Ascorbic acid and nitric oxide are known to play important roles in epilepsy. The aim of present study was to identify the involvement of nitric oxide (NO) in the anticonvulsant effects of ascorbic acid on penicillin-induced epileptiform activity in rats. Intracortical injection of penicillin (500, International Units (IU)) into the left sensorimotor cortex induced epileptiform activity within 2-5 min. Thirty minutes after penicillin injection, nitric oxide synthase (NOS) inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME, 100 mg/kg), neuronal nitric oxide synthase (nNOS) inhibitor 7-nitroindazole (7-NI, 40 mg/kg), NO Substrate, L-arginine (500 mg/kg) were administered with the most effective dose of ascorbic acid (100 mg/kg) intraperitoneally (i.p.), The administration Of L-arginine significantly decreased the frequency of epileptiform activity while administration of L-NAME did not influence the mean frequency of epileptiform activity. Injection of 7-NI decreased the mean frequency of epileptiform activity but did not influence amplitude. Ascorbic acid decreased both the mean frequency and amplitude of penicillin-induced epileptiform activity in rats. The application of L-NAME partially and temporarily reversed the anticonvulsant effects of ascorbic acid. The results Support the hypothesis of neuro-protective role for NO and ascorbic acid. The protective effect of ascorbic acid against epileptiform activity was partially and temporarily reversed by nonspecific nitric oxide synthase inhibitor L-NAME, but not selective neuronal nitric oxide synthase inhibitor 7-NI, indicating that ascorbic acid needs endothelial-NOS/NO route to decrease penicillin-induced epileptiform activity. (C) 2009 British Epilepsy Association. Published by Elsevier Ltd. All rights reserved.