Animal models of epilepsy are used to understand the pathophysiology of seizures and to develop new therapies to treat epilepsy ^3,14 . Penicillin is one of the most widely used drugs for inducing experimental seizures. With the administration of intracortical penicillin, γ-aminobutyric acid (GABA) type-A receptors are inhibited and this suppresses the chlorine entry inside the neurons resulting in focal seizure focus which is recorded from the surface of the cortex via ECoG ^2,3 .

Free radical production causes the accumulation of excitatory neurotransmitter glutamic acid and a decrease in GABA, an inhibitory neurotransmitter, in the cerebrum ¹⁴ . Free oxygen radicals are thought to play important roles in the formation of epilepsy and neuronal death following seizures ^5,15 . Obay et al. declared that in the cerebrum, MDA levels were increased, while SOD and GSH levels were decreased after pentylenetetrazole-induced seizures in rats ¹⁵ . Parallel to this study, we found an increase in MDA levels and a decrease in GPx and SOD levels observed in the cerebrum and cerebellum tissues of rats after penicillin-induced epileptiform activity.

Leptin has been found to increase nerve cell excitability in some studies ^3,7 . Arslan et al. reported that leptin increased epileptiform activity in the experimental epilepsy model induced by penicillin in rats ³ . Moreover, leptin increases excitability by increasing intracellular calcium and synaptic transmission ⁷ . Kutlu et al. reported that lipid peroxidase and glutathione levels were decreased in the cerebrum as a result of leptin administration ¹⁶ . Furthermore, the external application of leptin did not change MDA levels in a glial cell culture study ¹⁷ . We determined that the leptin administration to the non-epileptic animals did not have a significant effect on oxidative stress. However, leptin significantly increased the MDA levels in the cerebrum and cerebellum tissues of the rats compared to the penicillin group and significantly decreased the SOD and GPx levels suggesting that leptin may show its proconvulsant effect through oxidative stress probably by increasing intracellular Ca⁺² levels.

Cannabinoids CB1 and CB2 receptors are G protein-coupled receptors and belong to different families of cell membrane-bound proteins. CB1 receptors inhibit presynaptic N and P/Q type Ca⁺² channels and activate inflow rectifying K⁺ channels ¹⁸ . CB1 receptors are found extensively in the cerebrum, especially in the cerebral cortex, hippocampus, basal ganglia, and cerebellum ¹⁹ . Studies so far have shown that cannabinoids exert their behavioral and neuronal effects through cerebrum CB1 receptor activation ²⁰ . On the other hand, cannabinoids have been known to exert anticonvulsant effects through CB1 receptors ^2,3,21,22 . In the epileptic model created with pentylenetetrazole, ACEA was found to have an antiepileptic effect ⁹ and inhibited the proconvulsant effect of toxoplasmosis ²² .

Synthetic cannabinoids are full agonists that bind to CB1 and CB2 receptors with a higher potency and affinity ²³ . Since cannabinoids are known to exert their anticonvulsant effects on the CB1 receptor, selective CB1 receptor agonists and antagonists were used in the present study. For this reason, ACEA was preferred as the CB1 agonist and AM251 as the antagonist. Systemically administered cannabinoids undergo various pharmacokinetic interactions until they reach the cerebrum ²⁰ . Therefore, we preferred to administer cannabinoids by i.c.v. route. Di Giacomo et al. showed that cannabidiol and cannabigerol have neuroprotective and antioxidant effects in rat astrocytes and isolated cortexes ²⁴ . Marsicano et al. showed that cannabinoids have strong antioxidant effects in cerebellar granule cell cultures by inhibiting cell excitability with increasing K⁺ permeability and decreasing Ca⁺² permeability through the CB1 receptor ²⁵ .

In the present study, ACEA administration to the penicillin-injected rats decreased the MDA levels and increased SOD and GPx levels in the cerebrum and cerebellum tissues compared with the penicillin group. When AM251 was administered after penicillin, AM251 enhanced the activity of penicillin on MDA, SOD and GPx levels. Based on the data obtained, it is thought that ACEA, at a dose of 7.5 μg, acts on CB1 receptors presynaptically and reduces intracellular Ca⁺² levels, preventing oxidative stress and thus suppressing the epileptiform activity. Thus, AM251 is thought to have the opposite effect of ACEA by increasing the intracellular Ca⁺² level, increasing oxidative stress, and thus increasing the epileptiform activity induced by penicillin.

Recent studies have suggested that there may be a relationship between leptin and cannabinoids ^10-12 . Co-administration of ACEA and leptin significantly increased oxidative stress compared with the alone ACEA after penicillin injection. So, it was concluded that leptin reversed the effect of ACEA on oxidative stress. Combined administration of AM251 and leptin on our epileptic model no significant change was found in terms of oxidative stress compared to the AM251-administered group. From a pharmacological point of view, AM251 and leptin, which have proconvulsant effects when administered separately, are expected to significantly enhance oxidative stress when administered together. However, no significant differences were found. We suggested that AM251 and leptin have identical effects on oxidative stress by using similar pathways on Ca⁺². Although our study revealed that leptin and cannabinoids exert their effects on epilepsy through the oxidative system, further molecular and biochemical studies are needed to further elucidate the molecular mechanisms. Findings underlying these interactions may lead to the development of new therapeutic strategies for the treatment of epilepsy.