Evaluation of interactions between cannabinoid compounds and diazepam in electroshock-induced seizure model in mice

Several studies have shown that cannabinoids have anticonvulsant properties that are mediated through activation of the cannabinoid CB1 receptors. In addition, endogenous cannabinoid compounds (endocannabinoids) regulate synaptic transmission and dampen seizure activity via activation of the same receptors. The aim of this study was to evaluate the possible interactions between antiepileptic effects of cannabinoid compounds and diazepam using electroshock-induced model of seizure in mice. Electroconvulsions were produced by means of an alternating current (ear-clip electrodes, fixed current intensity 35 mA, stimulus duration 0.2 s) and tonic hindlimb extension was taken as the endpoint. All experiments were performed on groups of ten mice and the number of animals who did not display seizure reported as percent protection. Intraperitoneal (i.p.) administration of diazepam (0.25-2 mg/kg) and CB1 receptor agonist WIN55212-2 (0.5-4 mg/kg) dose dependently produced an antiepileptic effect evaluated in terms of increased percentage of protection against electroshock-induced seizure. Logistic regression analysis indicated synergistic interactions in anticonvulsant action after co-administration of diazepam and WIN55212-2 in fixed-ratio combination of 3:1 (diazepam:WIN55212-2), while an additive effect was resulted after co-administration of 1:1 and 1:3 fixed-ratio combinations. Administration of various doses of the endocannabinoid reuptake inhibitor, AM404, did not produce any effect on electroshock-induced seizure. Moreover, co-administration of AM404 and diazepam did not produce significant interaction in antiepileptic properties of these compounds. Administration of the fatty acid amide hydrolase inhibitor, URB597, produced significant antiepileptic effect. Co-administration of URB597 and diazepam led to an antagonistic interaction in protection against shock-induced seizure. Co-administration of different doses of the cannabinoid CB1 receptor antagonist, AM251 did not alter the antiepileptic effect of diazepam in the electroshock-induced seizure test. These results demonstrate that endocannabinoid system participates in the modulation of seizure and combination of small doses of exogenous CB1 receptor agonists with diazepam may have effective consequences in seizure control. Furthermore, inhibiting the endocannabinoid degradation could be more efficacious in modulating seizure than preventing their uptake. This study also suggests that the effects of cannabinoids on epilepsy depend on the relative cannabinoid responsiveness of GABAergic and glutamatergic neurotransmission. While, the antiepileptic effects of cannabinoid compounds are likely by affecting excitatory glutamate neurotransmission, the antagonistic interaction between cannabinoid compounds and diazepam to protect seizure is due to the cannabinoid action on inhibitory GABAergic system.