arachidonyl-2-chloroethylamide and Seizures

The influence of arachidonyl-2'-chloroethylamide (ACEA - a selective cannabinoid CB1 receptor agonist) on the anticonvulsant potency and acute adverse-effect potentials of clobazam, lacosamide, and pregabalin was determined in the maximal electroshock-induced seizure model and chimney test in mice. ACEA (2.5 mg/kg, i.p.) significantly enhanced the anticonvulsant potency of pregabalin in the mouse maximal electroshock-induced seizure model by decreasing the median effective dose (ED50 ) of pregabalin from 125.39 to 78.06 mg/kg (P < 0.05). In contrast, ACEA (2.5 mg/kg) had no significant impact on the anticonvulsant potency of clobazam and lacosamide in the mouse maximal electroshock-induced seizure model. On the other hand, ACEA (2.5 mg/kg) did not affect acute adverse effects of clobazam, lacosamide or pregabalin, and the median toxic doses (TD50 ) for the studied anti-epileptic drugs in combination with ACEA did not differ from the TD50 values as determined for the drugs administered alone in the chimney test. In conclusion, ACEA ameliorates the pharmacological profile of pregabalin, when considering both the anticonvulsant and the acute adverse effects of the drug in preclinical study on animals. The combination of pregabalin with ACEA can be of pivotal importance for patients with epilepsy as a potentially advantageous combination if the results from this study translate into clinical settings.

Topics: Acetamides; Animals; Anticonvulsants; Arachidonic Acids; Ataxia; Benzodiazepines; Brain; Clobazam; Dose-Response Relationship, Drug; Drug Synergism; Electroshock; Lacosamide; Lethal Dose 50; Male; Mice; Pregabalin; Psychomotor Performance; Receptor, Cannabinoid, CB1; Seizures

Cannabinoid system plays a pivotal role in the seizure threshold modulation which is mainly mediated through activation of the cannabinoid CB₁ receptor. There is also several evidence of interaction between cannabinoid system and α₂-adrenoceptors in different paradigms. Using model of clonic seizure induced by intravenous pentylenetetrazole (PTZ) in male mice, we investigated whether α₂-adrenoceptors is involved in the effects of cannabinoids on the seizure threshold. Injection of the selective cannabinoid CB₁ agonist ACEA (2 mg/kg) significantly (P<0.01) increased the seizure threshold which was prevented by pretreatment with the selective CB1 antagonist AM251 (1 mg/kg, i.p.). The highest doses of clonidine, a α₂ receptor agonist, (1 and 5 mg/kg) showed anticonvulsant effects while yohimbine, a α₂ receptor antagonist, (0.01, 0.1, 1, and 10 mg/kg) did not induce any significant effect on PTZ seizure threshold. Pretreatment with clonidine (0.1 and 0.5 mg/kg) significantly reversed the anticonvulsant effect of ACEA (2 mg/kg). Yohimbine (0.1, 1, and 10 mg/kg) pretreatment of mice enhanced the clonic seizure threshold of ACEA (1 mg/kg), significantly. Combination of non-effective doses of AM251 (0.1 mg/kg) and clonidine (0.01 mg/kg) showed additive effect in blocking the anticonvulsant effect of ACEA (2 mg/kg). In conclusion, our findings demonstrated that α₂-adrenoceptors could be involved in the anticonvulsant properties of the specific cannabinoid CB₁ agonist ACEA, suggesting that CB₁ cannabinoid and α₂ receptors have functional interactions in modulation of clonic seizure threshold.

Topics: Animals; Anticonvulsants; Arachidonic Acids; Cannabinoids; Clonidine; Dose-Response Relationship, Drug; Drug Interactions; Male; Mice; Pentylenetetrazole; Piperidines; Pyrazoles; Receptor, Cannabinoid, CB1; Receptors, Adrenergic, alpha-2; Seizures; Yohimbine

It has been known for centuries that exogenous cannabinoids, such as tetrahydrocannabinol have anticonvulsant activity. Recent studies have advanced our understanding of the endogenous cannabinoid system and renewed the interest in cannabinoids as a potential treatment for epilepsy. The endogenous cannabinoid system is rapidly activated after seizure activity but still little is known about the molecular mechanisms underlying the role of the cannabinoid system in epilepsy. In this study epileptiform activity was induced by kainic acid (KA) and effects of the CB1 receptor agonists N-(2-Chloroethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide (ACEA) on G-protein signaling using the agonist-stimulated [(35)S]GTPγS binding assay were evaluated. Control and KA treated rat hippocampus and cortex membranes were used. Our results showed that the ACEA displayed a high potency and efficacy in stimulating the G-proteins and when compared to the control animals, significant enhancements were observed in tissues from the KA treated animals. Potency and efficacy values were in particular increased in the hippocampus tissues. Furthermore, gene expression levels of the cannabinoid receptor 1 (CB1) receptor and cannabinoid receptor interacting protein 1 (CRIP1) were measured by RT-PCR, where both CB1 and CRIP1 expressions were found to be elevated in the KA treated animals.

Topics: Animals; Arachidonic Acids; Cannabinoids; Carrier Proteins; Dose-Response Relationship, Drug; Gene Expression Regulation; GTP-Binding Proteins; Kainic Acid; Male; Rats; Rats, Sprague-Dawley; Receptor, Cannabinoid, CB1; Seizures

Endogenous cannabinoid ligands and cannabinoid CB1 receptor agonists have been shown to exert anticonvulsant effects in various experimental models of epilepsy. The purpose of this study was to determine the effects of arachidonyl-2'-chloroethylamide (ACEA-a highly selective cannabinoid CB1 receptor agonist) on the protective action of clonazepam, ethosuximide, phenobarbital, and valproate against pentylenetetrazole (PTZ)-induced clonic seizures in mice. To ascertain any pharmacokinetic contribution of ACEA to the observed interactions between tested drugs, free (non-protein bound) plasma and total brain concentrations of the antiepileptic drugs were estimated. Additionally, acute adverse-effect profiles of the combination of ACEA and different classical antiepileptic drugs (clonazepam, ethosuximide, phenobarbital and valproate) with respect to motor performance, long-term memory and skeletal muscular strength were measured. Results indicated that ACEA (10mg/kg, i.p.) co-administered with phenylmethylsulfonyl fluoride (PMSF-a substance protecting ACEA against degradation by the fatty-acid hydrolase; 30mg/kg, i.p.) significantly potentiated the anticonvulsant activity of ethosuximide, phenobarbital and valproate in the mouse PTZ-induced clonic seizure model by reducing their median effective doses (ED(50) values) from 122.8mg/kg to 71.7mg/kg (P<0.01; for ethosuximide), from 13.77mg/kg to 5.26mg/kg (P<0.05; for phenobarbital), and from 142.7mg/kg to 87.3mg/kg (P<0.05; for valproate), respectively. In contrast, ACEA (10mg/kg, i.p.) in combination with PMSF (30mg/kg, i.p.) had no impact on the protective action of clonazepam against PTZ-induced seizures in mice. However, ACEA (10mg/kg)+PMSF (30mg/kg) considerably increased free plasma and total brain concentrations of ethosuximide and valproate in mice suggesting a pharmacokinetic nature of interaction between drugs. In contrast, free plasma and total brain concentrations of clonazepam and phenobarbital remained unchanged after ACEA+PMSF administration and thus, indicating pharmacodynamic interactions. Moreover, none of the examined combinations of ACEA (10mg/kg, i.p.)+PMSF (30mg/kg, i.p.) with clonazepam, ethosuximide, phenobarbital, and valproate (at their ED(50) values from the PTZ-induced seizure test) affected motor coordination in the chimney test, long-term memory in the passive avoidance task, and muscular strength in the grip-strength test in mice, indicating no possible acute adverse effects in anim

Topics: Animals; Anticonvulsants; Arachidonic Acids; Avoidance Learning; Cannabinoid Receptor Agonists; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Synergism; Drug Therapy, Combination; Hand Strength; Male; Mice; Motor Activity; Pentylenetetrazole; Phenylmethylsulfonyl Fluoride; Seizures

The aim of this study was to determine the influence of arachidonyl-2'-chloroethylamide (ACEA - a highly selective cannabinoid type 1 [CB1] receptor agonist) on the protective action and acute adverse effects of carbamazepine, lamotrigine, oxcarbazepine, phenobarbital, phenytoin, and topiramate in the maximal electroshock seizure model and chimney test in mice. Tonic hind limb extension (seizure activity) was evoked in adult male albino Swiss mice by a current (sine-wave, 25 mA, 500 V, 50 Hz, 0.2s stimulus duration) delivered via auricular electrodes. Acute adverse-effect profiles of the studied antiepileptic drugs with respect to motor coordination was assessed in the chimney test. Additionally, long-term memory and skeletal muscular strength were measured along with free plasma (non-protein bound) and total brain antiepileptic drug concentrations. To inhibit the rapid metabolic degradation of ACEA by the fatty-acid amide hydrolase, phenylmethylsulfonyl fluoride (PMSF) was used at a constant ineffective dose of 30 mg/kg. Results indicate that ACEA (2.5 mg/kg, i.p.) co-administered with PMSF (30 mg/kg, i.p.), significantly enhanced the anticonvulsant activity of phenobarbital, but not that of carbamazepine, lamotrigine, oxcarbazepine, phenytoin, or topiramate in the maximal electroshock seizure test in mice. Moreover, ACEA (2.5 mg/kg) with PMSF (30 mg/kg) had no significant impact on the acute adverse effects of all examined antiepileptic drugs in the chimney test in mice. The protective index values (as quotients of the respective TD(50) and ED(50) values denoted from the chimney and maximal electroshock seizure tests, respectively) for the combinations of ACEA (2.5 mg/kg) and PMSF (30 mg/kg) with carbamazepine, oxcarbazepine, phenobarbital, and topiramate were greater than those denoted for the antiepileptic drugs administered alone. Only, the protective index values for the combination of ACEA (2.5 mg/kg) and PMSF (30 mg/kg) with lamotrigine and phenytoin were lower than those determined for the antiepileptic drugs administered alone. Pharmacokinetic experiments revealed that ACEA (2.5 mg/kg) and PMSF (30 mg/kg) affected neither free plasma (non-protein bound) nor total brain concentrations of phenobarbital in mice. Moreover, ACEA and PMSF in combination with carbamazepine, lamotrigine, oxcarbazepine, phenobarbital, phenytoin, and topiramate did not alter long-term memory or skeletal muscular strength in experimental animals. In conclusion, the enhance

Topics: Animals; Anticonvulsants; Arachidonic Acids; Avoidance Learning; Disease Models, Animal; Drug Combinations; Electroshock; Enzyme Inhibitors; Male; Memory Disorders; Mice; Muscle Strength; Muscle, Skeletal; Phenylmethylsulfonyl Fluoride; Psychomotor Performance; Receptor, Cannabinoid, CB1; Seizures

Cannabinoid system plays a pivotal role in the seizure threshold modulation which is mainly mediated through activation of the cannabinoid CB(1) receptor. There is also several evidence of interaction between cannabinoid system and other neurotransmitters including nitric oxide (NO) system. Using model of clonic seizure induced by pentylenetetrazole (PTZ) in male NMRI mice, we investigated whether NO is involved in the effects of cannabinoids on the seizure threshold. Injection of the selective cannabinoid CB(1) agonist ACEA (2mg/kg, i.p.) significantly (P<0.01) increased the seizure threshold which was prevented (P<0.001) by pretreatment with the selective CB(1) antagonist AM251 (1mg/kg, i.p.). The NO precursor l-arginine (50 and 100mg/kg, i.p.) potentiated the anticonvulsant effects of the sub-effective dose of ACEA (1mg/kg, i.p.). Pretreatment with non-effective doses of the non-specific NOS inhibitor l-NAME (15 and 30mg/kg, i.p.) and the specific neuronal NOS inhibitor 7-NI (40 and 80mg/kg, i.p.) but not the inducible NOS inhibitor aminoguanidine (10, 50 and 100mg/kg, i.p.) prevented the anticonvulsant effect of ACEA (2mg/kg, i.p.). Co-administration of non-effective dose of AM251 (0.5mg/kg) with both low and per se non-effective doses of l-NAME (1mg/kg, i.p.) and 7-NI (10mg/kg, i.p.) had significant (P<0.01) effect in preventing the anticonvulsant effect of ACEA (2mg/kg, i.p.). Our findings demonstrated that central NO system could be involved in the anticonvulsant properties of the specific cannabinoid CB(1) agonist ACEA, emphasizing on the interaction between two systems in the seizure modulation.

Topics: Analysis of Variance; Animals; Animals, Inbred Strains; Anticonvulsants; Arachidonic Acids; Arginine; Brain; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Male; Mice; NG-Nitroarginine Methyl Ester; Nitric Oxide; Pentylenetetrazole; Piperidines; Pyrazoles; Receptor, Cannabinoid, CB1; Seizures

Several results support the conclusion that the cannabinoid system has a role in generation and cessation of epileptic seizures. The aim of this study was to evaluate the effects of intracerebroventricular AM-251 [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide], a CB1-receptor antagonist, and ACEA (arachidonyl-2-chloroethylamide), a CB1-receptor agonist, on penicillin-induced epileptiform activity in rats.. In the first set of experiments, 30 min after penicillin injection, AM-251, at doses of 0.125, 0.25, 0.5, and 1 μg, was administered intracerebroventricularly (i.c.v.). In the second set of experiments, 30 min after penicillin injection, ACEA, at doses of 2.5, 5, 7.5, and 15 μg (i.c.v.), was administered. In the third set of experiments, AM-251, at doses of 0.125 and 0.25 μg (i.c.v.), was administered 10 min before ACEA (7.5 μg, i.c.v.) injection.. ACEA, at a dose of 7.5 μg, significantly decreased the frequency of penicillin-induced epileptiform activity without changing the amplitude. ACEA, at doses of 2.5, 5, and 15 μg, had no impact on either frequency or amplitude of epileptiform activity. AM-251, at doses of 0.25 and 0.50 μg, significantly increased the frequency of epileptiform activity. AM-251, at a dose of 0.25 μg (i.c.v.), was the most effective in changing the frequency of penicillin-induced epileptiform activity, and it also caused status epilepticus-like activity. AM-251, at doses of 0.125 and 0.25 μg, 10 min before ACEA (7.5 μg), reversed the anticonvulsant action of ACEA.. The results of the present study provide electrophysiologic evidence for the role of CB1 receptors in regulating the frequency of epileptiform activity in the model of penicillin-induced epilepsy. To elucidate the precise mechanism of cannabinoid action in the brain during seizure, more advanced electrophysiologic and neurochemical studies are required.

Topics: Animals; Anticonvulsants; Arachidonic Acids; Brain; Cannabinoids; Cerebral Cortex; Disease Models, Animal; Dose-Response Relationship, Drug; Electroencephalography; Epilepsy; Injections, Intraventricular; Male; Penicillins; Piperidines; Pyrazoles; Rats; Receptor, Cannabinoid, CB1; Seizures

Cannabinoid compounds are anticonvulsant since they have inhibitory effects at micromolar doses, which are mediated by activated receptors coupling to G(i/o) proteins. Surprisingly, both the analgesic and anticonvulsant effects of opioids are enhanced by ultra-low doses (nanomolar to picomolar) of the opioid antagonist naltrexone and as opioid and cannabinoid systems interact, it has been shown that ultra-low dose naltrexone also enhances cannabinoid-induced antinociception. Thus, concerning the seizure modulating properties of both classes of receptors this study investigated whether the ultra-low dose opioid antagonist naltrexone influences cannabinoid anticonvulsant effects. The clonic seizure threshold was tested in separate groups of male NMRI mice following injection of vehicle, the cannabinoid selective agonist arachidonyl-2-chloroethylamide (ACEA) and ultra-low doses of the opioid receptor antagonist naltrexone and a combination of ACEA and naltrexone doses in a model of clonic seizure induced by pentylenetetrazole (PTZ). Systemic injection of ultra-low doses of naltrexone (1pg/kg to 1ng/kg, i.p.) significantly potentiated the anticonvulsant effect of ACEA (1mg/kg, i.p.). Moreover, the very low dose of naltrexone (500pg/kg) unmasked a strong anticonvulsant effect for very low doses of ACEA (10 and 100microg/kg). A similar potentiation by naltrexone (500pg/kg) of anticonvulsant effects of non-effective dose of ACEA (1mg/kg) was also observed in the generalized tonic-clonic model of seizure. The present data indicate that the interaction between opioid and cannabinoid systems extends to ultra-low dose levels and ultra-low doses of opioid receptor antagonist in conjunction with very low doses of cannabinoids may provide a potent strategy to modulate seizure susceptibility.

Topics: Animals; Anticonvulsants; Arachidonic Acids; Cannabinoid Receptor Agonists; Cannabinoids; Convulsants; Dose-Response Relationship, Drug; Drug Synergism; Epilepsy, Generalized; Epilepsy, Tonic-Clonic; Injections, Intraperitoneal; Male; Mice; Naltrexone; Narcotic Antagonists; Pentylenetetrazole; Piperidines; Pyrazoles; Seizures

Several lines of evidence suggest that cannabinoid compounds are anticonvulsant since they have inhibitory effects at micromolar doses, which are mediated by activated receptors coupling to Gi/o proteins. Surprisingly, both the analgesic and anticonvulsant effects of opioids are enhanced by ultra-low doses (nanomolar to picomolar) of the opioid antagonist naltrexone and as opioid and cannabinoid systems interact, it has been shown that ultra-low dose naltrexone also enhances cannabinoid-induced antinociception. However, regarding the seizure modulating properties of both classes of receptors this study investigated whether ultra-low dose cannabinoid antagonist AM251 influences cannabinoid anticonvulsant effects. The clonic seizure threshold (CST) was tested in separate groups of male NMRI mice following injection of vehicle, the cannabinoid selective agonist arachidonyl-2-chloroethylamide (ACEA) and ultra-low doses of the cannabinoid CB1 antagonist AM251 and a combination of ACEA and AM251 doses in a model of clonic seizure induced by pentylenetetrazole (PTZ). Systemic administration of ultra-low doses of AM251 (10 fg/kg-100 ng/kg) significantly potentiated the anticonvulsant effect of ACEA at 0.5 and 1 mg/kg. Moreover, inhibition of cannabinoid induced excitatory signaling by AM251 (100 pg/kg) unmasked a strong anticonvulsant effect for very low doses of ACEA (100 ng/kg-100 microg/kg), suggesting that a presumed inhibitory component of cannabinoid receptor signaling can exert strong seizure-protective effects even at very low levels of cannabinoid receptor activation. A similar potentiation by AM251 (100 pg/kg and 1 ng/kg) of anticonvulsant effects of non-effective dose of ACEA (0.5 and 1 mg/kg) was also observed in the generalized tonic-clonic model of seizure. The present data suggest that ultra-low doses of cannabinoid receptor antagonists may provide a potent strategy to modulate seizure susceptibility, especially in conjunction with very low doses of cannabinoids.

Topics: Animals; Anticonvulsants; Arachidonic Acids; Cannabinoid Receptor Agonists; Cannabinoid Receptor Antagonists; Convulsants; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Synergism; Male; Mice; Pentylenetetrazole; Piperidines; Pyrazoles; Seizures

Endogenous cannabinoid ligands and cannabinoid CB(1) receptor agonists have been shown to exert potent anticonvulsant effects in various experimental models of epilepsy. The purpose of this study was to determine the effects of arachidonyl-2'-chloroethylamide (ACEA; N-(2-chloroethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide, a highly selective cannabinoid CB(1) receptor agonist) on the threshold for electroconvulsions and the anticonvulsant activity of valproate in the maximal electroshock-induced seizures in mice. To inhibit the rapid metabolic degradation of ACEA by the fatty-acid amide hydrolase, phenylmethylsulfonyl fluoride (PMSF) was used at a constant ineffective dose of 30 mg/kg (i.p.). Moreover, the effects of ACEA and PMSF on the acute adverse-effect profile of valproate were determined in the chimney test. Additionally, the adverse-effect potentials of combination of ACEA, PMSF with valproate were examined in the step-through passive avoidance task (long-term memory) and grip-strength test (neuromuscular strength). To ascertain any pharmacokinetic contribution of ACEA and PMSF to the observed interaction between tested drugs, both free (non-protein bound) plasma and total brain concentrations of valproate were estimated. Results indicated that ACEA (5 and 7.5 mg/kg; i.p.) combined with PMSF increased significantly (P<0.001) the electroconvulsive threshold in mice. ACEA at low doses of 1.25 and 2.5 mg/kg, i.p., with PMSF had no impact on threshold for electroconvulsions. Similarly, neither PMSF (30 mg/kg) nor ACEA (15 mg/kg) administered alone affected the electroconvulsive threshold in mice. Moreover, ACEA (at a subthreshold dose of 2.5 mg/kg; i.p.) co-administered with PMSF potentiated significantly the antielectroshock activity of valproate by reducing its ED(50) from 258.3 to 195.1 mg/kg (P<0.01). Isobolographic transformation of data revealed that the interactions between valproate and ACEA (at 1.25 and 2.5 mg/kg) combined with PMSF were additive. In the chimney test, the combination of ACEA (2.5 mg/kg) and PMSF (30 mg/kg) had no effect on acute adverse effect of valproate and its TD(50) (356.4 mg/kg) did not differ significantly from that for valproate administered alone (TD(50)=404.4 mg/kg). Moreover, none of the examined drugs administered either alone or in combinations produced long-term memory deficits in the step-through passive avoidance task and impaired neuromuscular strength in the grip-strength test in mice. In contrast, ACEA (2.5 mg/kg

Topics: Amidohydrolases; Animals; Anticonvulsants; Arachidonic Acids; Avoidance Learning; Brain; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Synergism; Electroshock; Enzyme Inhibitors; Forelimb; Male; Memory; Mice; Molecular Structure; Muscles; Phenylmethylsulfonyl Fluoride; Receptor, Cannabinoid, CB1; Seizures; Time Factors; Valproic Acid