Evidence implicating alpha-2 adrenergic receptors in the anticonvulsant action of intranigral muscimol

The effects of systemically administered catecholamine receptor antagonists on the anticonvulsant action produced by local application of a gamma-aminobutyric acid agonist, muscimol, to the substantia nigra of rats were studied. Both electroshock and kindled seizures were studied. Two alpha-2 receptor antagonists, idazoxan and yohimbine, blocked the anticonvulsant action of intranigral muscimol in the electroshock model. Neither a beta nor an alpha-1 adrenergic nor a dopamine-2 receptor antagonist blocked this action. In contrast to electroshock seizures, an alpha-2 antagonist only partially reversed the anticonvulsant action of intranigral muscimol in the kindling model. We interpret the data to indicate that the interaction of norepinephrine with alpha-2 receptors is required for the anticonvulsant action of intranigral muscimol in the electroshock model. The only partial reversal found in the kindling model suggests that nigral control of seizure propagation involves more than alpha-2 receptor mediated neurotransmission.     

Regulation of kindling epileptogenesis by hippocampal galanin type 1 and type 2 receptors: The effects of subtype-selective agonists and the role of G-protein-mediated signaling

The search for antiepileptic drugs that are capable of blocking the progression of epilepsy (epileptogenesis) is an important problem of translational epilepsy research. The neuropeptide galanin effectively suppresses acute seizures. We examined the ability of hippocampal galanin receptor type 1 (GalR1) and type 2 (GalR2) to inhibit kindling epileptogenesis and studied signaling cascades that mediate their effects. Wistar rats received 24-h-long intrahippocampal infusion of a GalR1/2 agonist galanin(1-29), GalR1 agonist M617 [galanin(1-13)-Gln14-bradykinin(2-9)-amide], or GalR2 agonist galanin(2-11). The peptides were administered alone or combined with an inhibitor of Gi protein pertussis toxin (PTX), Gi-protein activated K+ channels (GIRK) inhibitor tertiapin Q (TPQ), G(q/11) protein inhibitor [D-Arg1,D-Trp(5,7,9),Leu11]-substance P (dSP), or an inhibitor of intracellular Ca2+ release dantrolene. Sixteen hours into drug delivery, the animals were subjected to rapid kindling-60 electrical trains administered to ventral hippocampus every 5 min. M617 delayed epileptogenesis, whereas galanin(1-29) and galanin(2-11) completely prevented the occurrence of full kindled seizures. TPQ abolished anticonvulsant effect of M617 but not of galanin(2-11). PTX blocked anticonvulsant effects of M617 and inversed the action of galanin(1-29) and galanin(2-11) to proconvulsant. dSP and dantrolene did not modify seizure suppression through GalR1 and GalR2, but eliminated the proconvulsant effect of PTX + galanin(1-29) and PTX + galanin(2-11) combinations. We conclude that hippocampal GalR1 exert their disease-modifying effect through the Gi-GIRK pathway. GalR2 is antiepileptogenic through the Gi mechanism independent of GIRK. A secondary proconvulsant pathway coupled to GalR2 involves G(q/11) and intracellular Ca2+. The data are important for understanding endogenous mechanisms regulating epileptogenesis and for the development of novel antiepileptogenic drugs.     

Prolonged attenuation of amygdala-kindled seizure measures in rats by convection-enhanced delivery of the N-type calcium channel antagonists omega-conotoxin GVIA and omega-conotoxin MVIIA

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

Alpha-2 receptors mediate an endogenous noradrenergic suppression of kindling development

We sought to elucidate the receptor subtype through which endogenous norepinephrine suppresses epileptogenesis in the rat kindling model. To this end we examined the effects of systemically administered selective antagonists and an alpha-2 agonist on kindling development and on kindled seizures. The alpha-2 adrenergic antagonists idazoxan, yohimbine and rauwolscine (0.1-10.0 mg/kg i.p.) dose-dependently facilitated amygdala kindling development. Central administration of idazoxan (20 micrograms/40 microliter i.c.v.) produced an equivalent facilitation. The facilitation was selective for alpha-2 antagonists because neither the alpha-1 antagonist corynanthine (0.1-10.0 mg/kg i.p.), nor the beta antagonist propranolol (0.1-10.0 mg/kg i.p.), nor the selective beta-1 antagonist ICI 89,406 (10 micrograms/40 microliter i.c.v.) nor the beta-2 antagonist ICI 118,551 (0.5-5.0 mg/kg i.p.) modified kindling development. The alpha-2 agonist clonidine (0.01-0.2 mg/kg i.p.) dose-dependently suppressed kindling development. In contrast to the effects on kindling development, neither the alpha-2 antagonists nor clonidine modified seizures elicited from previously kindled animals. We interpret the data to indicate that endogenous norepinephrine suppresses kindling development by activation of postsynaptic alpha-2 receptors. The selective inhibition of kindling development, but not kindled seizures, suggests that alpha-2 agonists may be effective antiepileptogenic, but not anticonvulsant, agents.     

Dantrolene ameliorates delayed cell death and concomitant DNA fragmentation in the rat hippocampal CA1 neurons subjected to mild ischemia.

The present study investigated whether dantrolene, which inhibits the Ca(2+) release from the intracellular Ca(2+) store sites, reduced nuclear DNA fragmentation and produced neuronal protection in a model of global forebrain ischemia. Male Wistar rats were subjected to four-vessel occlusion (4VO) for 5 min and then infused continuously with dantrolene or vehicle into the cerebral ventricle for 3 days. The intact rats did not undergo any intervention. The number of viable and DNA nick-end-labeled neurons in the hippocampal CA1 were evaluated 4 days after the ischemia. The number of viable neurons in the dantrolene-treated rats was significantly higher than that in the vehicle-treated rats and lower than that in the intact animals (P<0.01 and <0.05, respectively). The number of DNA nick-end-labeled nuclei was significantly lower in dantrolene-treated rats compared with the vehicle-treated animals (P<0.0001). No nick-end labeling was observed in the intact animals. A linear correlation was found between the number of viable cells and nick-end labeled nuclei in the CA1 (r=0.91, P<0.0001). These results suggest that the postischemic intraventricular dantrolene is effective in precluding neuronal death and concomitant nuclear DNA fragmentation following transient global ischemia.     

Anticonvulsant action of the beta-carboline abecarnil: studies in rodents and baboon, Papio papio

Abecarnil (ZK 112119; isopropyl-6-benzyloxy-4-methoxymethyl-beta-carboxylate) is a metabolically stable beta-carboline derivative with potent anxiolytic and few sedative and ataxic effects in rodents. The anticonvulsant and muscle relaxant actions of abecarnil have been evaluated in mice, rats, gerbils and baboons. Abecarnil raised the threshold for tonic electroconvulsions in mice after corneal but not after auricular application, had no effect on maximal electroshock-induced tonic convulsions triggered by either method, protected mice against the tonic hindlimb extension in PTZ-, picrotoxin- and 3-mercaptopropionate-induced seizures and blocked clonus after PTZ, DMCM (methyl-4-ethyl-6,7-dimethoxy-9H-pyrido-(3,4-b)-indol-3-carboxylate) and 3-mercaptopropionate. Abecarnil had no effect on convulsions induced by bicuculline and strychnine. Furthermore, abecarnil blocked kindled seizures after chronic administration of PTZ and FG 7142 (beta-carboline-3-carboxylic acid methylamide) and protected mice and rats against limbic convulsions induced by pilocarpine. Severity and afterdischarge duration of amygdala-kindled seizures were reduced in rats treated with abecarnil. Abecarnil also antagonized selectively convulsions induced by i.c.v. administration of kainate, but not those triggered by N-methyl-D-aspartate or quisqualate. In genetic models of reflex epilepsy, abecarnil was effective against sound-induced convulsions in DBA/2 mice, against air blast-induced generalized seizures in gerbils and against myoclonus in baboons Papio papio. The anticonvulsant effect of abecornil in a PTZ seizure model in mice was potentiated by ethosuximide, whereas no significant potentiation was found with diazepam, clonazepam, diphenylhydantoin, carbamazepine and phenobarbital. Electromyographic monitoring in a etorphine model of muscle rigidity in rats showed no or little muscle relaxant effect of abecarnil.(ABSTRACT TRUNCATED AT 250 WORDS)     

Wine polyphenols modulate calcium handling in rat aorta: involvement of nitric oxide pathway

The effects of short-term oral administration of red wine polyphenolic compounds (RWPCs) on blood pressure and vascular reactivity were investigated in rats. The consequence of RWPCs treatment on agonist-induced contractility of rat aorta with respect to Ca2+ handling was assessed, by examining both intracellular Ca2+ store and extracellular Ca2+ influx components of the response. Rats were treated daily for 7 days by intragastric administration of either 5% glucose, or RWPCs (20 mg/kg) [from two different sources, i.e. Provinols (SFD, Vallont Pont d'Arc, France) and RWPC1 (INRA, Montpellier, France)]. Administration of these compounds produced a decrease in systolic blood pressure. The consequence of RWPCs treatment on vascular smooth muscle was investigated in rat aorta without endothelium exposed to noradrenaline. In Ca(2+)-free medium, RWPC1 but not Provinols treatment induced an increase in noradrenaline-induced contraction. After depletion of intracellular Ca2+ stores by noradrenaline in Ca(2+)-free medium, addition of CaCl2 in the continuous presence of agonist induced an increase in contraction, which was not significantly different between control, Provinols- and RWPC-treated rats. The presence of an inhibitor of sarcoplasmic reticulum Ca(2+)-ATPase, thapsigargin, significantly reduced noradrenaline-induced contraction in Ca(2+)-free medium in RWPCs-treated aorta, as compared to that of control. Interestingly, the inhibitory effect of thapsigargin on the response linked to the release of Ca2+ from internal stores in RWPCs-treated vessels was completely prevented in the presence of NO-synthase inhibitor, L-nitro arginine methyl ester, the inhibitor of guanylyl cyclase, oxadiazolo-quinoxaline or the protein kinase G inhibitor, 8-Bromoguanosine-3'-5-cyclic mono-phosphorothioate, Rp isomer. These results suggest that short-term administration of RWPCs in rats induced subtle alteration of thapsigargin-sensitive component of agonist-induced contraction in rat aorta linked to Ca2+ release from intracellular store. Calcium release from intracellular stores sensitive to thapsigargin was implicated in this mechanism. The prevention of the inhibitory effect of thapsigargin by the inhibitors of NO/cyclic guanosine monophosphate pathway after RWPCs treatment highlights the role of NO in this phenomenon.     

Kindling as a model of drug-resistant partial epilepsy: selection of phenytoin-resistant and nonresistant rats.

Complex partial seizures comprise the major uncontrolled seizure type in adult patients with epilepsy. Any improvement of our understanding of the mechanisms through which these seizures are often refractory to antiepileptic drugs is therefore of considerable importance. By examining the effects of the anti-epileptic drug phenytoin in a large group of kindled rats, a widely used model of complex partial seizures, animals with different sensitivity to this drug were selected. Using determination of the focal seizure threshold for evaluation of phenytoin's anticonvulsant effect, most animals (about 60%) showed variable effects in response to phenytoin. However, about 20% of the animals ("phenytoin nonresponders") showed no increase in their focal seizure threshold at repeated test trials with phenytoin, and 20% ("phenytoin responders") exhibited reproducible increases in focal seizure threshold after injection of phenytoin. Phenytoin responders and nonresponders thus selected were used for subsequent experiments. The different response of focal seizures to phenytoin was not related to differences in pharmacokinetics or location of the stimulating electrode. Although phenytoin reproducibly increased the threshold for induction of afterdischarges in responders, it did not alter severity or duration of the elicited seizure response. In contrast to phenytoin, carbamazepine induced increases in focal seizure threshold in all kindled rats. Duration of seizures and afterdischarges were significantly reduced by carbamazepine in phenytoin responders, but not in nonresponders, although plasma levels of carbamazepine were the same in both groups. The difference in response of kindled rats to phenytoin was restricted to kindled seizures, because phenytoin induced the same anticonvulsant effect on the threshold for generalized tonic electroconvulsions (determined via transauricular electrodes) in both groups of kindled rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anticonvulsant and behavioral effects of two novel competitive N-methyl-D-aspartic acid receptor antagonists, CGP 37849 and CGP 39551, in the kindling model of epilepsy. Comparison with MK-801 and carbamazepine.

The orally active competitive N-methyl-D-aspartate (NMDA) receptor antagonists CGP 37849 (DL-[E]-2-amino-4-methyl-5-phosphono-3-pentenoic acid) and its ethyl ester CGP 39551 were evaluated in amygdala-kindled rats, a model for complex partial and secondarily generalized seizures. Anticonvulsant and behavioral effects of these novel compounds were compared with those of the noncompetitive NMDA receptor antagonist MK-801 [(+)-5-methyl-10,11-dihydroxy-5H-dibenzo(a,d)cyclohepten-5,10-imin e] and the antiepileptic drug carbamazepine, one of the major drugs for treatment of partial and generalized seizures in humans. For comparative evaluation, the compounds were injected i.p. at the following doses: 1 to 10 mg/kg (CGP 37849 or CGP 39551), 0.05 to 0.3 mg/kg (MK-801) and 20 to 40 mg/kg (carbamazepine), respectively. In contrast to carbamazepine, CGP 37849, CGP 39551 and MK-801 exerted only weak anticonvulsant effects in fully kindled rats and did not increase the focal seizure threshold. The weak anticonvulsant effects of the NMDA receptor antagonists in kindled rats were associated with profound untoward behavioral effects. The behavioral syndrome induced by the NMDA receptor antagonists in kindled rats was characterized by marked ataxia, hyperactivity and, in case of CGP 37849 and MK-801, stereotypies, such as head weaving. The low or absent effectiveness of the novel NMDA receptor antagonists against kindled seizures suggests that these compounds will not be clinically useful antiepileptics against partial and secondarily generalized seizures. Furthermore, in view of the recent clinical findings on psychotomimetic effects of MK-801 in epileptic patients, the similarities in the excitatory effects produced by CGP 39551, CGP 37849 and MK-801 in kindled rats may indicate that competitive NMDA receptor antagonists may also produce psychotomimetic effects in humans.     

Anticonvulsant activity of the low-affinity uncompetitive N-methyl-D- aspartate antagonist (+-)-5-aminocarbonyl-10,11-dihydro-5H- dibenzo[a,d]cyclohepten-5,10-imine (ADCI): comparison with the structural analogs dizocilpine (MK-801) and carbamazepine

(+-)-5-Aminocarbonyl-10,11-dihydro-5H-dibenzo[a,d] [a,d]cyclohepten-5,10-imine (ADCI), a tricyclic compound structurally related to dizocilpine (MK-801) and carbamazepine, was a potent anticonvulsant in the mouse maximal electroshock seizure test when administered i.p. (ED50, 8.9 mg/kg) or p.o. (ED50, 23.5 mg/kg), but failed to cause motor impairment except at substantially higher doses (TD50 values, 49.2 mg/kg i.p. and 293 mg/kg p.o.). ADCI was also protective against chemically induced seizures in mice, including those produced by 4-aminopyridine (ED50, 7.1 mg/kg s.c.) and pentylenetetrazol (ED50, 37.4 mg/kg s.c.). In addition, ADCI antagonized the behavioral effects and lethality of s.c. administered N-methyl-D-aspartate (NMDA: ED50, 15.2 mg/kg), but was a weaker antagonist of kainate-induced clonic seizures (ED50, 33.0 mg/kg), indicating that the drug is a selective functional NMDA antagonist. In common with other NMDA antagonists, ADCI retarded the development of amygdaloid kindled seizures in rats, but failed to attenuate the afterdischarge duration in fully kindled animals. Whole cell voltage clamp recordings from cultured hippocampal neurons demonstrated that ADCI selectively blocks inward current responses to NMDA in a use-dependent fashion without affecting responses to kainate or quisqualate, indicating that ADCI is a selective open channel (uncompetitive) blocker of the NMDA receptor-ionophore complex. ADCI blocked NMDA-evoked inward current responses with a potency (IC50, 14 microM) similar to that with which it displaces [3H]-1-[1-(2-thienyl)-cyclohexyl]piperidine from binding to NMDA receptor channels in rat brain homogenates (IC50, 11.3 microM).(ABSTRACT TRUNCATED AT 250 WORDS)     

The antiepileptic drug levetiracetam decreases the inositol 1,4,5-trisphosphate-dependent [Ca2+]I increase induced by ATP and bradykinin in PC12 cells

The present study explores the hypothesis that the new anti-epileptic drug levetiracetam (LEV) could interfere with the inositol 1,4,5-trisphosphate (IP(3))-dependent release of intracellular Ca(2+) initiated by G(q)-coupled receptor activation, a process that plays a role in triggering and maintaining seizures. We assessed the effect of LEV on the amplitude of [Ca(2+)](i) response to bradykinin (BK) and ATP in single Fura-2/acetoxymethyl ester-loaded PC12 rat pheochromocytoma cells, which express very high levels of LEV binding sites. LEV dose-dependently reduced the [Ca(2+)](i) increase, elicited either by 1 microM BK or by 100 microM ATP (IC(50), 0.39 +/- 0.01 microM for BK and 0.20 +/- 0.01 microM for ATP; Hill coefficients, 1.33 +/- 0.04 for BK and 1.38 +/- 0.06 for ATP). Interestingly, although the discharge of ryanodine stores by a process of calcium-induced calcium release also took place as part of the [Ca(2+)](i) response to BK, LEV inhibitory effect was mainly exerted on the IP(3)-dependent stores. In fact, the drug was still effective after the pharmacological blockade of ryanodine receptors. Furthermore, LEV did not affect Ca(2+) stored in the intracellular deposits since it did not reduce the amplitude of [Ca(2+)](i) response either to thapsigargin or to ionomycin. In conclusion, LEV inhibits Ca(2+) release from the IP(3)-sensitive stores without reducing Ca(2+) storage into these deposits. Because of the relevant implications of IP(3)-dependent Ca(2+) release in neuron excitability and epileptogenesis, this novel effect of LEV could provide a useful insight into the mechanisms underlying its antiepileptic properties.     

Systemic administration of adenosine ameliorates pentylenetetrazol-induced chemical kindling and secondary behavioural and biochemical changes in mice

Adenosine is one of the inhibitory neuromodulators in the brain and is considered to be responsible for seizure arrest and postictal refractoriness. Adenosine, adenosine receptor agonists, and adenosine uptake blockers are known to reduce the severity and duration of amygdala-kindled seizures. The present study was carried out to elucidate the anticonvulsant and neuromodulatory effect of systemic adenosine on the pentylenetetrazol (PTZ)-induced chemical kindling in mice. Kindling was induced by chronic administration of a subconvulsive dose of PTZ (40 mg/kg, i.p.) on every other day for a total period of 9 days. Adenosine was administered daily, 30 min before PTZ or vehicle. The kindling score was recorded immediately following PTZ administration according to a prevalidated scoring scale. Various behavioral and biochemical estimations were performed on day 10 (i.e. 24 h after the last dose of PTZ). Chronic PTZ treatment progressively increased the seizure score with the maximum score reached on day 9. Behavioral analysis found hyperlocomotor activity, anxiogenic response, hyperalgesia and amnesia in kindled mice. Biochemical analysis revealed that chronic treatment with PTZ significantly increased lipid peroxidation (malondialdehyde levels), nitrite (NO(2-) levels), adenosine deaminase (ADA) and total RNA levels and decreased catalase, reduced glutathione (GSH) levels in brain homogenates, and a depletion of adrenal ascorbic acid. Daily treatment with adenosine (25 and 50 mg/kg, i.p.) for 9 days led to a significant decrease in PTZ-induced kindling score and also reversed various behavioral and biochemical alterations produced by PTZ. The results of the present study suggested that systemic adenosine administration reversed the behavioral and biochemical alterations induced by chronic PTZ.     

CM 40907: a structurally novel anticonvulsant in mice, rats and baboons

CM 40907 [3-(4-hydroxypiperidyl)-6-(2'-chlorophenyl)-pyridazine] is a chemically original compound which possesses the pharmacological properties of a potent, p.o. active anticonvulsant. The anticonvulsant activity of CM 40907 was examined in mice, rats and photosensitive Papio-papio baboons and compared to that of phenobarbital, diphenylhydantoin, carbamazepine, sodium valproate and ethosuximide. In mice, CM 40907 antagonized electroconvulsive shock and chemically induced seizures with an overall potency comparable to that of carbamazepine and a therapeutic ratio (ED50 rotorod/ED50 electroshock) superior to that of ethosuximide, sodium valproate, phenobarbital and carbamazepine. In the rat CM 40907 suppressed completed kindled amygdaloid seizures and was approximately as active as phenobarbital. In naturally photosensitive Senegalese Papio-papio baboons CM 40907 antagonized myoclonus and cortical paroxysmal discharges. In this model CM 40907 was approximately one-fourth as potent as phenobarbital, twice as potent as carbamazepine and 6 times more potent than sodium valproate. In mice CM 40907, at anticonvulsant doses, increased the affinity of [3H]flunitrazepam for its central receptor site. Based on these results it is postulated that CM 40907 is a potent and relatively nonsedative anticonvulsant and may be of therapeutic benefit in epileptic disorders.     

Age-Dependent Changes in the Effects of Amiodarone on Rabbit Cardiac Myocyte Contractions

BACKGROUND: Intravenous amiodarone has increasingly been used to control life-threatening atrial and ventricular arrhythmias. In addition to its four antiarrhythmic properties, amiodarone may have complex effects on intracellular Ca(2+) stores and myocyte contractility. METHODS AND RESULTS: Contraction amplitude was recorded for cardiac ventricular myocytes isolated from neonatal and adult rabbits. Sarcoplasmic reticulum (SR) Ca(2+) stores were loaded to steady-state levels by a train of eight electric field stimulations. The SR Ca(2+) load was quantified by recording the contraction amplitude resulting from the complete depletion of SR Ca(2+) stores by exposing the cell to a 1-second pulse of 10 mmol/L caffeine. After the cells were exposed to 1 μmol/L amiodarone for 10 minutes, electrically stimulated contraction amplitudes significantly decreased in both adult and neonatal cells. Caffeine-induced cell contraction amplitudes were not affected by amiodarone in adult ventricular myocytes. By contrast, amiodarone markedly inhibited caffeine-induced contractions in neonatal ventricular myocytes. The inhibitory effect of amiodarone on the caffeine-induced contractions was not replicated by Ca(2+) channel blockade with diltiazem. CONCLUSIONS: Amiodarone markedly inhibits caffeine-induced contraction in neonatal myocytes but has no significant effect on adult myocytes. Ca(2+) influx through amiodarone-sensitive Ca(2+) channels may play a primary role in maintaining SR Ca(2+) stores in neonatal heart.     

Chronic self-administration of nicotine in rats impairs T cell responsiveness

Chronic exposure of rodents to nicotine via subcutaneously or intracerebroventricularly implanted miniosmotic pumps affects T cell function. However, this method of continuous nicotine administration does not replicate the self-motivated administration of nicotine in human smokers. To determine whether nicotine impairs the immune system under conditions pertinent to human smokers, we investigated the T cell responsiveness of male Lewis rats self-administering (SA) nicotine (0.03 mg/kg of body weight per injection) 40 to 50 times/day for 5 weeks, using a model of virtually unlimited access to nicotine. Compared with sham control animals, the concanavalin A-induced proliferation of spleen cells from SA rats was significantly decreased. Moreover, the ability of spleen cells to mobilize intracellular Ca(2+) after ligation of the T cell antigen receptor (TCR) with an anti-alphabeta TCR antibody was significantly less in SA than in control rats. In addition, inositol 1,4,5-trisphosphate (IP(3))-sensitive intracellular Ca(2+) stores were markedly depleted in spleen cells from SA animals. These results suggest that chronic nicotine self-administration suppresses T cell responsiveness, and this suppression may result from an impaired TCR-mediated signaling that stems from the depletion of IP(3)-sensitive intracellular Ca(2+) stores.     

Responses to reversible anoxia of intracellular free and bound Ca(2+) in rat cortical slices

Severe anoxia induces destabilisation of intracellular calcium homeostasis in neurones. The mechanism of this effect, and particularly the interrelationship between changes in intracellular concentration of free Ca(2+) ions and the content of the intracellular Ca(2+) stores, during and after anoxia, is not clear. We used a superfusion system of rat olfactory cortical slices for the fluorimetric estimation of changes in the intracellular concentration of free Ca(2+) ions and in the level of bound Ca(2+), utilising the fluorescent indicators Fura-2 and chlortetracycline, respectively. It was found that 10-min normoglycaemic anoxia results in simultaneous decrease in bound and increase in free Ca(2+) levels, whereas during 60-min reoxygenation, we detected an increase in both indices. The NMDA receptor antagonists MK-801 and APV attenuated changes in free Ca(2+) level during anoxia and reoxygenation and intensified anoxia-evoked decrease in bound Ca(2+) content, whereas a late post-anoxic increase in bound Ca(2+) was abolished. These data suggest that the influx of extracellular Ca(2+) to neurones via NMDA receptors, plays a critical role in the rise of intracellular free Ca(2+) concentration during and after anoxia. Biphasic changes in bound Ca(2+) content during anoxia and reoxygenation may reflect an anoxia-induced release of Ca(2+) from intracellular stores, followed later by a neuronal calcium overload and refilling of intracellular Ca(2+) binding sites.     

Effect of magnesium sulfate administration on blood–brain barrier in a rat model of intraperitoneal sepsis: a randomized controlled experimental study

Rapid and efficient cooling is the most important therapeutic objective in patients with heat stroke (HS). This article reviews the mechanism of action and rationale for the use of dantrolene as a potential supportive cooling method in the treatment of HS. Relevant studies were included to support discussion of the role of dantrolene for the treatment of HS. In some studies dantrolene was shown to accelerate cooling rate when administered after the development of exertional HS. Dantrolene was also found to be effective in reducing the extent of HS signs when given as pretreatment in an animal model. Accumulated data do not support the routine use of dantrolene as an adjuvant cooling technique in HS, but administration of this drug in severe cases, or in which no improvement is observed, appears rational. Further trials are needed in order to assess the true effectiveness of dantrolene in HS.     

The mechanism of action of dantrolene sodium.

Dantrolene sodium is thought to affect some step in the excitation-contraction coupling process. Using a point voltage clamp to determine mechanical thresholds of individual fibers of frog, rat and goat skeletal muscle preparations, we found that dantrolene both raises (moves to more positive potentials) the rheobasic potential and increases the steepness of the strength-duration curve for mechanical threshold. The effect of raising the rheobase reaches a maximum at 1.2 x 10-5M dantrolene whereas the steepness effect occurs only at a saturating concentration (3.8x10-5M),indicating that dantrolene has two sites of action. The rheobase effect is absent below 18 degrees C (suggesting a phase transition) and it is competitively antagnoized by the calcium ionophore A23187 which can release calcium from the sarcoplasmic reticulum. The data indicate that dantrolene raises the rheobase by directly decreasing the release of calcium from the sarcoplasmic reticulum. We propose that dantrolene inhibits the movement of a natural calcium ionophore. Measurements of the voltage-dependent charge movement associated with excitation-contraction coupling showed that dantrolene did not significantly change the amount of charge moved. The time to peak of the ON current was not significantly changed, but that of the OFF current was significantly shortened. This action of dantrolene on the kinetics of the OFF current may account for its effect on the steepness of the strength-duration curve.     

Characterization of long-term potentiation of primary afferent transmission at trigeminal synapses of juvenile rats: essential role of subtype 5 metabotropic glutamate receptors

Recent work has demonstrated that a brief high-frequency conditioning stimulation to the primary afferent nerve fibers can induce a long-term potentiation (LTP) of synaptic transmission in neurons in the superficial layer of the trigeminal caudal nucleus; however, the cellular and molecular mechanisms underlying this synaptic potentiation remain unclear. Using both extracellular field potential and whole-cell patch-clamp recordings in brainstem parasagital slices of juvenile rat with the mandibular nerve attached, we show here that the induction of trigeminal primary afferent LTP: (1) does not require the activation of ionotropic glutamate receptors; (2) is dependent on extracellular Ca(2+) and the release of Ca(2+) from intracellular stores; (3) is specifically prevented by the metabotropic glutamate receptor subtype 5 (mGluR5) antagonist 2-methyl-6-(phenylethynyl)pyridine but not the mGluR1 antagonist LY367385, group II mGluR antagonist LY341495 or group III mGluR antagonist MAP4; (4) is mimicked by the bath-applied group I mGluR agonist (S)-3,5-dihydroxyphenylglycine and mGluR5 agonist (RS)-2-chloro-5-hydroxyphenylglycine; (5) requires the activation of phospholipase C (PLC) and protein kinase C (PKC); and (6) is concomitantly with a decrease in paired-pulse depression. These results demonstrate that the activation of mGluR5 and in turn triggering a PLC/PKC-dependent signaling cascade may contribute to the induction of LTP of primary afferent synaptic transmission in the superficial layer of trigeminal caudal nucleus of juvenile rats. This may be relevant to the processing of nociceptive information.