-PAPE) and 4-[(hydroxyimino) methyl]-1-octylpyridinium bromide (4-PAO) appeared in the dialysate. Striatal extracellular fluid/blood concentration ratios were 0.039 {+-} 0.018 and 0.301 {+-} 0.183 (mean {+-} SEM), respectively, 1 h after treatment. This is the first report of BBB penetration of 4-PAPE, and the concentration ratio was smaller than that of 2-PAM.The mean BBB penetration of 4-PAO was approximately 30%, indicating that intravenous administration of 4-PAO may be effective for the reactivation of blocked cholinesterase in the brain. However, the toxicity of 4-PAO (LD{sub 50}; 8.89 mg/kg) was greater than that of 2-PAM. Further investigation is required to determine the effects of these alkylPAMs in organophosphate poisoning.

  2. Intravenous kinetics and metabolism of [15,16-3H]naltrexonium methiodide in the rat.


    Misra, A L; Pontani, R B; Vadlamani, N L


    After a 4 mg kg-1 bolus intravenous dose of [15,16-3H]naltrexonium methiodide to the rat, brain to plasma concentration ratios of the compound were 0.031 to 0.228 between 0.25 to 6 h after injection and the t 1/2 beta in plasma and brain were 2.92 and 7.61 h, respectively. Ethyl acetate-extracted radioactivity due to metabolites in plasma decayed with t 1/2 beta 1.83 h and the ratios of plasma concentration of metabolites to quaternary compound between 0.25 and 6 h were 0.014-0.026. Only unconjugated 7,8-dihydro-14-hydroxynormorphine, naltrexone and traces of 7,8-dihydro-14-hydroxynormorphinone were the metabolites in plasma. Naltrexone (but not normetabolites) was present only in traces in brain up to 0.5 h after injection and not at later times. PMID:2883290

  3. Cocaine-induced cardiovascular effects: lack of evidence for a central nervous system site of action based on hemodynamic studies with cocaine methiodide.


    Dickerson, L W; Rodak, D J; Kuhn, F E; Wahlstrom, S K; Tessel, R E; Visner, M S; Schaer, G L; Gillis, R A


    It has been suggested that cocaine acts directly in the brain to enhance central sympathetic outflow. However, some studies suggested that the cardiovascular effects of cocaine are related to a peripheral action. To characterize further the site of cocaine's cardiovascular effect, we compared the hemodynamic effects of cocaine (2 mg/kg, i.v. bolus) with those observed after administration of an equimolar dose (2.62 mg/kg, i.v. bolus) of cocaine methiodide, a quaternary derivative of cocaine that does not penetrate the blood-brain barrier, by using sufentanil-sedated dogs. Cocaine produced significant (p < 0.05) increases in heart rate (+37+/-11 beats/min), mean arterial pressure (+55+/-11 mm Hg), left ventricular end-diastolic pressure (+5.3+/-1.0 mm Hg), and cardiac output (+2.4+/-0.9 L/min). Cocaine methiodide produced increases in heart rate (+57+/-11 beats/min), mean arterial pressure (+45+/-11 mm Hg), left ventricular end-diastolic pressure (+3.4+/-1.0 mm Hg), and cardiac output (1.1+/-0.9 L/min), which were not significantly different from those observed with cocaine. Because opiate sedation potentially might have attenuated central sympathetic outflow, we further confirmed the qualitative similarity of the actions of cocaine and cocaine methiodide on heart rate and blood pressure in unsedated, conscious dogs. Our data suggest that the cardiovascular effects of cocaine result primarily from a peripheral site of action.

  4. Pyridine-2-aldoxime methiodide. A valuable agent for phosphate poisoning.




    Phosphate insecticide use is increasing as is concomitant human poisoning. Home insecticide bomb as well as agricultural, crop contamination and suicidal exposure are noted. Clinical poisoning may be chronic and severe. It may follow long exposure or short exposure with heavy dosages. Manifestations are those of excessive cholinergic activity.Adequate laboratory means for early, rapid diagnosis and screen testings are available.PAM is a valuable agent for this type of poisoning and is a much more adequate and complete antidote than atropine. It is available (under certain restrictive conditions presently). It is being widely used elsewhere in the world but with limited education and use in this country. Morbidity and mortality continue at a rate that could probably be corrected. Case reports, describing the use of this antidote in our hands, are included. Government and industry responsibility as well as physician education must be more clearly defined in prevention, recognition and treatment in what is often a life threatening situation.

  5. Refinement of structural leads for centrally acting oxime reactivators of phosphylated cholinesterases.


    Radić, Zoran; Sit, Rakesh K; Kovarik, Zrinka; Berend, Suzana; Garcia, Edzna; Zhang, Limin; Amitai, Gabriel; Green, Carol; Radić, Bozica; Fokin, Valery V; Sharpless, K Barry; Taylor, Palmer


    We present a systematic structural optimization of uncharged but ionizable N-substituted 2-hydroxyiminoacetamido alkylamine reactivators of phosphylated human acetylcholinesterase (hAChE) intended to catalyze the hydrolysis of organophosphate (OP)-inhibited hAChE in the CNS. Starting with the initial lead oxime RS41A identified in our earlier study and extending to the azepine analog RS194B, reactivation rates for OP-hAChE conjugates formed by sarin, cyclosarin, VX, paraoxon, and tabun are enhanced severalfold in vitro. To analyze the mechanism of intrinsic reactivation of the OP-AChE conjugate and penetration of the blood-brain barrier, the pH dependence of the oxime and amine ionizing groups of the compounds and their nucleophilic potential were examined by UV-visible spectroscopy, (1)H NMR, and oximolysis rates for acetylthiocholine and phosphoester hydrolysis. Oximolysis rates were compared in solution and on AChE conjugates and analyzed in terms of the ionization states for reactivation of the OP-conjugated AChE. In addition, toxicity and pharmacokinetic studies in mice show significantly improved CNS penetration and retention for RS194B when compared with RS41A. The enhanced intrinsic reactivity against the OP-AChE target combined with favorable pharmacokinetic properties resulted in great improvement of antidotal properties of RS194B compared with RS41A and the standard peripherally active oxime, 2-pyridinealdoxime methiodide. Improvement was particularly noticeable when pretreatment of mice with RS194B before OP exposure was combined with RS194B reactivation therapy after the OP insult.

  6. Conditioned Contribution of Peripheral Cocaine Actions to Cocaine Reward and Cocaine-Seeking

    PubMed Central

    Wang, Bin; You, Zhi-Bing; Oleson, Erik B; Cheer, Joseph F; Myal, Stephanie; Wise, Roy A


    Cocaine has actions in the peripheral nervous system that reliably precede—and thus predict—its soon-to-follow central rewarding effects. In cocaine-experienced animals, the peripheral cocaine signal is relayed to the central nervous system, triggering excitatory input to the ventral tegmental origin of the mesocorticolimbic dopamine system, the system that mediates the rewarding effects of the drug. We used cocaine methiodide, a cocaine analog that does not cross the blood–brain barrier, to isolate the peripheral actions of cocaine and determine their central and behavioral effects in animals first trained to lever-press for cocaine hydrochloride (the centrally acting and abused form of the drug). We first confirmed with fast-scan cyclic voltammetry that cocaine methiodide causes rapid dopamine release from dopamine terminals in cocaine hydrochloride-trained rats. We then compared the ability of cocaine hydrochloride and cocaine methiodide to establish conditioned place preferences in rats with self-administration experience. While cocaine hydrochloride established stronger place preferences, cocaine methiodide was also effective and its effectiveness increased (incubated) over weeks of cocaine abstinence. Cocaine self-administration was extinguished when cocaine methiodide or saline was substituted for cocaine hydrochloride in the intravenous self-administration paradigm, but cocaine hydrochloride and cocaine methiodide each reinstated non-rewarded lever-pressing after extinction. Rats extinguished by cocaine methiodide substitution showed weaker cocaine-induced reinstatement than rats extinguished by saline substitution. These findings suggest that the conditioned peripheral effects of cocaine can contribute significantly to cocaine-induced (but not stress-induced) cocaine craving, and also suggest the cocaine cue as an important target for cue-exposure therapies for cocaine addiction. PMID:23535778

  7. Naloxone improves functional recovery of myocardial stunning in conscious dogs through its action on the central nervous system.


    Weber, T P; Stypmann, J; Meissner, A; Hartlage, M G; Van Aken, H; Rolf, N


    This study tests the hypothesis that naloxone, but not its quarternary salt, naloxone methiodide (which does not enter the central nervous system), improves recovery from myocardial stunning in conscious dogs. Twenty dogs were chronically instrumented for measurement of heart rate, left atrial, aortic and left ventricular pressure (LVP), LV dP x dtmax(-1) and myocardial wall thickening fraction (WTF). Regional myocardial blood flow was determined with coloured microspheres. Occluder around the left anterior descending artery (LAD) allowed induction of reversible LAD ischaemia. Each of the 20 dogs underwent two LAD ischaemic challenges. Experiments (performed on separate days, in crossover fashion) were: (i) 10 min of LAD occlusion after application of naloxone 63 microg kg(-1) or naloxone methiodide 63 microg kg(-1) and (ii) occlusion without naloxone or naloxone methiodide. WTF was measured at baseline and until complete recovery occurred. LAD ischaemia significantly reduced LAD WTF with (mean (SD) 54 (15)% lower than baseline) and without naloxone (55 (16)% lower than baseline), without significant haemodynamic differences. Between I to 30 min of reperfusion, WTF was significantly higher with naloxone (P < 0.05). There was no difference in WTF with or without naloxone methiodide. We conclude that naloxone improved recovery from myocardial stunning in conscious dogs, and that this was centrally mediated. PMID:11573630

  8. Pilocarpine modulates the cellular electrical properties of mammalian hearts by activating a cardiac M3 receptor and a K+ current

    PubMed Central

    Wang, Huizhen; Shi, Hong; Lu, Yanjie; Yang, Baofeng; Wang, Zhiguo


    Pilocarpine, a muscarinic acetylcholine receptor (mAChR) agonist, is widely used for treatment of xerostomia and glaucoma. It can also cause many other cellular responses by activating different subtypes of mAChRs in different tissues. However, the potential role of pilocarpine in modulating cardiac function remained unstudied.We found that pilocarpine produced concentration-dependent (0.1–10 μM) decrease in sinus rhythm and action potential duration, and hyperpolarization of membrane potential in guinea-pig hearts. The effects were nearly completely reversed by 1 μM atropine or 2 nM 4DAMP methiodide (an M3-selective antagonist).Patch-clamp recordings in dispersed myocytes from guinea-pig and canine atria revealed that pilocarpine induces a novel K+ current with delayed rectifying properties. The current was suppressed by low concentrations of M3-selective antagonists 4DAMP methiodide (2–10 nM), 4DAMP mustard (4–20 nM, an ackylating agent) and p-F-HHSiD (20–200 nM). Antagonists towards other subtypes (M1, M2 or M4) all failed to alter the current.The affinity of pilocarpine (KD) at mAChRs derived from displacement binding of [3H]-NMS in the homogenates from dog atria was 2.2 μM (65% of the total binding) and that of 4DAMP methiodide was 2.8 nM (70% of total binding), consistent with the concentration of pilocarpine needed for the current induction and for the modulation of the cardiac electrical activity and the concentration of 4DAMP to block pilocarpine effects.Our data indicate, for the first time, that pilocarpine modulates the cellular electrical properties of the hearts, likely by activating a K+ current mediated by M3 receptors. PMID:10372814

  9. Peripherally mediated antinociception of the mu-opioid receptor agonist 2-[(4,5alpha-epoxy-3-hydroxy-14beta-methoxy-17-methylmorphinan-6beta-yl)amino]acetic acid (HS-731) after subcutaneous and oral administration in rats with carrageenan-induced hindpaw inflammation.


    Bileviciute-Ljungar, Indre; Spetea, Mariana; Guo, Yan; Schütz, Johannes; Windisch, Petra; Schmidhammer, Helmut


    Opioids induce analgesia by activating opioid receptors not only within the central nervous system but also on peripheral sensory neurons. This study investigated peripherally mediated antinociception produced by the mu-opioid receptor agonist 2-[(4,5alpha-epoxy-3-hydroxy-14beta-methoxy-17-methylmorphinan-6beta-yl)amino]acetic acid (HS-731) after s.c. and oral administration in rats with carrageenan-induced hindpaw inflammation. Antinociceptive effects after s.c. administration were assessed 3 h after intraplantar carrageenan injection and compared with those of centrally acting mu-opioid agonists 14-methoxymetopon and morphine. Opioid agonists caused dose-dependent increases in inflamed paw withdrawal latencies to mechanical and thermal stimulation. The time course of action was different, in that HS-731 (20 microg/kg s.c.) produced significant long-lasting effects up to 4 h after administration, whereas 14-methoxymetopon (20 microg/kg) and morphine (2 mg/kg) reached their peak of action at 10 to 30 min, and their effect declined rapidly thereafter. Subcutaneous administration of the peripherally selective opioid antagonist naloxone methiodide inhibited antinociception elicited by HS-731 (20 microg/kg s.c.), whereas it was ineffective against 14-methoxymetopon (20 microg/kg s.c.). Moreover, the antinociception produced by 100 microg/kg s.c. HS-731 was dose-dependently reversed by s.c. naloxone methiodide. This indicates that HS-731 preferentially activates peripheral opioid receptors, whereas 14-methoxymetopon mediates analgesia via central mechanisms. Orally administered HS-731 significantly reduced hyperalgesia in the inflamed paw induced by carrageenan, which was reversible by s.c. administered naloxone methiodide. These results show that systemic (s.c. and oral) treatment with the mu-opioid agonist HS-731 produces potent and long-lasting antinociception through peripheral mechanisms in rats with carrageenan-induced hindpaw inflammation.

  10. Differences in the morphine-induced inhibition of small and large intestinal transit: Involvement of central and peripheral μ-opioid receptors in mice.


    Matsumoto, Kenjiro; Umemoto, Hiroyuki; Mori, Tomohisa; Akatsu, Ryuya; Saito, Shinichiro; Tashima, Kimihito; Shibasaki, Masahiro; Kato, Shinichi; Suzuki, Tsutomu; Horie, Syunji


    Constipation is the most common side effect of morphine. Morphine acts centrally and on peripheral sites within the enteric nervous system. There are a few comprehensive studies on morphine-induced constipation in the small and large intestine by the activation of central and peripheral μ-opioid receptors. We investigated the differences in the inhibition of the small and large intestinal transit in normal and morphine-tolerant mice. Morphine reduced the geometric center in the fluorescein isothiocyanate-dextran assay and prolonged the bead expulsion time in a dose-dependent manner. The inhibitory effects of morphine were blocked by μ-opioid antagonist β-funaltrexamine, but not by δ- and κ-opioid antagonists. The peripheral opioid receptor antagonist, naloxone methiodide, partially blocked morphine's effect in the small intestine and completely blocked its effect in the large intestine. The intracerebroventricular administration of naloxone significantly reversed the delay of small intestinal transit but did not affect morphine-induced inhibition of large intestinal transit. Naloxone methiodide completely reversed the inhibition of large intestinal transit in normal and morphine-tolerant mice. Naloxone methiodide partially reversed the morphine-induced inhibition of small intestinal transit in normal mice but completely reversed the effects of morphine in tolerant mice. Chronic treatment with morphine results in tolerance to its inhibitory effect on field-stimulated contraction in the isolated small intestine but not in the large intestine. These results suggest that peripheral and central opioid receptors are involved in morphine-induced constipation in the small and large intestine during the early stage of treatment, but the peripheral receptors mainly regulate constipation during long-term morphine treatment.

  11. The pharmacology of the cholinoceptor in muscle preparations of Ascaris lumbricoides var. suum

    PubMed Central

    Natoff, I. L.


    1. The preparation of a muscle strip of Ascaris lumbricoides for the study of the effect of drugs in vitro is described. 2. Stimulant drugs which are classified as nicotine-like in mammalian pharmacology increased the isometric tension of this preparation. These drugs were, in descending order of potency: dimethylphenylpiperazinium, nicotine, acetylcholine, carbachol, decamethonium and pyridine-2-aldoxime methiodide. 3. Muscarine-like drugs (oxotremorine, methacholine, pilocarpine) had no activity. 4. Potassium and barium ions stimulated the tissue, while the anti-cholinesterases, dichlorvos and eserine, increased the resting tension of the preparation and potentiated the responses to acetylcholine. 5. Adrenaline neither stimulated the tissue nor affected the responses to nicotine-like drugs. 6. The relative potency of several blocking agents which antagonize the responses to nicotine-like drugs was assayed. These blocking agents were, in descending order of potency: mecamylamine, (+)-tubocurarine, hexamethonium, atropine and piperazine. Acetylcholine, dimethylphenylpiperazinium and pyridine-2-aldoxime methiodide apparently act on a common receptor, for each blocking agent had a similar molar inhibitory concentration against these stimulants. 7. It is concluded that the cholinoceptor in muscle preparations of Ascaris lumbricoides is pharmacologically similar to that of the mammalian autonomic ganglion. PMID:4390485

  12. Activation of Opioid μ-Receptors, but not δ- or κ-Receptors, Switches Pulmonary C-Fiber-Mediated Rapid Shallow Breathing into An Apnea in Anesthetized Rats

    PubMed Central

    Zhang, Zhenxiong; Zhang, Cancan; Zhou, Moxi; Xu, Fadi


    Rapid shallow breathing (RSB) is mainly mediated by bronchopulmonary C-fibers (PCFs). We asked whether this RSB could be modulated by opioid. In anesthetized rats right atrial bolus injection of phenylbiguanide (PBG) to evoke RSB was repeated after: 1) intravenously giving fentanyl (μ-receptor agonist), DPDPE (δ-receptor agonist), or U-50488H (κ-receptor agonist); 2) fentanyl (iv) following naloxone methiodide, a peripheral opioid receptor antagonist; 3) bilateral microinjection of fentanyl into the nodose ganglia; 4) fentanyl (iv) with pre-blocking histamine H1 and H2 receptors by diphenhydramine and ranitidine. Systemic fentanyl challenge, but not DPDPE or U-50488H, switched the PBG-induced RSB to a long lasting apnea. This switch was blocked by naloxone methiodide rather than diphenhydramine and ranitidine. After microinjecting fentanyl into the nodose ganglia, PBG also produced an apnea. Our results suggest that activating μ-receptors is capable of turning the PCF-mediated RSB into an apnea, at least partly, via facilitating PCFs’ activity and this switching effect appears independent of the released histamine. PMID:22796630

  13. [Investigation of functional groups of Cryptococcus albidus alpha-L-rhamnosidase].


    Gudzenko, O V; Varbanets', L D


    The effect of cations, anions and specific chemical reagents: 1-[3-(dimethylamino)propyl]-3-ethylcarbodimide methiodide, EDTA, o-phenantroline, dithiotreitol, L-cysteine, beta-mercaptoethanol, p-chlormercurybenzoate (p-ChMB), N-ethylmaleimide on the alpha-L-rhamnosidase activity of Cryptococcus albidus has been investigated. The essential role of Ag+ which inhibits the alpha-L-rhamnosidase activity by 72.5% was shown. Rhamnose at 1-5 mM protect the enzyme from the negative effect of Ag(+). It was expected that carboxyl group of C-terminal aminoacid and imidazole group of histidine would participate in the catalytic action of alpha-L-rhamnosidase on the basis of inhibition and kinetic analysis. PMID:23088096

  14. Clinical observation and comparison of the effectiveness of several oxime cholinesterase reactivators.


    Xue, S Z; Ding, X J; Ding, Y


    After passing toxicity and experimental therapeutic tests, four oxime cholinesterase reactivators [PAM (pyridine aldoxime methiodide), PAC (pralidoxime, pyridine aldoxime methylchloride), TMB4 (trimedoxime), and DMO4 (obidoxime, Toxogonin, LüH6)] were compared in clinical trials. All of them proved capable of restoring erythrocyte cholinesterase activity and relieving symptoms and signs of organophosphate insecticide poisoning. Mildly and moderately poisoned patients can be treated by several injections of any one of these drugs alone, but severe cases need the synergistic action of atropine, as well as treatments for two to three consecutive days. Although response to treatment is stronger with TMB4 and DMO4, they are not recommended for routine treatment because of their dangerous adverse side effects.

  15. Evidence That GABA Mediates Dopaminergic and Serotonergic Pathways Associated with Locomotor Activity in Juvenile Chinook Salmon (Oncorhynchus tshawytscha)

    USGS Publications Warehouse

    Clements, S.; Schreck, C.B.


    The authors examined the control of locomotor activity in juvenile salmon (Oncorhynchus tshawytscha) by manipulating 3 neurotransmitter systems-gamma-amino-n-butyric acid (GABA), dopamine, and serotonin-as well as the neuropeptide corticotropin releasing hormone (CRH). Intracerebroventricular (ICV) injections of CRH and the GABAAagonist muscimol stimulated locomotor activity. The effect of muscimol was attenuated by administration of a dopamine receptor antagonist, haloperidol. Conversely, the administration of a dopamine uptake inhibitor (4???,4??? -difluoro-3-alpha-[diphenylmethoxy] tropane hydrochloride [DUI]) potentiated the effect of muscimol. They found no evidence that CRH-induced hyperactivity is mediated by dopaminergic systems following concurrent injections of haloperidol or DUI with CRH. Administration of muscimol either had no effect or attenuated the locomotor response to concurrent injections of CRH and fluoxetine, whereas the GABAA antagonist bicuculline methiodide potentiated the effect of CRH and fluoxetine.

  16. Clinical observation and comparison of the effectiveness of several oxime cholinesterase reactivators.


    Xue, S Z; Ding, X J; Ding, Y


    After passing toxicity and experimental therapeutic tests, four oxime cholinesterase reactivators [PAM (pyridine aldoxime methiodide), PAC (pralidoxime, pyridine aldoxime methylchloride), TMB4 (trimedoxime), and DMO4 (obidoxime, Toxogonin, LüH6)] were compared in clinical trials. All of them proved capable of restoring erythrocyte cholinesterase activity and relieving symptoms and signs of organophosphate insecticide poisoning. Mildly and moderately poisoned patients can be treated by several injections of any one of these drugs alone, but severe cases need the synergistic action of atropine, as well as treatments for two to three consecutive days. Although response to treatment is stronger with TMB4 and DMO4, they are not recommended for routine treatment because of their dangerous adverse side effects. PMID:3914075

  17. Effects of a 60 Hz magnetic field on central cholinergic systems of the rat

    SciTech Connect

    Lai, H.; Carino, M.A.; Horita, A.; Guy, A.W. )


    The authors studied the effects of an acute exposure to a 60 Hz magnetic field on sodium-dependent, high-affinity choline uptake in the brain of the rat. Decreases in uptake were observed in the frontal cortex and hippocampus after the animals were exposed to a magnetic field at flux densities [>=] 0.75 mT. These effects of the magnetic field were blocked by pretreating the animals with the narcotic antagonist naltrexone, but not by the peripheral opioid antagonist, naloxone methiodide. These data indicate that the magnetic-field-induced decreases in high-affinity choline uptake in the rat brain were mediated by endogenous opioids in the central nervous systems.

  18. Plasmodium falciparum: growth response to potassium channel blocking compounds.


    Waller, Karena L; Kim, Kami; McDonald, Thomas V


    Potassium channels are essential for cell survival and regulate the cell membrane potential and electrochemical gradient. During its lifecycle, Plasmodium falciparum parasites must rapidly adapt to dramatically variant ionic conditions within the mosquito mid-gut, the hepatocyte and red blood cell (RBC) cytosols, and the human circulatory system. To probe the participation of K(+) channels in parasite viability, growth response assays were performed in which asexual stage P. falciparum parasites were cultured in the presence of various Ca(2+)-activated K(+) channel blocking compounds. These data describe the novel anti-malarial effects of bicuculline methiodide and tubocurarine chloride and the novel lack of effect of apamine and verruculogen. Taken together, the data herein imply the presence of K(+) channels, or other parasite-specific targets, in P. falciparum-infected RBCs that are sensitive to blockade with Ca(2+)-activated K(+) channel blocking compounds. PMID:18703053

  19. A comparison of affinity constants for muscarine-sensitive acetylcholine receptors in guinea-pig atrial pacemaker cells at 29 degrees C and in ileum at 29 degrees C and 37 degrees C.

    PubMed Central

    Barlow, R B; Berry, K J; Glenton, P A; Nilolaou, N M; Soh, K S


    1 The affinity of 17 compounds for muscarine-sensitive acetylcholine receptors in atrial pacemaker cells and ileum of the guinea-pig has been measured at 29 degrees C in Ringer-Locke solution. Measurements were also made at 37 degrees C with 7 of them. 2 Some of the compounds had much higher affinity for the receptors in the ileum than for those in the atria. For the most selective compound, 4-diphenylacetoxy-N-methylpiperidine methiodide, the difference was approximately 20-fold. The receptors in the atria are therefore different the structure from those in the ileum. 3 The effect of temperature on affinity are not the same for all the compounds, tested indicating different enthalpies and entropies of adsorption and accounting for some of the difficulty experienced in predicting the affinity of new compounds. PMID:1000135

  20. Dorsomedial hypothalamic GABA regulates anxiety in the social interaction test.


    Shekhar, A; Katner, J S


    Blockade of GABAA function in the region of the dorsomedial hypothalamus (DMH) of rats is known to elicit a constellation of physiologic responses including increases in heart rate (HR), mean arterial blood pressure (BP), respiratory rate, and plasma catecholamine levels, as well as behavioral responses such as increases in locomotor activity and anxiogenic-like effects as measured in a conflict test and the elevated plus-maze test. The aim of the present study was to test the effects of microinjecting GABAA antagonists bicuculline methiodide (BMI) and picrotoxin, as well as the GABAA agonist muscimol, into the DMH of rats placed in the social interaction (SI) test. Muscimol decreased HR and BP but increased SI, whereas the GABA antagonists increased HR and BP but decreased SI time. Blocking the HR changes elicited by GABAergic drugs injected into the DMH with systemic injections of atenolol and atropine methylbromide did not block their effects on SI.

  1. Binding and functional properties of antimuscarinics of the hexocyclium/sila-hexocyclium and hexahydro-diphenidol/hexahydro-sila-diphenidol type to muscarinic receptor subtypes.

    PubMed Central

    Waelbroeck, M.; Tastenoy, M.; Camus, J.; Christophe, J.; Strohmann, C.; Linoh, H.; Zilch, H.; Tacke, R.; Mutschler, E.; Lambrecht, G.


    1. In an attempt to assess the structural requirements for the muscarinic receptor selectivity of hexahydro-diphenidol (hexahydro-difenidol) and hexahydro-sila-diphenidol (hexahydro-sila-difenidol), a series of structurally related C/Si pairs were investigated, along with atropine, pirenzepine and methoctramine, for their binding affinities in NB-OK 1 cells as well as in rat heart and pancreas. 2. The action of these antagonists at muscarinic receptors mediating negative inotropic responses in guinea-pig atria and ileal contractions has also been assessed. 3. Antagonist binding data indicated that NB-OK 1 cells (M1 type) as well as rat heart (cardiac type) and pancreas (glandular/smooth muscle type) possess different muscarinic receptor subtypes. 4. A highly significant correlation was found between the binding affinities of the antagonists to muscarinic receptors in rat heart and pancreas, respectively, and the affinities to muscarinic receptors in guinea-pig atria and ileum. This implies that the muscarinic binding sites in rat heart and the receptors in guinea-pig atria are essentially similar, but different from those in pancreas and ileum. 5. The antimuscarinic potency of hexahydro-diphenidol and hexahydro-sila-diphenidol at the three subtypes was influenced differently by structural modifications (e.g. quaternization). Different selectivity profiles for the antagonists were obtained, which makes these compounds useful tools to investigate further muscarinic receptor heterogeneity. Indeed, the tertiary analogues hexahydro-diphenidol (HHD) and hexahydro-sila-diphenidol (HHSiD) had an M1 = glandular/smooth muscle greater than cardiac selectivity profile, whereas the quaternary analogues HHD methiodide and HHSiD methiodide were M1 preferring (M1 greater than glandular/smooth muscle, cardiac). PMID:2804545

  2. A partial L5 spinal nerve ligation induces a limited prolongation of mechanical allodynia in rats: an efficient model for studying mechanisms of neuropathic pain.


    Guan, Yun; Yuan, Frank; Carteret, Alene F; Raja, Srinivasa N


    The relationship between pain severity and the extent of injury to a peripheral nerve remains elusive. In this study, we compared the pain behavior resulting from partial (1/3-1/2 thickness) and full L5 spinal nerve ligation (SNL) in rats. The decrease in paw withdrawal threshold (PWT) to mechanical stimuli in the hindpaw ipsilateral to the injury was comparable in the two groups on days 3-21 post-injury. However, the decreased PWT recovered earlier in the partial SNL group than in the full SNL group. These observations suggest that the duration of neuropathic pain behavior, but not the early development of mechanical allodynia, is dependent on the extent of nerve injury. On days 6 and 15 post-injury, when the mechanical allodynia was similar in the two groups, systemic morphine induced a greater reduction of mechanical allodynia in the partial SNL group than in the full SNL group. Furthermore, in partial SNL rats, at post-injury time points when they had largely recovered from the neuropathic pain state, systemic administration of naloxone hydrochloride (day 53) or naloxone methiodide (a non-selective peripherally acting opioid receptor antagonist; day 64) or intra-plantar injection of naloxone methiodide rekindled mechanical pain hypersensitivity in the ipsilateral hindpaw, suggesting a prolonged activation of endogenous opioidergic pain-inhibition. Therefore, partial SNL in rats may represent an efficient model for studying the mechanisms of neuropathic pain, testing effects of analgesic/antihyperalgesic drugs, and understanding endogenous pain-inhibitory mechanisms that lead to reversal of the pain behavior with time.

  3. In vivo imaging of epileptic foci in rats using a miniature probe integrating diffuse optical tomography and electroencephalographic source localization

    PubMed Central

    Yang, Hao; Zhang, Tao; Zhou, Junli; Carney, Paul R.; Jiang, Huabei


    SUMMARY Objective The goal of this work is to establish a new dual-modal brain mapping technique based on diffuse optical tomography (DOT) and electroencephalographic source localization (ESL) that can chronically/intracranially record optical/EEG data to precisely map seizures and localize the seizure onset zone and associated epileptic brain network. Methods The dual-modal imaging system was employed to image seizures in an experimental acute bicuculline methiodide rat model of focal epilepsy. Depth information derived from DOT was used as constraint in ESL to enhance the image reconstruction. Groups of animals were compared based on localization of seizure foci, either at different positions or at different depths. Results This novel imaging technique successfully localized the seizure onset zone in rat induced by bicuculline methiodide injected at a depth of 1mm, 2mm and 3mm, respectively. The results demonstrated that the incorporation of the depth information from DOT into the ESL image reconstruction resulted in more accurate and reliable ESL images. Although the ESL images showed a horizontal shift of the source localization, the DOT identified the seizure focus accurately. In one case, when the BMI was injected at a site outside the field of view (FOV) of the DOT/ESL interface, ESL gives false positive detection of the focus while DOT shows negative detection. Significance This study represents the first to identify seizure onset zone using implantable DOT. In addition, the combination of DOT/ESL has never been documented in neuroscience and epilepsy imaging. This technology will enable us to precisely measure the neural activity and hemodynamic response at exactly the same tissue site and at both cortical and sub cortical levels. PMID:25524046

  4. Hypoxia-induced hypothermia mediated by GABA in the rostral parapyramidal area of the medulla oblongata.


    Osaka, T


    Hypoxia evokes a regulated decrease in the body core temperature (Tc) in a variety of animals. The neuronal mechanisms of this response include, at least in part, glutamatergic activation in the lateral preoptic area (LPO) of the hypothalamus. As the sympathetic premotor neurons in the medulla oblongata constitute a cardinal relay station in the descending neuronal pathway from the hypothalamus for thermoregulation, their inhibition can also be critically involved in the mechanisms of the hypoxia-induced hypothermia. Here, I examined the hypothesis that hypoxia-induced hypothermia is mediated by glutamate-responsive neurons in the LPO that activate GABAergic transmission in the rostral raphe pallidus (rRPa) and neighboring parapyramidal region (PPy) of the medulla oblongata in urethane-chloralose-anesthetized, neuromuscularly blocked, artificially ventilated rats. Unilateral microinjection of GABA (15nmol) into the rRPa and PPy regions elicited a prompt increase in tail skin temperature (Ts) and decreases in Tc, oxygen consumption rate (VO2), and heart rate. Next, when the GABAA receptor blocker bicuculline methiodide (bicuculline methiodide (BMI), 10pmol) alone was microinjected into the rRPa, it elicited unexpected contradictory responses: simultaneous increases in Ts, VO2 and heart rate and a decrease in Tc. Then, when BMI was microinjected bilaterally into the PPy, no direct effect on Ts was seen; and thermogenic and tachycardic responses were slight. However, pretreatment of the PPy with BMI, but not vehicle saline, greatly attenuated the hypothermic responses evoked by hypoxic (10%O2-90%N2, 5min) ventilation or bilateral microinjections of glutamate (5nmol, each side) into the LPO. The results suggest that hypoxia-induced hypothermia was mediated, at least in part, by the activation of GABAA receptors in the PPy. PMID:24607346

  5. Modulation of the release of norepinephrine by gamma-aminobutyric acid and morphine in the frontal cerebral cortex of the rat

    SciTech Connect

    Peoples, R.W.


    Agents that enhance gamma-aminobutyric acid, or GABA, neurotransmission modulate certain effects of opioids, such as analgesia. Opioid analgesia is mediated in part by norepinephrine in the forebrain. In this study, the interactions between morphine and GABAergic agents on release of ({sup 3}H) norepinephrine from rat frontal cerebral cortical slices were examined. GABA, 5 {times} 10{sup {minus}5}-10{sup {minus}3} M, enhanced potassium stimulated ({sup 3}H) norepinephrine release and reversed the inhibitory effect of morphine in a noncompetitive manner. GABA did not enhance release of ({sup 3}H) norepinephrine stimulated by the calcium ionophore A23187. The effect of GABA was reduced by the GABA{sub A} receptor antagonists bicuculline methiodide or picrotoxin, and by the selective inhibitor of GABA uptake SKF 89976A, but was blocked completely only when bicuculline methiodide and SKF 89976A were used in combination. The GABA{sub A} agonist muscimol, 10{sup {minus}4} M, mimicked the effect of GABA, but the GABA{sub B} agonist ({plus minus})baclofen, 10{sup {minus}4} M, did not affect the release of ({sup 3}H) norepinephrine in the absence or the presence of morphine. Thus GABA appears to produce this effect by stimulating GABA uptake and GABA{sub A}, but not GABA{sub B}, receptors. In contrast to the results that would be predicted for an event involving GABA{sub A} receptors, however, the effect of GABA did not desensitize, and benzodiazepine agonists did not enhance the effect of GABA at any concentration tested between 10{sup {minus}8} and 10{sup {minus}4} M. Thus these receptors may constitute a subclass of GABA{sub A} receptors. These results support a role of GABA uptake and GABA{sub A} receptors in enhancing the release of norepinephrine and modulating its inhibition by opioids in the frontal cortex of the rat.

  6. Characterization of muscarinic receptors in rat kidney.


    Blankesteijn, W M; Siero, H L; Rodrigues de Miranda, J F; van Megen, Y J; Russel, F G


    Muscarinic receptors in mammalian kidney seem to be involved in diuresis. In this study we give a detailed characterization of receptors in rat kidney. Specific binding of [3H](-)-quinuclidinylbenzilate ([3H]QNB) to membranes of rat kidney cortex was saturable and of high affinity. A dissociation constant of 0.063 +/- 0.003 nM and a receptor density of 1.46 +/- 0.07 pmol/g wet weight were obtained. The dissociation kinetics could be best described by assuming a mono-exponential function (k-1 = (0.52 +/- 0.1) x 10(-4) s-1). The binding of [3H]QNB reached a maximum in 60 min at 0.6 nM at 37 degrees C. Competition experiments with the enantiomers of benzetimide confirmed the muscarinic nature of the [3H]QNB binding sites. The inhibition constants of pirenzepine (0.23 +/- 0.02 microM), (+-)-hexahydrosiladifenidol (0.040 +/- 0.002 microM), AF-DX 116 (1.45 +/- 0.07 microM), methoctramine (1.67 +/- 0.02 microM) and gallamine (78 +/- 3 microM) classified this receptor as an M3 receptor. Inhibition of [3H]QNB binding by the agonists methylfurtrethonium, arecoline, isoarecoline methiodide, arecaidine propargyl ester and McN-A-343 displayed monophasic inhibition curves. With (+/-)-cis-2-methyl-4-dimethylaminomethyl-1,3- dioxolane methiodide in two out of four experiments a small (11%) population of high affinity agonist sites could be detected. The potassium sparing diuretic amiloride inhibited [3H]QNB binding (36 +/- 3 microM). Although in a way related to the amiloride binding site, the muscarinic receptors in rat kidney are unlikely to be the primary target of diuretic action of this drug. PMID:8420789

  7. Nucleophilic Polymers and Gels in Hydrolytic Degradation of Chemical Warfare Agents.


    Bromberg, Lev; Creasy, William R; McGarvey, David J; Wilusz, Eugene; Hatton, T Alan


    Water- and solvent-soluble polymeric materials based on polyalkylamines modified with nucleophilic groups are introduced as catalysts of chemical warfare agent (CWA) hydrolysis. A comparative study conducted at constant pH and based on the criteria of the synthetic route simplicity, aqueous solubility, and rate of hydrolysis of CWA mimic, diisopropylfluorophosphate (DFP), indicated that 4-aminopyridine-substituted polyallylamine (PAAm-APy) and polyvinylamine substituted with 4-aminopyridine (PVAm-APy) were advantageous over 4-pyridinealdoxime-modified PVAm and PAAm, poly(butadiene-co-pyrrolidinopyridine), and PAAm modified with bipyridine and its complex with Cu(II). The synthesis of PVAm-APy and PAAm-APy involved generation of a betaine derivative of acrylamide and its covalent attachment onto the polyalkylamine chain followed by basic hydrolysis. Hydrogel particles of PAAm-APy and PVAm-APy cross-linked by epichlorohydrin exhibited pH-dependent swelling and ionization patterns that affected the rate constants of DFP nucleophilic hydrolysis. Deprotonation of the aminopyridine and amine groups increased the rates of the nucleophilic hydrolysis. The second-order rate of nucleophilic hydrolysis was 5.5- to 10-fold higher with the nucleophile-modified gels compared to those obtained by cross-linking of unmodified PAAm, throughout the pH range. Testing of VX and soman (GD) was conducted in 2.5-3.7 wt % PVAm-APy suspensions or gels swollen in water or DMSO/water mixtures. The half-lives of GD in aqueous PVAm-APy were 12 and 770 min at pH 8.5 and 5, respectively. Addition of VX into 3.5-3.7 wt % suspensions of PVAm-APy in DMSO-d6 and D2O at initial VX concentration of 0.2 vol % resulted in 100% VX degradation in less than 20 min. The unmodified PVAm and PAAm were 2 orders of magnitude less active than PVAm-APy and PAAm-APy, with VX half-lives in the range of 24 h. Furthermore, the PVAm-APy and PAAm-APy gels facilitated the dehydrochlorination reaction of sulfur mustard

  8. Nucleophilic Polymers and Gels in Hydrolytic Degradation of Chemical Warfare Agents.


    Bromberg, Lev; Creasy, William R; McGarvey, David J; Wilusz, Eugene; Hatton, T Alan


    Water- and solvent-soluble polymeric materials based on polyalkylamines modified with nucleophilic groups are introduced as catalysts of chemical warfare agent (CWA) hydrolysis. A comparative study conducted at constant pH and based on the criteria of the synthetic route simplicity, aqueous solubility, and rate of hydrolysis of CWA mimic, diisopropylfluorophosphate (DFP), indicated that 4-aminopyridine-substituted polyallylamine (PAAm-APy) and polyvinylamine substituted with 4-aminopyridine (PVAm-APy) were advantageous over 4-pyridinealdoxime-modified PVAm and PAAm, poly(butadiene-co-pyrrolidinopyridine), and PAAm modified with bipyridine and its complex with Cu(II). The synthesis of PVAm-APy and PAAm-APy involved generation of a betaine derivative of acrylamide and its covalent attachment onto the polyalkylamine chain followed by basic hydrolysis. Hydrogel particles of PAAm-APy and PVAm-APy cross-linked by epichlorohydrin exhibited pH-dependent swelling and ionization patterns that affected the rate constants of DFP nucleophilic hydrolysis. Deprotonation of the aminopyridine and amine groups increased the rates of the nucleophilic hydrolysis. The second-order rate of nucleophilic hydrolysis was 5.5- to 10-fold higher with the nucleophile-modified gels compared to those obtained by cross-linking of unmodified PAAm, throughout the pH range. Testing of VX and soman (GD) was conducted in 2.5-3.7 wt % PVAm-APy suspensions or gels swollen in water or DMSO/water mixtures. The half-lives of GD in aqueous PVAm-APy were 12 and 770 min at pH 8.5 and 5, respectively. Addition of VX into 3.5-3.7 wt % suspensions of PVAm-APy in DMSO-d6 and D2O at initial VX concentration of 0.2 vol % resulted in 100% VX degradation in less than 20 min. The unmodified PVAm and PAAm were 2 orders of magnitude less active than PVAm-APy and PAAm-APy, with VX half-lives in the range of 24 h. Furthermore, the PVAm-APy and PAAm-APy gels facilitated the dehydrochlorination reaction of sulfur mustard





    Tyramine methiodide and dopamine methobromide have greater pressor effect (three- to five-times) in the spinal cat than the parent amines. Noradrenaline methochloride has little pressor effect. Dopamine methobromide is about four times as potent as nicotine; tyramine methiodide is about equiactive to nicotine; and noradrenaline methochloride has only one-tenth the potency of nicotine. Their pressor effects are usually abolished by hexamethonium but in some experiments the effect of noradrenaline methochloride persisted and was then abolished by tolazoline. Injected intravenously into the cat anaesthetized with chloralose, each of the three quaternary derivatives contracts the nictitating membrane; dopamine methobromide is again the most active, having more than six times the potency of nicotine. When the contractions of the nictitating membrane are induced by continuous stimulation of the preganglionic fibres of the cervical sympathetic nerve, intravenous injection of the quaternary derivatives of tyramine and dopamine has a biphasic effect; there is a block on which a contraction of the membrane appears to be superimposed. Noradrenaline methochloride produces only a further contraction of the membrane. On the isolated rectus abdominis muscle preparation of the frog, dopamine methobromide is the most active in contracting the muscle, being about twelve times as active as nicotine; noradrenaline methochloride is weakest, having only one-hundredth the activity of nicotine. These effects are antagonized by hexamethonium. On the isolated phrenic nerve-diaphragm preparation of the rat, the quaternary derivatives of tyramine and dopamine each have neuromuscular blocking properties, 0.7- and 3-times respectively that of nicotine. Noradrenaline methochloride has no effect. In the sciatic nerve-tibialis preparation of the cat, the quaternary derivatives of tyramine and dopamine are approximately equipotent in producing neuromuscular paralysis, having about three times the

  10. Muscarinic receptor subtypes mediating the mucosal response to neural stimulation of guinea pig ileum

    SciTech Connect

    Carey, H.V.; Tien, X.Y.; Wallace, L.J.; Cooke, H.J.


    Muscarinic receptors involved in the secretory response evoked by electrical stimulation of submucosal neutrons were investigated in muscle-stripped flat sheets of guinea pig ileum set up in flux chambers. Neural stimulation produced a biphasic increase in short-circuit current due to active chloride secretion. Atropine and 4-diphenylacetoxy-N-methylpiperadine methiodide (4-DAMP) (10/sup -7/ M) were more potent inhibitors of the cholinergic phase of the response than was pirenzepine. Dose-dependent increases in base-line short-circuit current were evoked by carbachol and bethanechol; 4-hydroxy-2-butynyl trimethylammonium chloride (McN A343) produced a much smaller effect. Tetrodotoxin abolished the effects of McN A343 but did not alter the responses of carbachol and bethanechol. McN A343 significantly reduced the cholinergic phase of the neurally evoked response and caused a rightward shift of the carbachol dose-response curve. All muscarinic compounds inhibited (/sup 3/H)quinuclidinyl benzilate binding to membranes from muscosal scrapings, with a rank order of potency of 4-DAMP > pirenzepine > McN A343 > carbachol > bethanechol. These results suggest that acetylcholine released from submucosal neurons mediates chloride secretion by interacting with muscarinic cholinergic receptors that display a high binding affinity for 4-DAMP. Activation of neural muscarinic receptors makes a relatively small contribution to the overall secretory response.

  11. Kinetics of in vivo binding of antagonist to muscarinic cholinergic receptor in the human heart studied by Positron Emission Tomography

    SciTech Connect

    Syrota, A.; Paillotin, G.; Davy, J.M.; Aumont, M.C.


    Positron Emission Tomography (PET) was used to analyze in vivo antagonist binding to human myocardial muscarinic cholinergic receptor. The methiodide salt of the muscarinic antagonist, quinuclidinyl benzilate (MQNB), was labeled with the positron emitter, Carbon-11, and injected intravenously to 8 normal subjects. /sup 11/C-MQNB concentration was determined in vivo in the ventricular septum from 40 cross-sectional images acquired at the same transverse level over a period of 70 minutes. In 4 subjects, various amounts of unlabeled atropine were rapidly injected at 20 minutes to study whether atropine competitively inhibited MQNB. The kinetics of binding of /sup 11/C-MQNB were not the same in vivo and in vitro. The apparent dissociation rate of /sup 11/C-MQNB in vivo was much slower (by 1 to 2 orders of magnitude) than that observed in vitro with /sup 3/H-QNB. After atropine injection, /sup 11/C-MQNB dissociated from its binding sites at a rate that apparently depended on the amount of atropine present. /sup 11/C-MQNB kinetics were analyzed with a mathematical model which assumes the existence of a boundary layer containing free ligand in the vicinity of the binding sites. The dissociation rate of the radioligand depends on the probability of its rebinding to a free receptor site. 11 references, 1 table.

  12. Attenuation of pain-related behavior in a rat model of trigeminal neuropathic pain by viral-driven enkephalin overproduction in trigeminal ganglion neurons.


    Meunier, Alice; Latrémolière, Alban; Mauborgne, Annie; Bourgoin, Sylvie; Kayser, Valérie; Cesselin, François; Hamon, Michel; Pohl, Michel


    Trigeminal neuropathic pain represents a real challenge to therapy because commonly used drugs are devoid of real beneficial effect or patients frequently become intolerant or refractory to some of these compounds. In a rat model of trigeminal neuropathic pain, which shares numerous similarities with human trigeminal neuralgia and trigeminal neuropathic pain, we used a genomic herpes simplex virus-derived vector (HSVLatEnk) to examine the possible effect of a local overproduction of proenkephalin A (PA) targeted to the trigeminal primary sensory neurons. Unilateral peripheral inoculation of recombinant vectors on the vibrissal pad territory resulted in an about ninefold increase in proenkephalin A mRNA levels in trigeminal ganglion ipsilateral to the infected side. Transgene-derived met-enkephalin accumulated in numerous nerve cell bodies of trigeminal ganglion and was transported through the sensory nerve fibers located in the infraorbital nerve. Bilateral mechanical hyperresponsiveness, which developed 2 weeks after chronic constrictive injury of the left infraorbital nerve, was significantly attenuated in animals overproducing PA in the trigeminal ganglion ipsilateral to the lesioned infraorbital nerve. This antiallodynic effect was reversed by both the opioid receptor antagonist naloxone and the peripherally acting antagonist naloxone methiodide. Our data demonstrate that the local overproduction of PA-derived peptides in trigeminal ganglion sensory neurons evoked a potent antiallodynic effect through the stimulation of mainly peripherally located opioid receptors and suggest that targeted delivery of endogenous opioids may be of interest for the treatment of some severe forms of neuropathic pain. PMID:15771963

  13. Increased GABA(A) inhibition of the RVLM after hindlimb unloading in rats

    NASA Technical Reports Server (NTRS)

    Moffitt, Julia A.; Heesch, Cheryl M.; Hasser, Eileen M.


    Attenuated baroreflex-mediated increases in renal sympathetic nerve activity (RSNA) in hindlimb unloaded (HU) rats apparently are due to changes within the central nervous system. We hypothesized that GABA(A) receptor-mediated inhibition of the rostral ventrolateral medulla (RVLM) is increased after hindlimb unloading. Responses to bilateral microinjection of the GABA(A) antagonist (-)-bicuculline methiodide (BIC) into the RVLM were examined before and during caudal ventrolateral medulla (CVLM) inhibition in Inactin-anesthetized control and HU rats. Increases in mean arterial pressure (MAP), heart rate (HR), and RSNA in response to BIC in the RVLM were significantly enhanced in HU rats. Responses to bilateral CVLM blockade were not different. When remaining GABA(A) inhibition in the RVLM was blocked by BIC during CVLM inhibition, the additional increases in MAP and RSNA were significantly greater in HU rats. These data indicate that GABA(A) receptor-mediated inhibition of RVLM neurons is augmented after hindlimb unloading. Effects of input from the CVLM were unaltered. Thus, after cardiovascular deconditioning in rodents, the attenuated increase in sympathetic nerve activity in response to hypotension is associated with greater GABA(A) receptor-mediated inhibition of RVLM neurons originating at least in part from sources other than the CVLM.



    Avdiyuk, K V; Varbanets, L D


    The effect of cations and anions on the activity of Aspergillus flavus var. oryzae and Bacillus subtilis α-amylases showed that the tested enzymes are sensitive to most of cations and resistant to anions. The most significant inhibitory effects on the activity of A. flavus var. oryzae α-amylase have been demonstrated by Al3+ and Fe3+ ions, while on the activity of B. subtilis α-amylase - Hg2+, Cu2+ and Fe3+ ions. Inactivation of A. flavus var. oryzae and B. subtilis α-amylases in the presence of EGTA is indicated on the presence within their structure of metal ions. An important role in the enzymatic catalysis of both enzymes play carboxyl groups as evidenced by their inhibition of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide methiodide. Inhibition of B. subtilis α-amylase by p-chloromercuribenzoate, N-ethylmaleimide and sodium sulfite is indicated on the probable involvement of the sulfhydryl groups in the functioning of the enzyme. Unlike most studied glycosidases the tested enzymes do not contain histidine imidazole group in the active center.

  15. Antitussive activity of Withania somnifera and opioid receptors.


    Nosálová, Gabriela; Sivová, Veronika; Ray, Bimalendu; Fraňová, Soňa; Ondrejka, Igor; Flešková, Dana


    Arabinogalactan is a polysaccharide isolated from the roots of the medicinal plant Withania somnifera L. It contains 65% arabinose and 18% galactose. The aim of the present study was to evaluate the antitussive activity of arabinogalactan in conscious, healthy adult guinea pigs and the role of the opioid pathway in the antitussive action. A polysaccharide extract was given orally in a dose of 50 mg/kg. Cough was induced by an aerosol of citric acid in a concentration 0.3 mol/L, generated by a jet nebulizer into a plethysmographic chamber. The intensity of cough response was defined as the number of cough efforts counted during a 3-min exposure to the aerosol. The major finding was that arabinogalactan clearly suppressed the cough reflex; the suppression was comparable with that of codeine that was taken as a reference drug. The involvement of the opioid system was tested with the use of a blood-brain barrier penetrable, naloxone hydrochloride, and non-penetrable, naloxone methiodide, to distinguish between the central and peripheral mu-opioid receptor pathways. Both opioid antagonists acted to reverse the arabinogalactan-induced cough suppression; the reversion was total over time with the latter antagonist. We failed to confirm the presence of a bronchodilating effect of the polysaccharide, which could be involved in its antitussive action. We conclude that the polysaccharide arabinogalactan from Withania somnifera has a distinct antitussive activity consisting of cough suppression and that this action involves the mu-opioid receptor pathways.



    Avdiyuk, K V; Varbanets, L D


    The effect of cations and anions on the activity of Aspergillus flavus var. oryzae and Bacillus subtilis α-amylases showed that the tested enzymes are sensitive to most of cations and resistant to anions. The most significant inhibitory effects on the activity of A. flavus var. oryzae α-amylase have been demonstrated by Al3+ and Fe3+ ions, while on the activity of B. subtilis α-amylase - Hg2+, Cu2+ and Fe3+ ions. Inactivation of A. flavus var. oryzae and B. subtilis α-amylases in the presence of EGTA is indicated on the presence within their structure of metal ions. An important role in the enzymatic catalysis of both enzymes play carboxyl groups as evidenced by their inhibition of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide methiodide. Inhibition of B. subtilis α-amylase by p-chloromercuribenzoate, N-ethylmaleimide and sodium sulfite is indicated on the probable involvement of the sulfhydryl groups in the functioning of the enzyme. Unlike most studied glycosidases the tested enzymes do not contain histidine imidazole group in the active center. PMID:26422920

  17. Regulation of sup 35 S-TBPS binding by bicuculline is region specific in rat brain

    SciTech Connect

    Peris, J.; Shawley, A.; Dawson, R.; Abendschein, K.H. )


    The allosteric regulation of specific {sup 35}S-TBPS binding to the convulsant site on the GABA{sub A} receptor/chloride (Cl{sup {minus}}) ionophore complex was studied in various brain regions in an attempt to characterize regional heterogeneity of the protein subunits forming the complex. Bicuculline methiodide (BIC), a GABA{sub A} antagonist, enhanced binding in cortex (CTX), substantia nigra (SN) and cerebellum (CBL), inhibited binding in inferior colliculus (IC) and did not affect binding in superior colliculus (SC). Similar results were found in CBL and IC using SR-95531, another GABA{sub A} antagonist. The levels of endogenous GABA in the different tissue samples could not account for the regional differences in binding. When the functional regulation of these receptors was measured using {sup 36}Cl{sup {minus}} uptake in microsomes, muscimol-stimulated uptake was completely blocked by BIC in CBL and IC but was not affected by BIC in SC. Additionally, picrotoxin completely blocked muscimol-stimulated uptake in CBL but had no effect in IC or SC. These findings provide a functional basis for regional heterogeneity of GABA{sub A} receptor.

  18. Intracerebroventricular injection of mu- and delta-opiate receptor antagonists block 60 Hz magnetic field-induced decreases in cholinergic activity in the frontal cortex and hippocampus of the rat.


    Lai, H; Carino, M


    In previous research, we have found that acute exposure to a 60 Hz magnetic field decreased cholinergic activity in the frontal cortex and hippocampus of the rat as measured by sodium-dependent high-affinity choline uptake activity. We concluded that the effect was mediated by endogenous opioids inside the brain because it could be blocked by pretreatment of rats before magnetic field exposure with the opiate antagonist naltrexone, but not by the peripheral antagonist naloxone methiodide. In the present study, the involvement of opiate receptor subtypes was investigated. Rats were pretreated by intracerebroventricular injection of the mu-opiate receptor antagonist, beta-funaltrexamine, or the delta-opiate receptor antagonist, naltrindole, before exposure to a 60 Hz magnetic field (2 mT, 1 hour). It was found that the effects of magnetic field on high-affinity choline uptake in the frontal cortex and hippocampus were blocked by the drug treatments. These data indicate that both mu- and delta-opiate receptors in the brain are involved in the magnetic field-induced decreases in cholinergic activity in the frontal cortex and hippocampus of the rat.

  19. Transient and reversible parkinsonism after acute organophosphate poisoning.


    Arima, Hajime; Sobue, Kazuya; So, MinHye; Morishima, Tetsuro; Ando, Hirkoshi; Katsuya, Hirotada


    Parkinsonism is a rare complication in patients with organophosphate poisoning. To date there have been two cases of transient parkinsonism after acute and severe cholinergic crisis, both of which were successfully treated using amantadine, an anti-parkinsonism drug. We report on an 81-year-old woman who was admitted for the treatment of acute severe organophosphate poisoning. Although acute cholinergic crisis was treated successfully with large doses of atropine and 2-pyridine aldoxime methiodide (PAM), extrapyramidal manifestations were noticed on hospital day 6. The neurological symptoms worsened, and the diagnosis of parkinsonism was made by a neurologist on hospital day 9. Immediately, biperiden (5mg), an anti-parkinsonism drug, was administered intravenously, and her symptoms markedly improved. From the following day, biperiden (5 mg/day) was given intramuscularly for eight days. Subsequently, neurological symptoms did not relapse, and no drugs were required. Our patient is the third case of parkinsonism developing after an acute severe cholinergic crisis and the first case successfully treated with biperiden. Patients should be carefully observed for the presence of neurological signs in this kind of poisoning. If present, an anti-parkinsonism drug should be considered.

  20. Across-frequency nonlinear inhibition by GABA in processing of interaural time difference.


    Mori, K


    The barn owl uses the interaural time difference (ITD) to determine the azimuth of a sound source. Narrowband ITD-sensitive neurons cannot distinguish a given ITD from those that produce the same interaural phase difference (phase ambiguity). Neurons in the external nucleus of the inferior colliculus (ICx) resolve the ambiguity by gathering ITD information across many frequencies, thereby suppressing false responses (side peaks, SP) relative to the true ITD (the main peak, MP) in a response versus ITD curve. This process was quantitatively studied by comparing the ITD curve for a pair of tones presented simultaneously (two-tone curve) to the simple sum (predicted curve) of the individual ITD curves for the same tones presented separately. Sixteen of the 39 neurons tested did not show a significant difference in MP and SP responses between these curves (category I); 14 neurons showed significant SP suppression (category II). During iontophoretic application of bicuculline methiodide, a GABA(A) antagonist, most (n = 7/8) category II neurons lost nonlinear SP suppression and became linear, whereas category I neurons retained linear summation (n = 3/3). Thus, the nonlinear cross-frequency interaction of ITD responses in ICx neurons was mediated mostly by GABAergic inhibition, which enhanced SP suppression, and helped resolve phase ambiguity. PMID:9307308

  1. Recovery of brain and plasma cholinesterase activities in ducklings exposed to organophosphorus pesticides

    USGS Publications Warehouse

    Fleming, W.J.


    Brain and plasma cholinesterase (ChE) activities were determined for mallard ducklings (Anas platyrhynchos) exposed to dicrotophos and fenthion. Recovery rates of brain ChE did not differ between ducklings administered a single oral dose vs. a 2-week dietary dose of these organophosphates. Exposure to the organophosphates, followed by recovery of brain ChE, did not significantly affect the degree of brain ChE inhibition or the recovery of ChE activity at a subsequent exposure. Recovery of brain ChE activity followed the general model Y = a + b(logX) with rapid recovery to about 50% of normal, followed by a slower rate of recovery until normal ChE activity levels were attained. Fenthion and dicrotophos-inhibited brain ChE were only slightly reactivated in vitro by pyridine-2-aldoxime methiodide, which suggested that spontaneous reactivation was not a primary method of recovery of ChE activity. Recovery of brain ChE activity can be modeled for interpretation of sublethal inhibition of brain ChE activities in wild birds following environmental applications of organophosphates. Plasma ChE activity is inferior to brain ChE activity for environmental monitoring, because of its rapid recovery and large degree of variation among individuals.

  2. Differential effects of petit mal anticonvulsants and convulsants on thalamic neurones: GABA current blockade.

    PubMed Central

    Coulter, D. A.; Huguenard, J. R.; Prince, D. A.


    1. Currents evoked by applications of gamma-aminobutyric acid (GABA) to acutely dissociated thalamic neurones were analysed by voltage-clamp techniques, and the effects of the anticonvulsant succinimides ethosuximide (ES) and alpha-methyl-alpha-phenylsuccinimide (MPS) and the convulsants tetramethylsuccinimide (TMS), picrotoxin, pentylenetetrazol (PTZ), and bicuculline methiodide were assessed. 2. TMS (1 microM-10 microM) reduced responses to iontophoretically applied GABA, as did picrotoxin (0.1-100 microM), PTZ (1-100 mM) and bicuculline (1-100 microM). 3. ES, in high concentrations (1-10 mM), reduced GABA responses to a lesser extent, and also occluded the reductions in GABA-evoked currents produced by TMS, picrotoxin, and PTZ. ES did not occlude the effects of bicuculline on GABA responses. Therefore, we propose that ES acts as a partial agonist at the picrotoxin GABA-blocking receptor. 4. MPS had no effect on GABA responses (at a concentration of 1 mM), and, like ES, occluded the GABA-blocking actions of TMS, apparently acting as a full antagonist. 5. The anticonvulsant actions of ES and MPS against TMS and PTZ-induced seizures may thus involve two independent mechanisms: (1) the occlusion of TMS and PTZ GABA-blocking effects; and (2) the previously described specific effect of ES and MPS on low-threshold calcium current of thalamic neurones. The latter cellular mechanism may be more closely related to petit mal anticonvulsant activity. PMID:2119843

  3. Local GABA receptor blockade reveals hindlimb responses in the SI forelimb-stump representation of neonatally amputated rats.


    Pluto, Charles P; Lane, Richard D; Rhoades, Robert W


    In adult rats that sustained forelimb amputation on the day of birth, there are numerous multi-unit recording sites in the forelimb-stump representation of primary somatosensory cortex (SI) that also respond to cutaneous stimulation of the hindlimb when cortical receptors for GABA are blocked. These normally suppressed hindlimb inputs originate in the SI hindlimb representation and synapse in the dysgranular cortex before exciting SI forelimb-stump neurons. In our previous studies, GABA (A + B) receptor blockade was achieved by topically applying a bicuculline methiodide/saclofen solution (BMI/SAC) to the cortical surface. This treatment blocks receptors throughout SI and does not allow determination of where along the above circuit the GABA-mediated suppression of hindlimb information occurs. In this study, focal injections of BMI/SAC were delivered to three distinct cortical regions that are involved in the hindlimb-to-forelimb-stump pathway. Blocking GABA receptors in the SI hindlimb representation and in the dysgranular cortex was largely ineffective in revealing hindlimb inputs ( approximately 10% of hindlimb inputs were revealed in both cases). In contrast, when the blockade was targeted at forelimb-stump recording sites, >80% of hindlimb inputs were revealed. Thus GABAergic interneurons within the forelimb-stump representation suppress the expression of reorganized hindlimb inputs to the region. A circuit model incorporating these and previous observations is presented and discussed.

  4. The effect of boric acid on acethylcholine, bethanechol and potasssium-evoked responses on ileum of rat.


    Ince, S; Turkmen, R; Yavuz, H


    1 The aim of this study was to clarify the effect of boric acid on contractions of rat isolated ileum. 2 Contractile responses expressed as Emax and pD2 for acetylcholine (10(-3)-10(-8) m, Ach), bethanechol (10(-3)-10(-8) m) and potassium (10-80 × 10(-3) m, KCl) were determined in the absence and presence of boric acid (10(-3); 5 × 10(-4); 10(-4) m). 3 The contractile response to Ach in the presence of verapamil (10(-6) or 10(-8) m) or in calcium-free Tyrode's solution was also determined in the absence and presence of boric acid. 4 Boric acid did not affect the contractile response to Ach, bethanechol or KCl. Single or cumulative treatment of boric acid did not affect ileum muscle contraction evoked by KCl. The atropine-resistant component of Ach-induced contraction and 4-diphenyl-acetoxy-N-methyl-piperidine methiodide-resistant component of bethanechol-induced contraction were not inhibited by boric acid (10(-3) m). The contractile response to Ach was reduced in calcium-free Tyrode's solution, and the contractile response was not affected by (10(-8) m). The addition of boric acid (10(-3) m) in combination with verapamil (10(-8) m) did not significantly affect the contractile response to Ach. 5 In conclusion, boric acid does not affect contractions induced by Ach, bethanechol or potassium in rat isolated ileum.

  5. Intraplantar injection of linalool reduces paclitaxel-induced acute pain in mice.


    Katsuyama, Soh; Kuwahata, Hikari; Yagi, Tomomi; Kishikawa, Yukinaga; Komatsu, Takaaki; Sakurada, Tsukasa; Nakamura, Hitoshi


    Linalool is the principal component of many essential oils known to possess biological activities. We previously reported that intraplantar injection of linalool reduces the nociceptive response as assayed by the capsaicin test. In this study, we sought to determine whether intraplantar injection of linalool could influence the induction of acute pain (allodynia and hyperalgesia) by paclitaxel in mice. Paclitaxel is widely used in cancer chemotherapy for the treatment of solid tumors, but it sometimes induces moderate to severe acute pain. Paclitaxel administered intraperitoneally as a single dose of 5, 10 or 20 mg/kg produced mechanical allodynia and hyperalgesia in mice. Paclitaxel-induced mechanical allodynia and hyperalgesia began 1 day after administration of paclitaxel and resolved within 7 days. Linalool injected into the hindpaw caused a significant reduction in paclitaxel-induced mechanical allodynia and hyperalgesia. Pretreatment with naloxone hydrochloride, an opioid receptor antagonist, or naloxone methiodide, a peripherally acting µ-opioid receptor-preferring antagonist, significantly reversed linalool-induced antiallodynia and antihyperalgesia. Our results provide evidence for the involvement of peripheral opioids in antiallodynia and antihyperalgesia induced by linalool. These results suggest that activation of peripheral opioid receptors may play an important role in reducing paclitaxel-induced mechanical allodynia and hyperalgesia.

  6. Effect of plantar subcutaneous administration of bergamot essential oil and linalool on formalin-induced nociceptive behavior in mice.


    Katsuyama, Soh; Otowa, Akira; Kamio, Satomi; Sato, Kazuma; Yagi, Tomomi; Kishikawa, Yukinaga; Komatsu, Takaaki; Bagetta, Giacinto; Sakurada, Tsukasa; Nakamura, Hitoshi


    This study investigated the effect of bergamot essential oil (BEO) or linalool, a major volatile component of BEO, on the nociceptive response to formalin. Plantar subcutaneous injection of BEO or linalool into the ipsilateral hindpaw reduced both the first and late phases of the formalin-induced licking and biting responses in mice. Plantar subcutaneous injection of BEO or linalool into the contralateral hindpaw did not yield an antinociceptive effect, suggesting that the antinociceptive effect of BEO or linalool in the formalin test occurred peripherally. Intraperitoneal and plantar subcutaneous injection pretreatment with naloxone hydrochloride, an opioid receptor antagonist, significantly attenuated both BEO- and linalool-induced antinociception. Pretreatment with naloxone methiodide, a peripherally acting opioid receptor antagonists, also significantly antagonized the antinociceptive effects of BEO and linalool. Our results provide evidence for the involvement of peripheral opioids in antinociception induced by BEO and linalool. These results suggest that activation of peripheral opioid receptors may play an important role in reducing formalin-induced nociception.

  7. JAK-STAT1/3-induced expression of signal sequence-encoding proopiomelanocortin mRNA in lymphocytes reduces inflammatory pain in rats

    PubMed Central


    Background Proopiomelanocortin (POMC)-derived beta-endorphin1-31 from immune cells can inhibit inflammatory pain. Here we investigated cytokine signaling pathways regulating POMC gene expression and beta-endorphin production in lymphocytes to augment such analgesic effects. Results Interleukin-4 dose-dependently elevated POMC mRNA expression in naïve lymph node-derived cells in vitro, as determined by real-time PCR. This effect was neutralized by janus kinase (JAK) inhibitors. Transfection of Signal Transducer and Activator of Transcription (STAT) 1/3 but not of STAT6 decoy oligonucleotides abolished interleukin-4 induced POMC gene expression. STAT3 was phosphorylated in in vitro interleukin-4 stimulated lymphocytes and in lymph nodes draining inflamed paws in vivo. Cellular beta-endorphin increased after combined stimulation with interleukin-4 and concanavalin A. Consistently, in vivo reduction of inflammatory pain by passively transferred T cells improved significantly when donor cells were pretreated with interleukin-4 plus concanavalin A. This effect was blocked by naloxone-methiodide. Conclusion Interleukin-4 can amplify endogenous opioid peptide expression mediated by JAK-STAT1/3 activation in mitogen-activated lymphocytes. Transfer of these cells leads to inhibition of inflammatory pain via activation of peripheral opioid receptors. PMID:23146666

  8. Tonic inhibitory control exerted by opioid peptides in the paraventricular nuclei of the hypothalamus on regional hemodynamic activity in rats.


    Lessard, Andrée; Bachelard, Hélène


    1. Systemic and regional cardiovascular changes were measured following bilateral microinjection of specific and selective opioid-receptor antagonists into the paraventricular nuclei of the hypothalamus (PVN) of awake, freely moving rats. 2. PVN microinjection of increasing doses of the specific opioid antagonist naloxone - methiodide (1 - 5.0 nmol), or a selective mu-opioid receptor antagonist, beta-funaltrexamine (0.05 - 0.5 nmol), evoked important cardiovascular changes characterized by small and transient increases in heart rate (HR) and mean arterial pressure (MAP), vasoconstriction in renal and superior mesenteric vascular beds and vasodilation in the hindquarter vascular bed. 3. No significant cardiovascular changes were observed following PVN administration of the highly selective delta-opioid-receptor antagonist, ICI 174864 (0.1 - 1 nmol), or the selective kappa-opioid-receptor antagonist, nor-binaltorphine (0.1 - 1 nmol). 4. Most of the cardiovascular responses to naloxone (3 nmol) and beta-funaltrexamine (0.5 nmol) were attenuated or abolished by an i.v. treatment with a specific vasopressin V(1) receptor antagonist. 5. These results suggest that endogenous opioid peptides and mu-type PVN opioid receptors modulate a tonically-active central depressor pathway acting on systemic and regional haemodynamic systems. Part of this influence could involve a tonic inhibition of vasopressin release.

  9. Tonic inhibitory control exerted by opioid peptides in the paraventricular nuclei of the hypothalamus on regional hemodynamic activity in rats

    PubMed Central

    Lessard, Andrée; Bachelard, Hélène


    Systemic and regional cardiovascular changes were measured following bilateral microinjection of specific and selective opioid-receptor antagonists into the paraventricular nuclei of the hypothalamus (PVN) of awake, freely moving rats.PVN microinjection of increasing doses of the specific opioid antagonist naloxone – methiodide (1 – 5.0 nmol), or a selective μ-opioid receptor antagonist, β-funaltrexamine (0.05 – 0.5 nmol), evoked important cardiovascular changes characterized by small and transient increases in heart rate (HR) and mean arterial pressure (MAP), vasoconstriction in renal and superior mesenteric vascular beds and vasodilation in the hindquarter vascular bed.No significant cardiovascular changes were observed following PVN administration of the highly selective δ-opioid-receptor antagonist, ICI 174864 (0.1 – 1 nmol), or the selective κ-opioid-receptor antagonist, nor-binaltorphine (0.1 – 1 nmol).Most of the cardiovascular responses to naloxone (3 nmol) and β-funaltrexamine (0.5 nmol) were attenuated or abolished by an i.v. treatment with a specific vasopressin V1 receptor antagonist.These results suggest that endogenous opioid peptides and μ-type PVN opioid receptors modulate a tonically-active central depressor pathway acting on systemic and regional haemodynamic systems. Part of this influence could involve a tonic inhibition of vasopressin release. PMID:12086985

  10. Role of central and peripheral opiate receptors in the effects of fentanyl on analgesia, ventilation and arterial blood-gas chemistry in conscious rats

    PubMed Central

    Henderson, Fraser; May, Walter J.; Gruber, Ryan B.; Discala, Joseph F.; Puscovic, Veljko; Young, Alex P.; Baby, Santhosh M.; Lewis, Stephen J.


    This study determined the effects of the peripherally restricted µ-opiate receptor (µ-OR) antagonist, naloxone methiodide (NLXmi) on fentanyl (25 µg/kg, i.v.)-induced changes in (1) analgesia, (2) arterial blood gas chemistry (ABG) and alveolar-arterial gradient (A-a gradient), and (3) ventilatory parameters, in conscious rats. The fentanyl-induced increase in analgesia was minimally affected by a 1.5 mg/kg of NLXmi but was attenuated by a 5.0 mg/kg dose. Fentanyl decreased arterial blood pH, pO2 and sO2 and increased pCO2 and A-a gradient. These responses were markedly diminished in NLXmi (1.5 mg/kg)-pretreated rats. Fentanyl caused ventilatory depression (e.g., decreases in tidal volume and peak inspiratory flow). Pretreatment with NLXmi (1.5 mg/kg, i.v.) antagonized the fentanyl decrease in tidal volume but minimally affected the other responses. These findings suggest that (1) the analgesia and ventilatory depression caused by fentanyl involve peripheral µ-ORs and (2) NLXmi prevents the fentanyl effects on ABG by blocking the negative actions of the opioid on tidal volume and A-a gradient. PMID:24284037

  11. Respiratory, metabolic and cardiac functions are altered by disinhibition of subregions of the medial prefrontal cortex

    PubMed Central

    Hassan, Sarah F; Cornish, Jennifer L; Goodchild, Ann K


    The prefrontal cortex (PFC) is referred to as the visceral motor cortex; however, little is known about whether this region influences respiratory or metabolic outflows. The aim of this study was to describe simultaneous changes in respiratory, metabolic and cardiovascular functions evoked by disinhibition of the medial PFC (mPFC) and adjacent lateral septal nucleus (LSN). In urethane-anaesthetized rats, bicuculline methiodide was microinjected (2 mm; GABA-A receptor antagonist) into 90 sites in the mPFC at 0.72–4.00 mm from bregma. Phrenic nerve amplitude and frequency, arterial pressure, heart rate, splanchnic and lumbar sympathetic nerve activities (SNA), expired CO2, and core and brown adipose tissue temperatures were measured. Novel findings included disturbances to respiratory rhythm evoked from all subregions of the mPFC. Injections into the cingulate cortex evoked reductions in central respiratory function exclusively, whereas in ventral sites, particularly the infralimbic region, increases in respiratory drive and frequency, and metabolic and cardiac outflows were evoked. Disinhibition of sites in surrounding regions revealed that the LSN could evoke cardiovascular changes accompanied by distinct oscillations in SNA, as well as increases in respiratory amplitude. We show that activation of neurons within the mPFC and LSN influence respiratory, metabolic and cardiac outflows in a site-dependent manner. This study has implications with respect to the altered PFC neuronal activity seen in stress-related and mental health disorders, and suggests how basic physiological systems may be affected. PMID:24042503

  12. Physiology and pharmacology of acetylcholinergic responses of interneurons in the antennal lobes of the moth Manduca sexta.


    Waldrop, B; Hildebrand, J G


    1. Neurons in the antennal lobe (AL) of the moth Manduca sexta respond to the application, via pressure injection into the neuropil, of acetylcholine (ACh). When synaptic transmission is not blocked, both excitatory (Fig. 2) and inhibitory (Fig. 3) responses are seen. 2. Responses to ACh appear to be receptor-mediated, as they are associated with an increase in input conductance (Figs. 2B and 3B) and are dose-dependent (Fig. 2 C). 3. All neurons responsive to ACh are also excited by nicotine. Responses to nicotine are stronger and more prolonged than responses to ACh (Fig. 4C). No responses are observed to the muscarinic agonist, oxotremorine (Fig. 4 B). 4. Curare blocks responses of AL neurons to applied ACh, while atropine and dexetimide are only weakly effective at reducing ACh responses (Figs. 5 and 6). 5. Curare is also more effective than atropine or dexetimide at reducing synaptically-mediated responses of AL neurons (Fig. 7). 6. In one AL neuron, bicuculline methiodide (BMI) blocked the IPSP produced by electrical stimulation of the antennal nerve, but it did not reduce the inhibitory response to application of ACh (Fig. 8). PMID:2926690

  13. The pharmacology of spontaneously open alpha 1 beta 3 epsilon GABA A receptor-ionophores.


    Maksay, Gábor; Thompson, Sally A; Wafford, Keith A


    Human alpha(1)beta(3) epsilon GABA(A) receptors were expressed in Xenopus oocytes and examined using the conventional two-electrode voltage-clamp technique and compared to alpha(1)beta(3)gamma(2) receptors. The effects of several GABA(A) agonists were studied, and the allosteric modulation of the channel by a number of GABAergic modulators investigated. The presence of the epsilon subunit increased the potency and efficacy of direct activation by partial GABA(A) agonists (piperidine-4-sulphonic acid and thio-4-PIOL), pentobarbital and neuro-steroids. Direct activation by 3-hydroxylated neurosteroids was restricted to 3alpha epimers, while chirality at C5 was indifferent. The 3beta-sulfate esters of pregnenolone and dehydroepiandrosterone inhibited the spontaneous currents with efficacies higher, while bicuculline methiodide and SR 95531 did so lower than picrotoxin and TBPS. Furosemide, fipronil, triphenylcyanoborate and Zn(2+) blocked the spontaneous currents of alpha(1)beta(3) epsilon receptors with different efficacies. Flunitrazepam and 4'-chlorodiazepam inhibited the spontaneous currents with micromolar potencies. In conclusion, spontaneously active alpha(1)beta(3) epsilon GABA(A) receptors can be potentiated and blocked by GABAergic agents within a broad range of efficacy.

  14. Human embryonic stem cell-derived neuronal cells form spontaneously active neuronal networks in vitro.


    Heikkilä, Teemu J; Ylä-Outinen, Laura; Tanskanen, Jarno M A; Lappalainen, Riikka S; Skottman, Heli; Suuronen, Riitta; Mikkonen, Jarno E; Hyttinen, Jari A K; Narkilahti, Susanna


    The production of functional human embryonic stem cell (hESC)-derived neuronal cells is critical for the application of hESCs in treating neurodegenerative disorders. To study the potential functionality of hESC-derived neurons, we cultured and monitored the development of hESC-derived neuronal networks on microelectrode arrays. Immunocytochemical studies revealed that these networks were positive for the neuronal marker proteins beta-tubulin(III) and microtubule-associated protein 2 (MAP-2). The hESC-derived neuronal networks were spontaneously active and exhibited a multitude of electrical impulse firing patterns. Synchronous bursts of electrical activity similar to those reported for hippocampal neurons and rodent embryonic stem cell-derived neuronal networks were recorded from the differentiated cultures until up to 4 months. The dependence of the observed neuronal network activity on sodium ion channels was examined using tetrodotoxin (TTX). Antagonists for the glutamate receptors NMDA [D(-)-2-amino-5-phosphonopentanoic acid] and AMPA/kainate [6-cyano-7-nitroquinoxaline-2,3-dione], and for GABAA receptors [(-)-bicuculline methiodide] modulated the spontaneous electrical activity, indicating that pharmacologically susceptible neuronal networks with functional synapses had been generated. The findings indicate that hESC-derived neuronal cells can generate spontaneously active networks with synchronous communication in vitro, and are therefore suitable for use in developmental and drug screening studies, as well as for regenerative medicine.

  15. Photomagnetism in cyano-bridged bimetal assemblies.


    Ohkoshi, Shin-ichi; Tokoro, Hiroko


    The study of photoinduced phase-transition materials has implications for the fields of inorganic chemistry, solid-state chemistry, and materials science. Cyano-bridged bimetal assemblies are promising photomagnetic materials. Because cyano-bridged bimetal assemblies possess various absorption bands in the visible light region, their electronic and spin states can be controlled by visible light irradiation. Moreover, the selection of magnetic metal ions and organic ligands provide a way of controlling spin-spin interactions through a cyano bridge. In this Account, we describe cyano-bridged bimetal assemblies developed in our laboratory. Cu(II)(2)[Mo(IV)(CN)(8)]·8H(2)O (CuMo), Rb(I)Mn(II)[Fe(III)(CN)(6)] (RbMnFe), and Co(II)(3)[W(V)(CN)(8)](2)·(pyrimidine)(4)·6H(2)O (CoW) induce photomagnetism via photoinduced metal-to-metal charge transfers (MM'CT), while Fe(II)(2)[Nb(IV)(CN)(8)]·(4-pyridinealdoxime)(8)·2H(2)O (FeNb) exhibits a photoinduced magnetization via a photoinduced spin crossover. Irradiation with 473 nm light causes the CuMo system to exhibit a spontaneous magnetization with a Curie temperature (T(C)) of 25 K, but irradiation with 532, 785, and 840 nm light reduces the magnetization. In this reversible photomagnetic process, excitation of the MM'CT from Mo(IV) to Cu(II) produces a ferromagnetic mixed-valence isomer of Cu(I)Cu(II)[Mo(V)(CN)(8)]·8H(2)O (CuMo'). CuMo' returns to CuMo upon irradiation in the reverse-M'MCT band. RbMnFe shows a charge transfer (CT)-induced phase transition from the Mn(II)-Fe(III) phase to the Mn(III)-Fe(II) phase. Irradiation with 532 nm light converts the Mn(III)-Fe(II) phase into the Mn(II)-Fe(III) phase, and we observe photodemagnetization. In contrast, irradiation of the Mn(II)-Fe(III) phase with 410 nm light causes the reverse phase transition. A CT-induced Jahn-Teller distortion is responsible for this visible light-induced reversible photomagnetic effect. In the CoW system, a CT-induced spin transition causes the

  16. Inhibition by levetiracetam of a non-GABAA receptor-associated epileptiform effect of bicuculline in rat hippocampus

    PubMed Central

    Margineanu, Doru Georg; Wülfert, Ernst


    Extracellular recording of field potentials, evoked by commissural stimulation in hippocampal area CA3 of anaesthetized rats, was performed in order to study the mode of action of the novel antiepileptic drug levetiracetam (ucb LO59). The amplitude of orthodromic field population spike (PS2) markedly increased and repetitive population spikes appeared when the recording micropipette contained either bicuculline methiodide (BMI), or the specific GABAA antagonist gabazine (SR-95531). BMI-induced increases in PS2 were reduced in a dose-dependent manner by 1 to 320 μmol kg−1 levetiracetam i.v., with a U-shape dose-response relationship. However, levetiracetam did not reduce the increases in PS2 produced by gabazine. Clonazepam (1 mg kg−1, i.p.), carbamazepine (20 mg kg−1, i.p.) and valproate (200 mg kg−1, i.v.) were ineffective in preventing BMI-induced increases in PS2, while the calcium channel antagonist flunarizine, 50 μmol kg−1, i.p., reduced PS2 increments caused by BMI. The L-type calcium channel blocker nifedipine, 100 μmol kg−1, i.p., was without effect. Similar to levetiracetam, flunarizine did not reduce the increases in PS2 induced by gabazine. These data suggest that the increased excitability of CA3 neurones, caused by BMI administered in situ, involves calcium-dependent processes not associated with blockade of GABAA receptors. The inhibition by levetiracetam of this calcium-dependent effect of BMI might contribute to the antiepileptic effects of the drug. PMID:9401779

  17. Effects of chain-length and unsaturation on affinity and selectivity at muscarinic receptors.

    PubMed Central

    Barlow, R. B.; Holdup, D. W.; Harris, G.; Veale, M. A.; Williams, A.


    1. Lengthening the chain in diphenylacetylcholine decreases affinity for muscarinic cholinoceptors in guinea-pig ileum. Diphenylacetoxypropyldimethylamine and its quaternary trimethylammonium salt are roughly equiactive: the dimethylamine and the piperidine have some selectivity for ileum compared with atria, but are not as active nor as selective as 4-diphenylacetoxy-N-methylpiperidine (4-DAMP) methobromide (MeBr). With the weaker diphenylacetoxybutyl compounds the base is more active than the quaternary salt. 2. The diphenylacetoxybutyl-, cis-butenyl and trans-butenyl compounds have similar affinities. The quaternary salts are less active than the tertiary bases, but they are less selective than the butynyl analogues studied in earlier work. 3. 1,1-Diphenyl-1-hydroxy-2,4-hexadiynyl dimethylamine and its trimethylammonium salt are inactive in concentrations below 100 microM, as are the (+)-camphor-sulphonyl ester of 4-hydroxy-N-methyl piperidine and its methiodide. The (+/-)-phenylcyclopentylacetyl ester of 4-hydroxy-N-methylpiperidine methobromide is more active than its cyclohexyl analogue and than 4-DAMP MeBr but it is less selective than 4-DAMP MeBr. 4. The high selectivity of p-fluoro-hexahydrosila-diphenidol is confirmed but this compound has relatively low affinity (for ileum log K = 7.8). 5. The results indicate steric constraints to binding at muscarinic receptors which could be used to check molecular modelling of the receptor based on its known amino acid sequence. The group binding the charged nitrogen is probably at the mouth of a cavity which can accommodate two large rings (as in 4-DAMP MeBr) but with a depth less than about 7 A so that the rod-like hexadiynes cannot fit.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2331586

  18. The Sensory Impact of Nicotine on Noradrenergic and Dopaminergic Neurons of the Nicotine Reward - Addiction Neurocircuitry

    PubMed Central

    Rose, Jed E; Dehkordi, Ozra; Manaye, Kebreten F; Millis, Richard M; Cianaki, Salman Ameri; Jayam-Trouth, Annapurni


    The sensory experience of smoking is a key component of nicotine addiction known to result, in part, from stimulation of nicotinic acetylcholine receptors (nAChRs) at peripheral sensory nerve endings. Such stimulation of nAChRs is followed by activation of neurons at multiple sites in the mesocorticolimbic reward pathways. However, the neurochemical profiles of CNS cells that mediate the peripheral sensory impact of nicotine remain unknown. In the present study in mice, we first used c-Fos immunohistochemistry to identify CNS cells stimulated by nicotine (NIC, 40 μg/kg, IP) and by a peripherally-acting analog of nicotine, nicotine pyrrolidine methiodide (NIC-PM, 30 μg/kg, IP). Sequential double-labelling was then performed to determine whether noradrenergic and dopaminergic neurons of the nicotine reward-addiction circuitry were primary targets of NIC and NIC-PM. Double-labelling of NIC and/or NIC-PM activated c-Fos immunoreactive cells with tyrosine hydroxylase (TH) showed no apparent c-Fos expression by the dopaminergic cells of the ventral tegmental area (VTA). With the exception of sparse numbers of TH immunoreactive D11 cells, dopamine-containing neurons in other areas of the reward-addiction circuitry, namely periaqueductal gray, and dorsal raphe, were also devoid of c-Fos immunoreactivity. Noradrenergic neurons of locus coeruleus (LC), known to innervate VTA, were activated by both NIC and NIC-PM. These results demonstrate that noradrenergic neurons of LC are among the first structures that are stimulated by single acute IP injection of NIC and NIC-PM. Dopaminergic neurons of VTA and other CNS sites, did not respond to acute IP administration of NIC or NIC-PM by induction of c-Fos. PMID:27347434

  19. Neuroanatomical circuitry mediating the sensory impact of nicotine in the central nervous system.


    Dehkordi, Ozra; Rose, Jed E; Asadi, Sadegh; Manaye, Kebreten F; Millis, Richard M; Jayam-Trouth, Annapurni


    Direct actions of nicotine in the CNS appear to be essential for its reinforcing properties. However, activation of nicotinic acetylcholine receptors (nAChRs) on afferent sensory nerve fibers is an important component of addiction to, and withdrawal from, cigarette smoking. The aim of the present study was to identify the neuroanatomical substrates activated by the peripheral actions of nicotine and to determine whether these sites overlap brain structures stimulated by direct actions of nicotine. Mouse brains were examined by immunohistochemistry for c-Fos protein after intraperitoneal injection of either nicotine hydrogen tartrate salt (NIC; 30 and 40 μg/kg) or nicotine pyrrolidine methiodide (NIC-PM; 20 and 30 μg/kg). NIC-PM induced c-Fos immunoreactivity (IR) at multiple brain sites. In the brainstem, c-Fos IR was detected in the locus coeruleus, laterodorsal tegmental nucleus, and pedunculotegmental nucleus. In the midbrain, c-Fos IR was observed in areas overlapping the ventral tegmental area (VTA), which includes the paranigral nucleus, parainterfascicular nucleus, parabrachial pigmental area, and rostral VTA. Other structures of the nicotine brain-reward circuitry activated by NIC-PM included the hypothalamus, paraventricular thalamic nucleus, lateral habenular nucleus, hippocampus, amygdala, accumbens nucleus, piriform cortex, angular insular cortex, anterior olfactory nucleus, lateral septal nucleus, bed nucleus of stria terminalis, cingulate and medial prefrontal cortex, olfactory tubercle, and medial and lateral orbital cortex. NIC, acting through central and peripheral nAChRs, produced c-Fos IR in areas that overlapped NIC-PM-induced c-Fos-expressing sites. These neuroanatomical data are the first to demonstrate that the CNS structures that are the direct targets of nicotine are also anatomical substrates for the peripheral sensory impact of nicotine.

  20. Nuclear medicine progress report for quarter ending September 30, 1984

    SciTech Connect

    Knapp, F.F. Jr.; Ambrose, K.R.; Goodman, M.M.; Srivastava, P.C.


    The preparation and animal testing of a new radio-iodinated p-iodophenylamine-linked dihydropyridine system is described. The model agent, 1-methyl-3-(N-(..beta..-(4-(/sup 125/I)iodophenyl)ethyl)carbamoyl)-1,4-dihydropyridine, was prepared by coupling 4-(/sup 125/I)iodoaniline with the methiodide salt succinimidyl ester of nicotinic acid followed by dithionite reduction to the lipid soluble product. The dihydropyridine agent showed good brain uptake in rats (5 min, 1.14% injected dose/gm; 60 min, 1.12% dose/gm) and good brain to blood ratios (5 min 3.9:1, 60 min, 3.5:1). In contrast the quaternary ammonium compound, prior to reduction, showed only moderate brain uptake (5 min, 0.63; 60 min, 0.46) and low brain to blood ratios (5 min, 0.05; 60 min, 0.06). Also described is further investigation of the effects of fasting on the relative myocardial retention of straight-chain iodovinyl fatty acids. 18-(/sup 125/)Iodo-17-octadecenoic acid showed good retention in unfasted rats. Studies have now been reported for fasted rats where this agent showed rapid myocardial wash-out. In fasted rats, approx. 70% wash-out at 30 min, and in unfasted rats, approx. 15% wash-out at 30 min was observed. During this period several shipments were made to Medical Cooperative investigators including three samples of /sup 191/Os-potassium osmate (Children's Hospital, Boston, and the University of Liege, Belgium) and 15-(p-(/sup 131/I)iodophenyl)-3-R,S-methylpentadecanoic acid (University of Massachusetts and Brookhaven National Laboratory).

  1. Defense-like behaviors evoked by pharmacological disinhibition of the superior colliculus in the primate

    PubMed Central

    DesJardin, Jacqueline T.; Holmes, Angela L.; Forcelli, Patrick A.; Cole, Claire E.; Gale, John T.; Wellman, Laurie L.; Gale, Karen; Malkova, Ludise


    Stimulation of the intermediate and deep layers of superior colliculus (DLSC) in rodents evokes both tracking/pursuit (approach) and avoidance/flight (defensive) responses (Dean et al., 1989). These two classes of response are subserved by distinct output projections associated with lateral (approach) and medial (defensive) DLSC (Comoli et al., 2012). In nonhuman primates, DLSC has been examined only with respect to orienting/approach behaviors, especially eye movements, however, defense-like behaviors have not been reported. Here we examined the profile of behavioral responses to the activation of DLSC by unilateral intracerebral infusions of the GABAA receptor antagonist, bicuculline methiodide (BIC), in nine freely moving macaques. Across animals, the most consistently evoked behavior was cowering (all animals), followed by increased vocalization and escape-like behaviors (seven animals), and attack of objects (three animals). The effects of BIC were dose-dependent within the range 2.5-14nmol (threshold dose of 4.6nmol). The behaviors and their latency to onset did not vary across different infusion sites within DLSC. Cowering and escape-like behaviors resembled the defense-like responses reported after DLSC stimulation in rats, but in the macaques these responses were evoked from both medial and lateral sites within DLSC. Our findings are unexpected in the context of an earlier theoretical perspective (Dean et al., 1989) that emphasized a preferential role of the primate DLSC for approach rather than defensive responses. Our data provide the first evidence for induction of defense-like behaviors by activation of DLSC in monkeys, suggesting that the role of DLSC in responding to threats is conserved across species. PMID:23283329

  2. Cardiovascular and behavioral effects produced by administration of liposome-entrapped GABA into the rat central nervous system.


    Vaz, G C; Bahia, A P C O; de Figueiredo Müller-Ribeiro, F C; Xavier, C H; Patel, K P; Santos, R A S; Moreira, F A; Frézard, F; Fontes, M A P


    Liposomes are nanosystems that allow a sustained release of entrapped substances. Gamma-aminobutyric acid (GABA) is the most prevalent inhibitory neurotransmitter of the central nervous system (CNS). We developed a liposomal formulation of GABA for application in long-term CNS functional studies. Two days after liposome-entrapped GABA was injected intracerebroventricularly (ICV), Wistar rats were submitted to the following evaluations: (1) changes in mean arterial pressure (MAP), heart rate (HR) and renal sympathetic nerve activity (RSNA) to ICV injection of bicuculline methiodide (BMI) in anesthetized rats; (2) changes in cardiovascular reactivity to air jet stress in conscious rats; and (3) anxiety-like behavior in conscious rats. GABA and saline-containing pegylated liposomes were prepared with a mean diameter of 200 nm. Rats with implanted cannulas targeted to lateral cerebral ventricle (n = 5-8/group) received either GABA solution (GS), empty liposomes (EL) or GABA-containing liposomes (GL). Following (48 h) central microinjection (2 μL, 0.09 M and 99 g/L) of liposomes, animals were submitted to the different protocols. Animals that received GL demonstrated attenuated response of RSNA to BMI microinjection (GS 48 ± 9, EL 43 ± 9, GL 11 ± 8%; P < 0.05), blunted tachycardia in the stress trial (ΔHR: GS 115 ± 14, EL 117 ± 10, GL 74 ± 9 bpm; P<0.05) and spent more time in the open arms of elevated plus maze (EL 6 ± 2 vs. GL 18 ± 5%; P = 0.028) compared with GS and EL groups. These results indicate that liposome-entrapped GABA can be a potential tool for exploring the chronic effects of GABA in specific regions and pathways of the central nervous system.

  3. New insights on amygdala: Basomedial amygdala regulates the physiological response to social novelty.


    Mesquita, Laura Tavares; Abreu, Aline Rezende; de Abreu, Alessandra Rezende; de Souza, Aline Arlindo; de Noronha, Sylvana Rendeiro; Silva, Fernanda Cacilda; Campos, Glenda Siqueira Viggiano; Chianca, Deoclecio Alves; de Menezes, Rodrigo Cunha


    The amygdala has been associated with a variety of functions linked to physiological, behavioral and endocrine responses during emotional situations. This brain region is comprised of multiple sub-nuclei. These sub-nuclei belong to the same structure, but may be involved in different functions, thereby making the study of each sub-nuclei important. Yet, the involvement of the basomedial amygdala (BMA) in the regulation of emotional states has yet to be defined. Therefore, the aim of our study was to investigate the regulatory role of the BMA on the responses evoked during a social novelty model and whether the regulatory role depended on an interaction with the dorsomedial hypothalamus (DMH). Our results showed that the chemical inhibition of the BMA by the microinjection of muscimol (γ-aminobutyric acid (GABAA) agonist) promoted increases in mean arterial pressure (MAP) and heart rate (HR), whereas the chemical inhibition of regions near the BMA did not induce such cardiovascular changes. In contrast, the BMA chemical activation by the bilateral microinjection of bicuculline methiodide (BMI; GABAA antagonist), blocked the increases in MAP and HR observed when an intruder rat was suddenly introduced into the cage of a resident rat, and confined to the small cage for 15min. Additionally, the increase in HR and MAP induced by BMA inhibition were eliminated by DMH chemical inhibition. Thus, our data reveal that the BMA is under continuous GABAergic influence, and that its hyperactivation can reduce the physiological response induced by a social novelty condition, possibly by inhibiting DMH neurons. PMID:27261213

  4. The prefrontal cortex and oculomotor delayed response: a reconsideration of the "mnemonic scotoma".


    Tsujimoto, Satoshi; Postle, Bradley R


    The concept of the "mnemonic scotoma," a spatially circumscribed region of working memory impairment produced by unilateral lesions of the PFC, is central to the view that PFC is critical for the short-term retention of information. Presented here, however, are previously unpublished data that offer an alternative, nonmnemonic interpretation of this pattern of deficit. In their study, Wajima and Sawaguchi [Wajima, K., & Sawaguchi, T. The role of GABAergic inhibiton in suppressing perseverative responses in the monkey prefrontal cortex. Neuroscience Research, 50(Suppl. 1), P3-P317, 2004] applied the GABA(A) antagonist bicuculline methiodide unilaterally to the PFC of two monkeys while they performed an oculomotor delayed-response task. Consistent with previous studies, errors for the initial memory-guided saccade were markedly higher when the cued location fell into the region of the visual field affected by the infusion. These erroneous saccades tended to select an alternative target location (out of a possible 16) that had not been cued on that trial. By extending the analysis window, however, it was observed that the second, "corrective" saccade often acquired the location that had been cued on that trial. Further analysis of the erroneous initial saccades indicated that they tended to be directed to a location that had been relevant on the previous trial. Thus, the deficit was not one of "forgetting" the cued location. Rather, it was one of selecting between currently and previously relevant locations. These findings suggest a need for a reconsideration of the concept of the mnemonic scotoma, which in turn invites a reconsideration of functional interpretations of sustained neuronal activity in PFC. PMID:22098265

  5. Reduced striatal acetylcholine efflux in the R6/2 mouse model of Huntington's disease: an examination of the role of altered inhibitory and excitatory mechanisms.


    Farrar, Andrew M; Callahan, Joshua W; Abercrombie, Elizabeth D


    Huntington's disease (HD) is a genetic neurodegenerative disorder that is characterized by the progressive onset of cognitive, psychiatric, and motor symptoms. In parallel, the neuropathology of HD is characterized by progressive loss of projection neurons in cortex and striatum; striatal cholinergic interneurons are relatively spared. Nonetheless, there is evidence that striatal acetylcholine (ACh) function is altered in HD. The present study is the first to examine striatal ACh function in awake, behaving animals, using the R6/2 mouse model of HD, which is transgenic for exon 1 of the mutant huntingtin gene. Physiological levels of extracellular striatal ACh were monitored in R6/2 mice and wild type controls using in vivo microdialysis. Results indicate that spontaneous ACh release is reduced in R6/2 mice relative to controls. Intrastriatal application of the GABA(A) antagonist bicuculline methiodide (10.0 μM) significantly elevated ACh levels in both R6/2 mice and wild type controls, while overall ACh levels were reduced in the R6/2 mice compared to the wild type group. In contrast, systemic administration of the D(1) dopamine receptor partial agonist, SKF-38393 (10.0mg/kg, IP), elevated ACh levels in control animals, but not R6/2 mice. Taken together, the present results suggest that GABA-mediated inhibition of striatal ACh release is intact in R6/2 mice, further demonstrating that cholinergic interneurons are capable of increased ACh release, whereas D(1) receptor-dependent activation of excitatory inputs to striatal cholinergic interneurons is dysfunctional in R6/2 mice. Reduced levels of extracellular striatal ACh in HD may reflect abnormalities in the excitatory innervation of cholinergic interneurons, which may have implications ACh-dependent processes that are altered in HD, including corticostriatal plasticity.

  6. Muscarinic acetylcholine receptor modulation of mu (mu) opioid receptors in adult rat sphenopalatine ganglion neurons.


    Margas, Wojciech; Mahmoud, Saifeldin; Ruiz-Velasco, Victor


    The sphenopalatine ganglion (SPG) neurons represent the parasympathetic branch of the autonomic nervous system involved in controlling cerebral blood flow. In the present study, we examined the coupling mechanism between mu (mu) opioid receptors (MOR) and muscarinic acetylcholine receptors (mAChR) with Ca(2+) channels in acutely dissociated adult rat SPG neurons. Successful MOR activation was recorded in approximately 40-45% of SPG neurons employing the whole cell variant of the patch-clamp technique. In addition, immunofluorescence assays indicated that MOR are not expressed in all SPG neurons while M(2) mAChR staining was evident in all neurons. The concentration-response relationships generated with morphine and [d-Ala2-N-Me-Phe4-Glycol5]-enkephalin (DAMGO) showed IC(50) values of 15.2 and 56.1 nM and maximal Ca(2+) current inhibition of 26.0 and 38.7%, respectively. Activation of MOR or M(2) mAChR with morphine or oxotremorine-methiodide (Oxo-M), respectively, resulted in voltage-dependent inhibition of Ca(2+) currents via coupling with Galpha(i/o) protein subunits. The acute prolonged exposure (10 min) of neurons to morphine or Oxo-M led to the homologous desensitization of MOR and M(2) mAChR, respectively. The prolonged stimulation of M(2) mAChR with Oxo-M resulted in heterologous desensitization of morphine-mediated Ca(2+) current inhibition, and was sensitive to the M(2) mAChR blocker methoctramine. On the other hand, when the neurons were exposed to morphine or DAMGO for 10 min, heterologous desensitization of M(2) mAChR was not observed. These results suggest that in rat SPG neurons activation of M(2) mAChR likely modulates opioid transmission in the brain vasculature to adequately maintain cerebral blood flow. PMID:19889856

  7. Peripheral endothelin B receptor agonist-induced antinociception involves endogenous opioids in mice.


    Quang, Phuong N; Schmidt, Brian L


    Endothelin-1 (ET-1) produced by various cancers is known to be responsible for inducing pain. While ET-1 binding to ETAR on peripheral nerves clearly mediates nociception, effects from binding to ETBR are less clear. The present study assessed the effects of ETBR activation and the role of endogenous opioid analgesia in carcinoma pain using an orthotopic cancer pain mouse model. mRNA expression analysis showed that ET-1 was nearly doubled while ETBR was significantly down-regulated in a human oral SCC cell line compared to normal oral keratinocytes (NOK). Squamous cell carcinoma (SCC) cell culture treated with an ETBR agonist (10(-4)M, 10(-5)M, and 10(-6) M BQ-3020) significantly increased the production of beta-endorphin without any effects on leu-enkephalin or dynorphin. Cancer inoculated in the hind paw of athymic mice with SCC induced significant pain, as indicated by reduction of paw withdrawal thresholds in response to mechanical stimulation, compared to sham-injected and NOK-injected groups. Intratumor administration of 3mg/kg BQ-3020 attenuated cancer pain by approximately 50% up to 3h post-injection compared to PBS-vehicle and contralateral injection, while intratumor ETBR antagonist BQ-788 treatment (100 and 300microg/kg and 3mg/kg) had no effects. Local naloxone methiodide (500microg/kg) or selective mu-opioid receptor antagonist (CTOP, 500microg/kg) injection reversed ETBR agonist-induced antinociception in cancer animals. We propose that these results demonstrate that peripheral ETBR agonism attenuates carcinoma pain by modulating beta-endorphins released from the SCC to act on peripheral opioid receptors found in the cancer microenvironment.

  8. Rational Basis for the Use of Bergamot Essential Oil in Complementary Medicine to Treat Chronic Pain.


    Rombolà, L; Amantea, D; Russo, R; Adornetto, A; Berliocchi, L; Tridico, L; Corasaniti, M T; Sakurada, S; Sakurada, T; Bagetta, G; Morrone, L A


    In complementary medicine, aromatherapy uses essential oils to improve agitation and aggression observed in dementia, mood, depression, anxiety and chronic pain. Preclinical research studies have reported that the essential oil obtained from bergamot (BEO) fruit (Citrus bergamia, Risso) modifies normal and pathological synaptic plasticity implicated, for instance, in nociceptive and neuropathic pain. Interestingly, recent results indicated that BEO modulates sensitive perception of pain in different models of nociceptive, inflammatory and neuropathic pain modulating endogenous systems. Thus, local administration of BEO inhibited the nociceptive behavioral effect induced by intraplantar injection of capsaicin or formalin in mice. Similar effects were observed with linalool and linalyl acetate, major volatile components of the phytocomplex, Pharmacological studies showed that the latter effects are reversed by local or systemic pretreatment with the opioid antagonist naloxone hydrochloride alike with naloxone methiodide, high affinity peripheral μ-opioid receptor antagonist. These results and the synergistic effect observed following systemic or intrathecal injection of an inactive dose of morphine with BEO or linalool indicated an activation of peripheral opioid system. Recently, in neuropathic pain models systemic or local administration of BEO or linalool induced antiallodynic effects. In particular, in partial sciatic nerve ligation (PSNL) model, intraplantar injection of the phytocomplex or linalool in the ipsilateral hindpaw, but not in the contralateral, reduced PSNL-induced extracellularsignal- regulated kinase (ERK) activation and mechanical allodynia. In neuropathic pain high doses of morphine are needed to reduce pain. Interestingly, combination of inactive doses of BEO or linalool with a low dose of morphine induced antiallodynic effects in mice. Peripheral cannabinoid and opioid systems appear to be involved in the antinociception produced by

  9. Site-selective chemical modification of chymotrypsin using peptidyl derivatives bearing optically active diphenyl 1-amino-2-phenylethylphosphonate: Stereochemical effect of the diphenyl phosphonate moiety.


    Ono, Shin; Nakai, Takahiko; Kuroda, Hirofumi; Miyatake, Ryuta; Horino, Yoshikazu; Abe, Hitoshi; Umezaki, Masahito; Oyama, Hiroshi


    Diphenyl (α-aminoalkyl)phosphonates act as mechanism-based inhibitors against serine proteases by forming a covalent bond with the hydroxy group of the active center Ser residue. Because the covalent bond was found to be broken and replaced by 2-pyridinaldoxime methiodide (2PAM), we employed a peptidyl derivative bearing diphenyl 1-amino-2-phenylethylphosphonate moiety (Phe(p) (OPh)2 ) to target the active site of chymotrypsin and to selectively anchor to Lys175 in the vicinity of the active site. Previously, it was reported that the configuration of the α-carbon of phosphorus in diphenyl (α-aminoalkyl)phosphonates affects the inactivation reaction of serine proteases, i.e., the (R)-enantiomeric diphenyl phosphonate is comparable to l-amino acids and it effectively reacts with serine proteases, whereas the (S)-enantiomeric form does not. In this study, we evaluated the stereochemical effect of the phosphonate moiety on the selective chemical modification. Epimeric dipeptidyl derivatives, Ala-(R or S)-Phe(p) (OPh)2 , were prepared by separation with RP-HPLC. A tripeptidyl (R)-epimer (Ala-Ala-(R)-Phe(p) (OPh)2 ) exhibited a more potent inactivation ability against chymotrypsin than the (S)-epimer. The enzyme inactivated by the (R)-epimer was more effectively reactivated with 2PAM than the enzyme inactivated by the (S)-epimer. Finally, N-succinimidyl (NHS) active ester derivatives, NHS-Suc-Ala-Ala- (R or S)-Phe(p) (OPh)2 , were prepared, and we evaluated their action when modifying Lys175 in chymotrypsin. We demonstrated that the epimeric NHS derivative that possessed the diphenyl phosphonate moiety with the (R)-configuration effectively modified Lys175 in chymotrypsin, whereas that with the (S)-configuration did not. These results demonstrate the utility of peptidyl derivatives that bear an optically active diphenyl phosphonate moiety as affinity labeling probes in protein bioconjugation. © 2015 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 521-530, 2016.

  10. 2-Deoxy-D-glucose-induced hypothermia in anesthetized rats: Lack of forebrain contribution and critical involvement of the rostral raphe/parapyramidal regions of the medulla oblongata.


    Osaka, Toshimasa


    Systemic or central administration of 2-deoxy-d-glucose (2DG), a competitive inhibitor of glucose utilization, induces hypothermia in awake animals and humans. This response is mediated by the central nervous system, though the neural mechanism involved is largely unknown. In this study, I examined possible involvement of the forebrain, which contains the hypothalamic thermoregulatory center, and the medullary rostral raphe/parapyramidal regions (rRPa/PPy), which mediate hypoxia-induced heat-loss responses, in 2DG-induced hypothermia in urethane-chloralose-anesthetized, neuromuscularly blocked, artificially ventilated rats. The intravenous injection of 2DG (250mgkg(-1)) elicited an increase in tail skin temperature and decreases in body core temperature and the respiratory exchange ratio, though it did not induce any significant change in the metabolic rate. These results indicate that the hypothermic response was caused by an increase in heat loss, but not by a decrease in heat production and that it was accompanied by a decrease in carbohydrate utilization and/or an increase in lipid utilization as energy substrates. Complete surgical transection of the brainstem between the hypothalamus and the midbrain had no effect on the 2DG-induced hypothermic responses, suggesting that the hindbrain, but not the forebrain, was sufficient for the responses. However, pretreatment of the rRPa/PPy with the GABAA receptor blocker bicuculline methiodide, but not with vehicle saline, greatly attenuated the 2DG-induced responses, suggesting that the 2DG-induced hypothermia was mediated, at least in part, by GABAergic neurons in the hindbrain and activation of GABAA receptors on cutaneous sympathetic premotor neurons in the rRPa/PPy. PMID:26146232

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

    NASA Technical Reports Server (NTRS)

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


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

  12. The role of GABAergic inhibition in processing of interaural time difference in the owl's auditory system.


    Fujita, I; Konishi, M


    The barn owl uses interaural time differences (ITDs) to localize the azimuthal position of sound. ITDs are processed by an anatomically distinct pathway in the brainstem. Neuronal selectivity for ITD is generated in the nucleus laminaris (NL) and conveyed to both the anterior portion of the ventral nucleus of the lateral lemniscus (VLVa) and the central (ICc) and external (ICx) nuclei of the inferior colliculus. With tonal stimuli, neurons in all regions are found to respond maximally not only to the real ITD, but also to ITDs that differ by integer multiples of the tonal period. This phenomenon, phase ambiguity, does not occur when ICx neurons are stimulated with noise. The main aim of this study was to determine the role of GABAergic inhibition in the processing of ITDs. Selectivity for ITD is similar in the NL and VLVa and improves in the ICc and ICx. Iontophoresis of bicuculline methiodide (BMI), a selective GABAA antagonist, decreased the ITD selectivity of ICc and ICx neurons, but did not affect that of VLVa neurons. Responses of VLVa and ICc neurons to unfavorable ITDs were below the monaural response levels. BMI raised both binaural responses to unfavorable ITDs and monaural responses, though the former remained smaller than the latter. During BMI application, ICx neurons showed phase ambiguity to noise stimuli and no longer responded to a unique ITD. BMI increased the response magnitude and changed the temporal discharge patterns in the VLVa, ICc, and ICx. Iontophoretically applied GABA exerted effects opposite to those of BMI, and the effects could be antagonized with simultaneous application of BMI. These results suggest that GABAergic inhibition (1) sharpens ITD selectivity in the ICc and ICx, (2) contributes to the elimination of phase ambiguity in the ICx, and (3) controls response magnitude and temporal characteristics in the VLVa, ICc, and ICx. Through these actions, GABAergic inhibition shapes the horizontal dimension of the auditory receptive

  13. In Vivo Voltage-Sensitive Dye Study of Lateral Spreading of Cortical Activity in Mouse Primary Visual Cortex Induced by a Current Impulse

    PubMed Central

    Fehérvári, Tamás Dávid; Sawai, Hajime; Yagi, Tetsuya


    In the mammalian primary visual cortex (V1), lateral spreading of excitatory potentials is believed to be involved in spatial integrative functions, but the underlying cortical mechanism is not well understood. Visually-evoked population-level responses have been shown to propagate beyond the V1 initial activation site in mouse, similar to higher mammals. Visually-evoked responses are, however, affected by neuronal circuits prior to V1 (retina, LGN), making the separate analysis of V1 difficult. Intracortical stimulation eliminates these initial processing steps. We used in vivo RH1691 voltage-sensitive dye (VSD) imaging and intracortical microstimulation in adult C57BL/6 mice to elucidate the spatiotemporal properties of population-level signal spreading in V1 cortical circuits. The evoked response was qualitatively similar to that measured in single-cell electrophysiological experiments in rodents: a fast transient fluorescence peak followed by a fast and a slow decrease or hyperpolarization, similar to EPSP and fast and slow IPSPs in single cells. The early cortical response expanded at speeds commensurate with long horizontal projections (at 5% of the peak maximum, 0.08–0.15 m/s) however, the bulk of the VSD signal propagated slowly (at half-peak maximum, 0.05–0.08 m/s) suggesting an important role of regenerative multisynaptic transmission through short horizontal connections in V1 spatial integrative functions. We also found a tendency for a widespread and fast cortical response suppression in V1, which was eliminated by GABAA-antagonists gabazine and bicuculline methiodide. Our results help understand the neuronal circuitry involved in lateral spreading in V1. PMID:26230520

  14. Antinociceptive and anti-allodynic effects of oral PL37, a complete inhibitor of enkephalin-catabolizing enzymes, in a rat model of peripheral neuropathic pain induced by vincristine.


    Thibault, Karine; Elisabeth, Bonnard; Sophie, Dubacq; Claude, Fournié-Zaluski Marie; Bernard, Roques; Bernard, Calvino


    Vincristine is a common anti-cancer therapy administered for the treatment of many types of tumors. Its dose-limiting side effect is the production of peripheral neuropathy, resulting in chronic neuropathic pain in many patients. An animal model of vincristine-induced sensory neuropathy was developed after repeated intraperitoneal injection in male rats and used in the present work to study the effects of PL37, an orally active complete dual inhibitor of enkephalin-catabolizing enzymes, on mechanical hypersensitivity and allodynia and on cold allodynia. We used the Electronic Von Frey filament (mechanical static allodynia), acetone test (cold allodynia), and a new behavioural test we first describe in this study, the "paint-brush test" which evaluates dynamic mechanical allodynia and dynamic mechanical hypersensitivity. We used a smooth paint brush leading to an innocuous stimulus, and a rough-one leading to an intense mechanical stimulus. Mechanical hypersensitivity and allodynia due to vincristine-induced neuropathy, but not cold allodynia, are strongly reduced by oral or i.p. injected PL37, the dose-dependent effects being reversed by naloxone-methiodide supporting the peripheral action of the dual inhibitor. These results show that enkephalins protected from degradation by PL37 could bind to peripheral opioid receptors expressed only on C- and Adelta-mechanonociceptors but not on cold thermonociceptors. The fact that PL37 is also active on small intensity mechanical stimulus could reveal an expression of opioid receptors on low threshold mechanoreceptors in the vincrisitine-evoked pathological conditions. Thus the increase in endogenous enkephalin levels induced by PL37 offers a new way to reduced neuropathic pain without the possible side effects of opiates. PMID:18938155

  15. GABAergic inhibition and modifications of taste responses in the cortical taste area in rats.


    Ogawa, H; Hasegawa, K; Otawa, S; Ikeda, I


    Using multibarrel electrodes, recordings were made in the cortical taste area (CTA), specifically in the granular and dysgranular parts of the insular cortex (areas GI and DI), of urethane-anesthetized rats. The effects of an iontophoretic application of gamma-aminobutylic acid (GABA) and bicuculline methiodide (BMI), a specific antagonist to the GABA(A) receptor, were tested. GABA decreased background discharges in ca. 69% of 509 neurons in both areas, and in ca. 58% of 64 taste neurons. BMI antagonized the inhibitory action of GABA in CTA neurons and facilitated background discharges in ca. 51% of the 390 neurons tested, including ca. 69% of the 52 taste neurons, which indicates that CTA neurons have GABA(A) receptors to receive inhibitory inputs from interneurons. In both areas, the effects of BMI (6-20 nA) on taste responses of the 85 CTA neurons (49 and 36 in areas GI and DI, respectively) to the four basic taste stimuli were examined: 65 neurons were recognized in the absence of BMI, whereas 20 only in the presence of the drug. BMI increased taste responses in 25 of the former group and changed the type of their response profiles in 25 including 12 neurons whose responses were increased. It also changed the best stimulus in 34 neurons. The drug affected the receptive fields in almost all cases examined (n = 23) and increased the size in 78.2% when the value for all four basic taste stimuli were totaled. New receptive fields were uncovered by BMI in varying regions of the oral cavity depending on the taste stimulus. But the drug decreased taste responses in several neurons (n = 8). These findings indicate that the GABAergic inhibitory system apparently contributes to modifying or selecting taste information in both areas of the CTA. PMID:9831255

  16. The effect of atropine on the activation of 5-hydroxytryptamine3 channels in rat nodose ganglion neurons.


    Fan, P; Weight, F F


    It has been suggested that changes in brain 5-hydroxytryptamine3 receptor function may contribute to some behavior disorders, such as anxiety, schizophrenia and drug abuse. We are using the whole-cell version of the patch-clamp technique to study the function of 5-hydroxytryptamine3 channels in neurons freshly dissociated from rat nodose ganglion. In these cells, 5-hydroxytryptamine elicits an inward current over the concentration range of 0.25-100 microM (EC50 = 2.62 microM) by activating 5-hydroxytryptamine3 receptors. The muscarinic cholinergic antagonist atropine reduced the amplitude of 5-hydroxytryptamine activated inward current in a concentration-dependent manner. Other muscarinic antagonists, scopolamine, dexetimide, the M1 muscarinic receptor antagonist pirenzepine, the M2 receptor antagonist methoctramine and the M3 receptor antagonist 4-DAMP methiodide also inhibited 5-hydroxytryptamine-induced inward current. Atropine did not appear to change the reversal potential of this current. In the presence of 5 microM atropine, the concentration-response curve for 5-hydroxytryptamine current was shifted to the right in a parallel fashion. The EC50 value for 5-hydroxytryptamine was increased from 2.62 to 8.76 microM. Schild plots of increasing atropine and 5-hydroxytryptamine concentrations revealed a pA2 value of 5.74 for atropine (apparent KD = 1.8 microM). These observations suggest that atropine competitively antagonizes the activation of a receptor for the neurotransmitter serotonin, a novel action of muscarinic antagonists in the nervous system. This effect of atropine may contribute to the clinical symptoms seen in severe atropine intoxication. PMID:7531305

  17. Fluctuations in central and peripheral temperatures induced by intravenous nicotine: central and peripheral contributions.


    Tang, Jeremy S; Kiyatkin, Eugene A


    Nicotine (NIC) is a highly addictive substance that interacts with different subtypes of nicotinic acetylcholine receptors widely distributed in the central and peripheral nervous systems. While the direct action of NIC on central neurons appears to be essential for its reinforcing properties, the role of peripheral actions of this drug remains a matter of controversy. In this study, we examined changes in locomotor activity and temperature fluctuations in the brain (nucleus accumbens and ventral tegmental area), temporal muscle, and skin induced by intravenous (iv) NIC at low human-relevant doses (10 and 30μg/kg) in freely moving rats. These effects were compared to those induced by social interaction, an arousing procedure that induces behavioral activation and temperature responses via pure neural mechanisms, and iv injections of a peripherally acting NIC analog, NIC pyrrolidine methiodide (NIC-PM) used at equimolar doses. We found that NIC at 30μg/kg induces a modest locomotor activation, rapid and strong decrease in skin temperature, and weak increases in brain and muscle temperature. While these effects were qualitatively similar to those induced by social interaction, they were much weaker and showed a tendency to increase with repeated drug administrations. In contrast, NIC-PM did not affect locomotion and induced much weaker than NIC increases in brain and muscle temperatures and decreases in skin temperature; these effects showed a tendency to be weaker with repeated drug administrations. Our data indicate that NIC's actions in the brain are essential to induce locomotor activation and brain and body hyperthermic responses. However, rapid peripheral action of NIC on sensory afferents could be an important factor in triggering its central effects, contributing to neural and physiological activation following repeated drug use. PMID:21295014

  18. Rapid Sensitization of Physiological, Neuronal, and Locomotor Effects of Nicotine: Critical Role of Peripheral Drug Actions

    PubMed Central

    Lenoir, Magalie; Tang, Jeremy S.; Woods, Amina S.


    Repeated exposure to nicotine and other psychostimulant drugs produces persistent increases in their psychomotor and physiological effects (sensitization), a phenomenon related to the drugs' reinforcing properties and abuse potential. Here we examined the role of peripheral actions of nicotine in nicotine-induced sensitization of centrally mediated physiological parameters (brain, muscle, and skin temperatures), cortical and VTA EEG, neck EMG activity, and locomotion in freely moving rats. Repeated injections of intravenous nicotine (30 μg/kg) induced sensitization of the drug's effects on all these measures. In contrast, repeated injections of the peripherally acting analog of nicotine, nicotine pyrrolidine methiodide (nicotinePM, 30 μg/kg, i.v.) resulted in habituation (tolerance) of the same physiological, neuronal, and behavioral measures. However, after repeated nicotine exposure, acute nicotinePM injections induced nicotine-like physiological responses: powerful cortical and VTA EEG desynchronization, EMG activation, a large brain temperature increase, but weaker hyperlocomotion. Additionally, both the acute locomotor response to nicotine and nicotine-induced locomotor sensitization were attenuated by blockade of peripheral nicotinic receptors by hexamethonium (3 mg/kg, i.v.). These data suggest that the peripheral actions of nicotine, which precede its direct central actions, serve as a conditioned interoceptive cue capable of eliciting nicotine-like physiological and neural responses after repeated nicotine exposure. Thus, by providing a neural signal to the CNS that is repeatedly paired with the direct central effects of nicotine, the drug's peripheral actions play a critical role in the development of nicotine-induced physiological, neural, and behavioral sensitization. PMID:23761889

  19. Low-dose morphine elicits ventilatory excitant and depressant responses in conscious rats: Role of peripheral μ-opioid receptors.


    Henderson, Fraser; May, Walter J; Gruber, Ryan B; Young, Alex P; Palmer, Lisa A; Gaston, Benjamin; Lewis, Stephen J


    The systemic administration of morphine affects ventilation via a mixture of central and peripheral actions. The aims of this study were to characterize the ventilatory responses elicited by a low dose of morphine in conscious rats; to determine whether tolerance develops to these responses; and to determine the potential roles of peripheral μ-opioid receptors (μ-ORs) in these responses. Ventilatory parameters were monitored via unrestrained whole-body plethysmography. Conscious male Sprague-Dawley rats received an intravenous injection of vehicle or the peripherally-restricted μ-OR antagonist, naloxone methiodide (NLXmi), and then three successive injections of morphine (1 mg/kg) given 30 min apart. The first injection of morphine in vehicle-treated rats elicited an array of ventilatory excitant (i.e., increases in frequency of breathing, minute volume, respiratory drive, peak inspiratory and expiratory flows, accompanied by decreases in inspiratory time and end inspiratory pause) and inhibitory (i.e., a decrease in tidal volume and an increase in expiratory time) responses. Subsequent injections of morphine elicited progressively and substantially smaller responses. The pattern of ventilatory responses elicited by the first injection of morphine was substantially affected by pretreatment with NLXmi whereas NLXmi minimally affected the development of tolerance to these responses. Low-dose morphine elicits an array of ventilatory excitant and depressant effects in conscious rats that are subject to the development of tolerance. Many of these initial actions of morphine appear to involve activation of peripheral μ-ORs whereas the development of tolerance to these responses does not. PMID:24900948

  20. Effects of dihydroavermectin B1a and analogs on stretcher muscle of the lined shore crab, Pachygrapsus crassipes.


    Bowman, J W; Lee, B L; Whaley, H A; Thompson, D P


    1. Dihydroavermectin B1a (DHAVM, Ivermectin) at 1 microM reduces excitatory and inhibitory postsynaptic potentials (EPSPs and IPSPs, respectively) in stretcher muscle fibres of the lined shore crab, Pachygrapsus crassipes. IPSPs decline faster and more extensively than EPSPs and, unlike EPSPs, do not recover upon replacement of DHAVM with picrotoxinin-containing medium. 2. Intracellular recordings show DHAVM reduces membrane resistance (Rin) and hyperpolarizes muscle fibres in a concentration-dependent manner, beginning at 10 nM. The rate and magnitude of DHAVM effects on Rin mirror its effects on EPSPs. 3. The decline in Rin due to DHAVM is sustained over time (i.e. there is no tendency for desensitization); it is also irreversible and not affected by coadministration of 1 mM gamma-aminobutyric acid (GABA), 0.1 mM bicuculline methiodide or addition of 20 mM Co2+ to the recording medium. 4. Replacement of DHAVM-containing medium with medium containing Cl- channel blockers (picrotoxinin or lindane) results in partial recovery of Rin, while channel blockers specific for other ions (TTX, TEA, 4-AP or verapamil) are without effect. The decline of Rin following application of DHAVM is attenuated in Cl(-)-free medium. 5. Results of tests using compounds structurally related to DHAVM reveal that relatively minor changes in the molecule often reduce biological activity significantly. Removal of one sugar, for instance, results in a ten-fold reduction in potency. 6. In general, avermectins that stimulate conductance in shore crab muscle also possess anthelmintic activity at similar concentrations, based on studies using the free-living nematode, Caenorhabditis elegans.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1685403

  1. Ligand binding and functional characterization of muscarinic acetylcholine receptors on the TE671/RD human cell line

    SciTech Connect

    Bencherif, M.; Lukas, R.J. )


    Cells of the TE671/RD human clonal line express a finite number ((Bmax) of about 350 fmol/mg of membrane protein) of apparently noninteracting, high-affinity binding sites (KD of 0.07 nM and a Hill coefficient close to unity, nH = 0.94) for the muscarinic acetylcholine receptor (mAChR) radio antagonist, tritium-labeled quinuclidinyl benzilate ({sup 3}H-QNB). The rank order potency of selective antagonists that inhibit specific {sup 3}HQNB binding is: atropine greater than 4-DAMP (4-diphenylacetoxy-N-methylpiperidine methiodide) greater than pirenzepine greater than methoctramine greater than AFDx-116 (11-2(2-((diethylamino)methyl)-1-(piperidinyl) acetyl)-5,11-dihydro-6H-pyrido(2,3-b)(1,4)benzodiazepin-6-one). Functional studies indicate that phosphoinositide (PIns) hydrolysis in TE671/RD cells is increased by carbachol (EC50 of 10 microM), but not by nicotine (to concentrations as high as 1 mM). Agonist-stimulated PIns metabolism is inhibited by antagonists with the same rank order potency as for inhibition of {sup 3}HQNB binding. Functional responses are augmented in the presence of a nonhydrolyzable GTP analog, are strongly inhibited after 24-hr exposure to cholera toxin, but are only slightly inhibited after long-term exposure to pertussis toxin or forskolin. These studies identify a pharmacologically-defined M3-subtype of mAChR strongly coupled via a cholera toxin-sensitive mechanism to PIns hydrolysis in these cells. Within 1 hr of treatment of TE671/RD cells with 1 mM dibutyryl cyclic AMP or with 10 microM phorbol-12-myristate-13-acetate (PMA), there is a 30 to 50% decrease in carbachol-stimulated PIns responsiveness that recovers to control values after 5 days of continued drug treatment. However, a comparable and more persistent inhibition of mAChR function is observed on cell treatment with 20 nM PMA.

  2. Thermodynamics of antagonist binding to rat muscarinic M2 receptors: antimuscarinics of the pridinol, sila-pridinol, diphenidol and sila-diphenidol type.

    PubMed Central

    Waelbroeck, M.; Camus, J.; Tastenoy, M.; Lambrecht, G.; Mutschler, E.; Kropfgans, M.; Sperlich, J.; Wiesenberger, F.; Tacke, R.; Christophe, J.


    1. We studied the effect of temperature on the binding to rat heart M2 muscarinic receptors of antagonists related to the carbon/silicon pairs pridinol/sila-pridinol and diphenidol/sila-diphenidol (including three germanium compounds) and six structurally related pairs of enantiomers [(R)- and (S)-procyclidine, (R)- and (S)-trihexyphenidyl, (R)- and (S)-tricyclamol, (R)- and (S)-trihexyphenidyl methiodide, (R)- and (S)-hexahydro-diphenidol and (R)- and (S)-hexbutinol]. Binding affinities were determined in competition experiments using [3H]-N-methyl-scopolamine chloride as radioligand. The reference drugs were scopolamine and N-methyl-scopolamine bromide. 2. The affinity of the antagonists either increased or decreased with temperature. van't Hoff plots were linear in the 278-310 degrees K temperature range. Binding of all antagonists was entropy driven. Enthalpy changes varied from large negative values (down to -29 kJ mol-1) to large positive values (up to +30 kJ mol-1). 3. (R)-configurated drugs had a 10 to 100 fold greater affinity for M2 receptors than the corresponding (S)-enantiomers. Enthalpy and entropy changes of the respective enantiomers were different but no consistent pattern was observed. 4. When silanols (R3SiOH) were compared to carbinols (R3COH), the affinity increase caused by C/Si exchange varied between 3 and 10 fold for achiral drugs but was negligible in the case of chiral drugs. Silanols induced more favourable enthalpy and less favourable entropy changes than the corresponding carbinols when binding. Organogermanium compounds (R4Ge) when compared to their silicon counterparts (R4Si) showed no significant difference in affinity as well as in enthalpy and entropy changes. 5. Exchange of a cyclohexyl by a phenyl moiety was associated with an increase or a decrease in drug affinity (depending on the absolute configuration in the case of chiral drugs) and generally also with a more favourable enthalpy change and a less favourable entropy change

  3. [Dmt(1)]DALDA is highly selective and potent at mu opioid receptors, but is not cross-tolerant with systemic morphine.


    Riba, Pal; Ben, Yong; Nguyen, Thi M-D; Furst, Susanna; Schiller, Peter W; Lee, Nancy M


    The clinical effectiveness of morphine is limited by several side effects, including the development of tolerance and dependence. Most of these side effects are believed to be mediated by central opioid receptors; therefore, hydrophilic opioids, which don't cross the blood-brain barrier, may have advantages over morphine in some clinical applications. We recently synthesized several analogues of DALDA (Tyr-D-Arg-Phe-Lys-NH2), a highly hydrophilic peptide derived from the endogenous opioid peptide dermorphin; all of them, particularly [Dmt(1)] DALDA (Dmt - 2',6'-dimethyl tyrosine), had high potency and selectivity at mu receptors, the target of morphine, in activity assays. Here we report the pharmacological characterization of [Dmt(1)] DALDA in the whole animal. [Dmt(1)]DALDA was 40 times more potent than morphine in inducing antinociception in mice when both drugs were given s.c., and 6-14 times more potent than DAMGO, a selective m agonist, when both drugs were given it. However, [Dmt(1)]DALDA showed poor cross-tolerance to morphine; thus chronic morphine treatment of animals increased the antinociceptive AD(50) of systemic [Dmt(1)]DALDA two fold or less, as compared to an 8-9-fold increase for morphine and a 4-5-fold increase for DAMGO. The antinociceptive activity of [Dmt(1)]DALDA (i.t) was blocked by CTAP, a selective mu antagonist, but not by TIPP psi, a selective delta antagonist, nor by nor-BNI, a selective kappa antagonist. [Dmt(1)]DALDA-induced antinociception was also blocked by naloxone methiodide, an antagonist that does not cross the blood-brain barrier, when agonist and antagonist were given i.t. or i.c.v., but not when they were given s.c. We conclude that [Dmt(1)] DALDA is a highly potent analgesic acting at mu receptors. Though it appears to penetrate the blood-brain barrier, it exhibits low cross-tolerance to morphine, suggesting that it may have advantages over the latter in certain clinical applications.

  4. Role of medullary GABA signal transduction on parasympathetic reflex vasodilatation in the lower lip.


    Kawakami, So; Izumi, Hiroshi; Masaki, Eiji; Kuchiiwa, Satoshi; Mizuta, Kentaro


    In the orofacial area, noxious stimulation of the orofacial structure in the trigeminal region evokes parasympathetic reflex vasodilatation, which occurs via the trigeminal spinal nucleus (Vsp) and the inferior/superior salivatory nucleus (ISN/SSN). However, the neurotransmitter involved in the inhibitory synaptic inputs within these nuclei has never been described. This parasympathetic reflex vasodilatation is suppressed by GABAergic action of volatile anesthetics, such as isoflurane, sevoflurane, and halothane, suggesting that medullary GABAergic mechanism exerts its inhibitory effect on the parasympathetic reflex via an activation of GABA receptors. The aim of the present study was to determine the role of GABA(A) and GABA(B) receptors in the Vsp and the ISN in regulating the lingual nerve (LN)-evoked parasympathetic reflex vasodilatation in the lower lip. Under urethane anesthesia (1g/kg), change in lower lip blood flow elicited by electrical stimulation of the LN was recorded in cervically vago-sympathectomized rats. Microinjection of GABA (10 μM; 0.3 μl/site) into the Vsp or the ISN significantly and reversibly attenuated the LN-evoked parasympathetic reflex vasodilatation. Microinjection of the GABA(A) receptor-selective agonist muscimol (100 μM; 0.3 μl/site) or the GABA(B) receptor-selective agonist baclofen (100 μM; 0.3 μl/site) into the Vsp or the ISN significantly and irreversibly reduced this reflex vasodilatation, and these effects were attenuated by pretreatment with microinjection of each receptor-selective antagonists [GABA(A) receptor selective antagonist bicuculline methiodide (1mM; 0.3 μl/site) or GABA(B) receptor selective antagonist CGP-35348 (1mM; 0.3 μl/site)] into the Vsp or the ISN. Microinjection of these antagonists alone into the Vsp or the ISN had no significant effect on this reflex vasodilatation. In addition, microinjection (0.3 μl/site) of the mixture of muscimol (100 μM) and baclofen (100 μM) into the Vsp or the ISN also

  5. Ventral tegmental area neurons are either excited or inhibited by cocaine’s actions in the peripheral nervous system

    PubMed Central

    Mejías-Aponte, Carlos A.; Kiyatkin, Eugene A.


    Cocaine’s multiple pharmacological substrates are ubiquitously present in the peripheral and central nervous system. Thus, upon its administration, cocaine acts in the periphery before directly acting in the brain. We determined whether cocaine alters ventral tegmental area (VTA) neuronal activity via peripheral actions, and whether this precedes its central actions. In urethane-anesthetized rats, we recorded VTA neurons responses to intravenous injections of two cocaine analogs: cocaine-hydrochloride (HCl, 0.25 mg/kg) that readily cross the blood-brain barrier (BBB) and cocaine-methiodide (MI, 0.33 mg/kg) that does not cross the BBB. Both cocaine analogs produced sustained changes in discharge rates that began 5s after the initiation of a 10s drug infusion. Within the first 90s post-injection the magnitudes of neuronal responsive of both cocaine analogs were comparable, but later in time the effects of cocaine-HCl were stronger and persisted longer than those of cocaine-MI. The proportion of neurons responsive to cocaine-HCl was twice to that of cocaine-MI (74% and 35% respectively). Both analogs also differed in the response onsets. Cocaine-MI rarely evoked responses after 1 min whereas cocaine-HCl continued to evoke responses within 3 min post-injection. VTA neurons were either excited or inhibited by both cocaine analogs. Most units responsive to cocaine-MI, regardless of excitation or inhibition, had electrophysiological characteristics of putative DA neurons. Units inhibited by cocaine-HCl also had characteristic of DA neurons whereas excited neurons had widely varying action potential durations and discharge rates. Cocaine-MI and cocaine-HCl each produced changes in VTA neuron activity under full DA receptor blockade. However, the duration of inhibition was shortened, the number of excitations increased, and they occurred with an earlier onset during DA receptor blockade. These findings indicate that cocaine acts peripherally with a short latency and

  6. Intrinsic and Network Mechanisms Constrain Neural Synchrony in the Moth Antennal Lobe.


    Lei, Hong; Yu, Yanxue; Zhu, Shuifang; Rangan, Aaditya V


    Projection-neurons (PNs) within the antennal lobe (AL) of the hawkmoth respond vigorously to odor stimulation, with each vigorous response followed by a ~1 s period of suppression-dubbed the "afterhyperpolarization-phase," or AHP-phase. Prior evidence indicates that this AHP-phase is important for the processing of odors, but the mechanisms underlying this phase and its function remain unknown. We investigate this issue. Beginning with several physiological experiments, we find that pharmacological manipulation of the AL yields surprising results. Specifically, (a) the application of picrotoxin (PTX) lengthens the AHP-phase and reduces PN activity, whereas (b) the application of Bicuculline-methiodide (BIC) reduces the AHP-phase and increases PN activity. These results are curious, as both PTX and BIC are inhibitory-receptor antagonists. To resolve this conundrum, we speculate that perhaps (a) PTX reduces PN activity through a disinhibitory circuit involving a heterogeneous population of local-neurons, and (b) BIC acts to hamper certain intrinsic currents within the PNs that contribute to the AHP-phase. To probe these hypotheses further we build a computational model of the AL and benchmark our model against our experimental observations. We find that, for parameters which satisfy these benchmarks, our model exhibits a particular kind of synchronous activity: namely, "multiple-firing-events" (MFEs). These MFEs are causally-linked sequences of spikes which emerge stochastically, and turn out to have important dynamical consequences for all the experimentally observed phenomena we used as benchmarks. Taking a step back, we extract a few predictions from our computational model pertaining to the real AL: Some predictions deal with the MFEs we expect to see in the real AL, whereas other predictions involve the runaway synchronization that we expect when BIC-application hampers the AHP-phase. By examining the literature we see support for the former, and we perform some

  7. Acetylcholine mediates the release of IL-8 in human bronchial epithelial cells by a NFkB/ERK-dependent mechanism.


    Profita, Mirella; Bonanno, Anna; Siena, Liboria; Ferraro, Maria; Montalbano, Angela M; Pompeo, Flora; Riccobono, Loredana; Pieper, Michael P; Gjomarkaj, Mark


    Acetylcholine may play a role in cell activation and airway inflammation. We evaluated the levels of both mRNA and protein of muscarinic M(1), M(2), M(3) receptors in human bronchial epithelial cell line (16HBE). 16HBE cells were also stimulated with acetylcholine and extracellular signal-regulated kinase1/2 (ERK1/2) and NFkB pathway activation as well as the IL-8 release was assessed in the presence or absence of the inhibitor of Protein-kinase (PKC) (GF109203X), of the inhibitor of mitogenic activated protein-kinase kinase (MAPKK) (PDO9805), of the inhibitor of kinaseB-alpha phosphorilation (pIkBalpha) (BAY11-7082), and of muscarinic receptor antagonists tiotropium bromide, 4-Diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP), telenzepine, gallamine. Additionally, we tested the IL-8-mediated neutrophil chemotactic activity of 16HBE supernatants stimulated with acetylcholine in the presence or absence of tiotropium. 16HBE cells expressed both protein and mRNA for muscarinic M(3), M(2) and M(1) receptors with levels of muscarinic M(3) receptor>muscarinic M(1) receptor>muscarinic M(2) receptor. Acetylcholine (10 microM) significantly stimulated ERK1/2 and NFkB activation as well as IL-8 release in 16HBE cells when compared to basal values. Furthermore, while the use of tiotropium, 4-DAMP, GF109203X, PDO98059, BAY11-7082 completely abolished these events, the use of telenzepine and gallamine were only partially able to downregulate these effects. Additionally, acetylcholine-mediated IL-8 release from 16HBE cells significantly increased chemotaxis toward neutrophils and this effect was blocked by tiotropium. In conclusion, acetylcholine activates the release of IL-8 from 16HBE involving PKC, ERK1/2 and NFkB pathways via muscarinic receptors, suggesting that it is likely to contribute to IL-8 related neutrophilic inflammatory disorders in the airway. Thus, muscarinic antagonists may contribute to control inflammatory processes in airway diseases.

  8. Differential Effects of the Gβ5-RGS7 Complex on Muscarinic M3 Receptor–Induced Ca2+ Influx and Release

    PubMed Central

    Karpinsky-Semper, Darla; Volmar, Claude-Henry; Brothers, Shaun P.


    The G protein β subunit Gβ5 uniquely forms heterodimers with R7 family regulators of G protein signaling (RGS) proteins (RGS6, RGS7, RGS9, and RGS11) instead of Gγ. Although the Gβ5-RGS7 complex attenuates Ca2+ signaling mediated by the muscarinic M3 receptor (M3R), the route of Ca2+ entry (i.e., release from intracellular stores and/or influx across the plasma membrane) is unknown. Here, we show that, in addition to suppressing carbachol-stimulated Ca2+ release, Gβ5-RGS7 enhanced Ca2+ influx. This novel effect of Gβ5-RGS7 was blocked by nifedipine and 2-aminoethoxydiphenyl borate. Experiments with pertussis toxin, an RGS domain–deficient mutant of RGS7, and UBO-QIC {L-threonine,(3R)-N-acetyl-3-hydroxy-L-leucyl-(aR)-a-hydroxybenzenepropanoyl-2,3-idehydro-N-methylalanyl-L-alanyl-N-methyl-L-alanyl-(3R)-3-[[(2S,3R)-3-hydroxy-4- methyl-1-oxo-2-[(1-oxopropyl)amino]pentyl]oxy]-L-leucyl-N,O-dimethyl-,(7→1)-lactone (9CI)}, a novel inhibitor of Gq, showed that Gβ5-RGS7 modulated a Gq-mediated pathway. These studies indicate that Gβ5-RGS7, independent of RGS7 GTPase-accelerating protein activity, couples M3R to a nifedipine-sensitive Ca2+ channel. We also compared the action of Gβ5-RGS7 on M3R-induced Ca2+ influx and release elicited by different muscarinic agonists. Responses to Oxo-M [oxotremorine methiodide N,N,N,-trimethyl-4-(2-oxo-1-pyrrolidinyl)-2-butyn-1-ammonium iodide] were insensitive to Gβ5-RGS7. Pilocarpine responses consisted of a large release and modest influx components, of which the former was strongly inhibited whereas the latter was insensitive to Gβ5-RGS7. McN-A-343 [(4-hydroxy-2-butynyl)-1-trimethylammonium-3-chlorocarbanilate chloride] was the only compound whose total Ca2+ response was enhanced by Gβ5-RGS7, attributed to, in part, by the relatively small Ca2+ release this partial agonist stimulated. Together, these results show that distinct agonists not only have differential M3R functional selectivity, but also confer specific

  9. Neural mechanism of activity spread in the cat motor cortex and its relation to the intrinsic connectivity.


    Capaday, Charles; van Vreeswijk, Carl; Ethier, Christian; Ferkinghoff-Borg, Jesper; Weber, Doug


    Motor cortical points are linked by intrinsic horizontal connections having a recurrent network topology. However, it is not known whether neural activity can propagate over the area covered by these intrinsic connections and whether there are spatial anisotropies of synaptic strength, as opposed to synaptic density. Moreover, the mechanisms by which activity spreads have yet to be determined. To address these issues, an 8 × 8 microelectrode array was inserted in the forelimb area of the cat motor cortex (MCx). The centre of the array had a laser etched hole ∼500 μm in diameter. A microiontophoretic pipette, with a tip diameter of 2-3 μm, containing bicuculline methiodide (BIC) was inserted in the hole and driven to a depth of 1200-1400 μm from the cortical surface. BIC was ejected for ∼2min from the tip of the micropipette with positive direct current ranging between 20 and 40 nA in different experiments. This produced spontaneous nearly periodic bursts (0.2-1.0 Hz) of multi-unit activity in a radius of about 400 μm from the tip of the micropipette. The bursts of neural activity spread at a velocity of 0.11-0.24 ms⁻¹ (mean=0.14 mm ms⁻¹, SD=0.05)with decreasing amplitude.The area activated was on average 7.22 mm² (SD=0.91 mm²), or ∼92% of the area covered by the recording array. The mode of propagation was determined to occur by progressive recruitment of cortical territory, driven by a central locus of activity of some 400 μm in radius. Thus, activity did not propagate as a wave. Transection of the connections between the thalamus and MCx did not significantly alter the propagation velocity or the size of the recruited area, demonstrating that the bursts spread along the routes of intrinsic cortical connectivity. These experiments demonstrate that neural activity initiated within a small motor cortical locus (≤ 400 μm in radius) can recruit a relatively large neighbourhood in which a variety of muscles acting at several forelimb joints are

  10. Down-regulation of muscarinic receptors and the m3 subtype in white-footed mice by dietary exposure to parathion

    USGS Publications Warehouse

    Jett, D.A.; Hill, E.F.; Fernando, J.C.; Eldefrawi, M.E.; Eldefrawi, A.T.


    The effect of ad libitum dietary exposure (as occurs in the field) to parathion for 14 d was investigated on the muscarinic acetylcholine receptor (mAChR) in brains and submaxillary glands of adults of a field species, the white-footed mouse Peromyscus leucopus. Immunoprecipitation using subtype selective antibodies revealed that the relative ratios of the m1-m5 mAChR subtypes in Peromyscus brain were similar to those in rat brain. There was little variability in acetylcholinesterase (AChE) activity in control mice brains but large variability in 39 exposed mice, resulting from differences in food ingestion and parathion metabolism. Accordingly, data on radioligand binding to mAChRs in each mouse brain were correlated with brain AChE activity in the same mouse, and AChE inhibition served as a biomarker of exposure reflecting in situ paraoxon concentrations. Exposure to parathion for 14 d reduced maximal binding (Bmax) of [3H]quinuclidinyl benzilate ([3H]QNB), [3H]-N-methylscopolamine ([3H]NMS), and [3H]-4-diphenylacetoxy-N-methylpiperidine methiodide ([3H]-4-DAMP) by up to approximately 58% without affecting receptor affinities for these ligands. Maximal reduction in Bmax of [3H]QNB and [3H]-4-DAMP binding occurred in mice with highest AChE inhibition, while equivalent maximal reduction in Bmax of [3H]NMS occurred in mice with only approximately 10% AChE inhibition, without further change at higher parathion doses. This is believed to be due to the hydrophilicity of [3H]NMS, which limits its accessibility to internalized desensitized receptors. In submaxillary glands (mAChRs are predominantly m3 subtype), there were significant dose-dependent reductions in [3H]QNB binding and m3 mRNA levels in exposed mice, revealed by Northern blot analyses. The reduction in m3 receptors is suggested to result mostly from reduced synthesis at the transcription level, rather than from translational or posttranslational events. The data suggest that down-regulation of mAChRs occurs

  11. Mechanisms underlying orientation selectivity of neurons in the primary visual cortex of the macaque.

    PubMed Central

    Sato, H; Katsuyama, N; Tamura, H; Hata, Y; Tsumoto, T


    1. Effects of blocking intracortical inhibition by microiontophoretic administration of bicuculline methiodide (BMI), a selective antagonist for GABAA receptors, on orientation selectivity of 109 neurones were studied in the primary visual cortex (V1) of anaesthetized and paralysed monkeys. 2. The averaged orientation tuning of visual responses of cells was poor in cytochrome oxidaserich blobs of layer II/III and in layer IVc beta, moderate in layers IVb, IVc alpha and V, and sharp in the interblob region of layer II/III and in layers IVa and VI. 3. Iontophoretic administration of BMI reduced the sharpness of orientation tuning of cells to a varying extent in each layer. In most cells, furthermore, the originally ineffective stimuli induced visual responses during the BMI administration, suggesting that excitatory inputs evoked by the non-optimally oriented stimuli were masked by GABAergic inhibition. Nevertheless, the maximal facilitation was observed in the response to the optimally or near-optimally oriented stimuli. 4. There was a difference in such an effect of BMI among layers. Orientation selectivity of cells in interblobs in layer II/III and in layer IVb was sensitive to BMI whereas that of cells in layer VI was relatively insensitive to BMI, suggesting a larger contribution of excitatory mechanisms to the orientation selectivity in this layer. 5. In the orientation-selective cells, an analysis of the magnitude of excitation and inhibition evoked by stimuli at various orientations suggests that both inputs tune around the optimal orientation and their magnitudes are almost proportional to each other except at the optimal orientation. This analysis also indicates that the orientation tuning of inhibition had a less prominent peak around the optimal orientation than that of excitation. This dominance of excitation over inhibition around the optimal orientation may function to accentuate the response to the optimally oriented stimulus. 6. These results suggest

  12. Pharmacological comparison of the cloned human and rat M2 muscarinic receptor genes expressed in the murine fibroblast (B82) cell line.


    Kovacs, I; Yamamura, H I; Waite, S L; Varga, E V; Roeske, W R


    The coding sequence of the human m2 receptor gene was amplified by polymerase chain reaction and stably transfected into a murine fibroblast cell line (B82). We have compared the human M2 clonal cell line (HM2-B10) with the previously established B82 cell line (M2LKB2-2) expressing the rat M2 receptor to assess drug specificity, drug selectivity and effector coupling. Both transfected cell lines showed a high level of specific, saturable [3H](-)-N-methyl-3-quinuclidinyl benzilate binding with Kd values of 243 pM (155-352 pM) and 345 pM (234-539 pM) and Bmax values of 97 +/- 4 and 338 +/- 16 fmol/10(6) cells, respectively. Inhibition of [3H](-)-N-methyl-3-quinuclidinyl benzilate binding to HM2-B10 cells and M2LKB2-2 cells showed the same rank order of potency for the antagonists: atropine > dexetimide > 4-diphenylacetoxy-N-methylpiperidine methiodide > himbacine > methoctramine > 11-[[2-[(diethylamino) methyl]-1-piperidinyl]acetyl]-5,11-dihidro-6H-pyrido-[2,3-b](1, 4)-benzodiazepine-6-one > hexahydro-sila-difenidol hydro-chloride > pirenzepine. Correlation analysis of the pKi values indicate that the expressed human and rat M2 receptors have nearly identical ligand-binding characteristics. Carbachol inhibited forskolin-stimulated cAMP formation with similar potency in both cell lines [EC50 = 2.4 microM (0.2-2.8) and 1.1 microM (0.2-5.3) for the human and rat M2 receptor, respectively]. In the M2LKB2-2 cells, carbachol slightly stimulated the [3H]inositol monophosphate formation but had no significant effect in HM2-B10 cells. In conclusion, the human and rat M2 receptors expressed in the B82 cell line have very similar binding properties but exhibit slight differences in effector coupling mechanisms. PMID:9454790

  13. Intrinsic and Network Mechanisms Constrain Neural Synchrony in the Moth Antennal Lobe

    PubMed Central

    Lei, Hong; Yu, Yanxue; Zhu, Shuifang; Rangan, Aaditya V.


    Projection-neurons (PNs) within the antennal lobe (AL) of the hawkmoth respond vigorously to odor stimulation, with each vigorous response followed by a ~1 s period of suppression—dubbed the “afterhyperpolarization-phase,” or AHP-phase. Prior evidence indicates that this AHP-phase is important for the processing of odors, but the mechanisms underlying this phase and its function remain unknown. We investigate this issue. Beginning with several physiological experiments, we find that pharmacological manipulation of the AL yields surprising results. Specifically, (a) the application of picrotoxin (PTX) lengthens the AHP-phase and reduces PN activity, whereas (b) the application of Bicuculline-methiodide (BIC) reduces the AHP-phase and increases PN activity. These results are curious, as both PTX and BIC are inhibitory-receptor antagonists. To resolve this conundrum, we speculate that perhaps (a) PTX reduces PN activity through a disinhibitory circuit involving a heterogeneous population of local-neurons, and (b) BIC acts to hamper certain intrinsic currents within the PNs that contribute to the AHP-phase. To probe these hypotheses further we build a computational model of the AL and benchmark our model against our experimental observations. We find that, for parameters which satisfy these benchmarks, our model exhibits a particular kind of synchronous activity: namely, “multiple-firing-events” (MFEs). These MFEs are causally-linked sequences of spikes which emerge stochastically, and turn out to have important dynamical consequences for all the experimentally observed phenomena we used as benchmarks. Taking a step back, we extract a few predictions from our computational model pertaining to the real AL: Some predictions deal with the MFEs we expect to see in the real AL, whereas other predictions involve the runaway synchronization that we expect when BIC-application hampers the AHP-phase. By examining the literature we see support for the former, and we

  14. Spatiotemporal dynamics of excitation in rat insular cortex: intrinsic corticocortical circuit regulates caudal-rostro excitatory propagation from the insular to frontal cortex.


    Fujita, S; Adachi, K; Koshikawa, N; Kobayashi, M


    The insular cortex (IC), composing unique anatomical connections, receives multi-modal sensory inputs including visceral, gustatory and somatosensory information from sensory thalamic nuclei. Axonal projections from the limbic structures, which have a profound influence on induction of epileptic activity, also converge onto the IC. However, functional connectivity underlying the physiological and pathological roles characteristic to the IC still remains unclear. The present study sought to elucidate the spatiotemporal dynamics of excitatory propagation and their cellular mechanisms in the IC using optical recording in urethane-anesthetized rats. Repetitive electrical stimulations of the IC at 50 Hz demonstrated characteristic patterns of excitatory propagation depending on the stimulation sites. Stimulation of the granular zone of the IC (GI) and other surrounding cortices such as the motor/primary sensory/secondary sensory cortices evoked round-shaped excitatory propagations, which often extended over the borders of adjacent areas, whereas excitation of the agranular and dysgranular zones in the IC (AI and DI, respectively) spread along the rostrocaudal axis parallel to the rhinal fissure. Stimulation of AI/DI often evoked excitation in the dorsolateral orbital cortex, which exhibited spatially discontinuous topography of excitatory propagation in the IC. Pharmacological manipulations using 6,7-dinitroquinoxaline-2,3(1H,4H)-dione (DNQX), a non-NMDA receptor antagonist, D-2-amino-5-phosphonovaleric acid (D-APV), an NMDA receptor antagonist, and bicuculline methiodide, a GABA(A) receptor antagonist, indicate that excitatory propagation was primarily regulated by non-NMDA and GABA(A) receptors. Microinjection of lidocaine or incision of the supragranular layers of the rostrocaudally middle part of excitatory regions suppressed excitation in the remote regions from the stimulation site, suggesting that the excitatory propagation in the IC is largely mediated by

  15. GABA and glutamate receptors in the horizontal limb of diagonal band of Broca (hDB): effects on cardiovascular regulation.


    Nasimi, Ali; Hatam, Masoumeh


    The horizontal limb of diagonal band of Broca (hDB) is a part of the limbic system. It has been shown that microinjection of L-glutamate into the hDB elicited cardiovascular depressive responses in anesthetized rats and pressor effect in unanesthetized rats. But the role of glutamate receptor subtypes has not yet been investigated. In addition the role of the GABAergic system of the hDB in cardiovascular responses is not known. Therefore, we examined the cardiovascular responses elicited by glutamate and GABA receptors in the hDB by using their agonists and antagonists. Drugs (50 nl) were microinjected into the hDB of anaesthetized rats. Blood pressure and heart rate were recorded before and throughout each experiment. The average changes in the mean arterial pressure and heart rate at different intervals were compared both within each case group and between the case and control groups using repeated measures of ANOVA. Microinjection of GABA(A) receptor antagonist, bicuculline methiodide (BMI, 1 mM) increased both the mean arterial pressure and heart rate, and muscimole, a GABA(A) agonist (500 pmol) caused a significant decrease in the mean arterial pressure and heart rate. Microinjection of L-glutamate (0.25 M) into the hDB resulted in a maximum decrease of the mean arterial pressure of 24.4 +/- 3.7 mmHg and heart rate of 25.2 +/- 3.08 beats/min. Injection of AP5, an antagonist of glutamate NMDA receptor (1 and 2.5 mM), and CNQX, an antagonist of glutamate AMPA receptor (0.5 and 1 mM) caused small, nonsignificant changes of the heart rate and the blood pressure. Either AP5 or CNQX when coinjected with glutamate abolished the depressor effect of glutamate, suggesting that simultaneous activation of both glutamate receptors is necessary for the effect of glutamate to emerge. The depressor effect of the glutaminergic system of the hDB on the cardiovascular system was similar to the previous studies. For the first time, the effects of CNQX, AP5, BMI, and muscimole

  16. Amino acids modify thalamo-cortical response transformation expressed by neurons of the ventrobasal complex.


    Vahle-Hinz, C; Hicks, T P; Gottschaldt, K M


    The hypothesis has been tested that inhibitory mechanisms, active spatially and temporally between the input and the output of thalamic neurons, determine the nature of the information transmitted to the cerebral cortex. To enable this assessment, in barbiturate-anesthetized cats and urethane-anesthetized rats juxtacellular recordings were performed together with microiontophoretic ejection of transmitter agonists and antagonists. The effects of these drugs were studied on responses evoked by mechanical stimulation of cutaneous receptive fields (RFs) of neurons in the thalamic ventrobasal complex (VB). Neurons from different parts of the VB were investigated: 29 units were located medially, in the ventral posteromedial nucleus (VPM; facial RFs), and 11 units were located laterally, in the ventral posterolateral nucleus (VPL; forepaw and body RFs). A further eleven VB units had no detectable RF. Twenty-six neurons were tested with electrical stimulation of the somatosensory cortex (SI), 17 of these being identified as thalamo-cortical relay neurons and 5 being classified as presumed interneurons; the remaining 4 could not be activated. Four additional recordings were from trigemino-thalamic or thalamo-cortical fibers. For the quantitative assessment of the neurons' input and output, neuronal activity was induced by feedback-controlled, mechanical trapezoidal and/or sinusoidal stimuli applied to sinus hairs, fur or skin and the numbers of prepotentials and soma spikes were compared in peristimulus time histograms (PSTHs) generated simultaneously for both types of signal from 'DC' recordings. Iontophoretic administration of excitatory amino acids (EAAs) or bicuculline methiodide (BMI) increased output-input ratios in 87% of the cases tested, due to a higher rate of conversion of prepotentials into soma spikes taking place. In cases of neurons exhibiting a sustained-to-transient response pattern, changes to sustained-to-sustained patterns were demonstrated. Tests with

  17. Thermodynamic and kinetic control of charged, amphiphilic triblock copolymer assembly via interaction with organic counterions in solvent mixtures

    NASA Astrophysics Data System (ADS)

    Cui, Honggang


    assembly process include THF/water ratio, PS block length, the type and amount of organic counterions, and the mixing pathway. Their formation mechanism has been investigated from three aspects: (i) the chain structure of organic counterions, including spacer length, chain hydrophobicity between ionizable groups and the number of ionizable groups (amine group); (ii) molecular structure of the triblock copolymer, including block length of polystyrene and chain architecture; (iii) relative variation of the components, such as different ratios of THF to water and the different ratios of amine groups to acid groups. The first example of a novel micelle formed was the toroidal micelle. The toroidal micelle morphology, which is theoretically predicted but rarely observed, has been produced by the self assembly of PAA99- b-PMA73-b-PS66 in combination with 2,2-(ethylenedioxy)diethylamine (EDDA) and mixed THF/H2O solvent. It was found that toroids can be constructed by two mechanisms: elimination of energetically unfavored cylindrical micelle endcaps or perforation of disk-like micelles. Three-fold junctions were formed as intermediate structures to facilitate toroidal formation from cylindrical micelles. In order to construct toroids from cylindrical micelles, three requirements must be met: lower bending modulus (flexibility of cylinders), selfattraction between cylinders, and extra endcapping energy originating from chain packing frustration. Extremely high energy spheres can also fuse into toroids. Disk-like micelles can transform into a toroidal morphology when cylindrical packing geometry was initiated along the rims of disk-like micelles via solvent mixing that eventually perforated the disk center. The toroidal morphology can be kinetically trapped by either ridding the system of organic solvent or chemically crosslinking the PAA corona with EDDA via addition of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide methiodide (DPEM). The interaction of positively