The present study was conducted to identify the role of nitric oxide (NO) in the anticonvulsant effects of pyridoxine hydrochloride on penicillin-induced epileptiform activity in rats. A single microinjection of penicillin (500 units) into the left sensorimotor cortex induced epfleptiform activity within 2-4 min, progressing to full seizure activity lasting about 3-5 h. Thirty minutes after penicillin injection, 20, 40, 80, and 160 mg/kg of pyridoxine hydrochloride was administered intraperitoneatty (i.p.). Pyridoxine significantly reduced the frequency of penicittin-induced epileptiform activity. A low dose of pyridoxine (40 mg/kg) was the most effective in reducing both the frequency and amplitude of epileptiform activity. The effect of systemic administration of nitric oxide synthase (NOS) inhibitors, non-selective N-G-nitro-L-arginine methyl ester (L-NAME), selective neuronal NOS inhibitor, 7-nitroindazole (7-NI) and NO substrate, L-arginine on anticonvulsive effects of pyridoxine was investigated. The administration Of L-arginine (500 mg/kg, i.p.) and 7-NI (25 and 50 mg/kg, i.p.) significantly decreased the frequency of epileptiform electrocorticographical (ECoG) activity white administration of L-NAME (60 mg/kg, i.p.) and the inactive form of arginine (D-arginine) did not influence it. The administration Of L-NAME (60mg/kg, i.p.) 15 min before pyridoxine (40 mg/kg i.p.) application reversed the anticonvulsant effects of pyridoxine whereas 7-NI (25 and 50 mg/kg, i.p.) did not influence it. The same dose of its inactive enantiomer N-G-nitro-D-arginine methyl ester (D-NAME) failed to reverse the anticonvu[sant effects of pyridoxine. The administration Of L-arginine (500 mg/kg, i.p.) did not affect the frequency of epileptiform ECoG activity in the pyridoxine administered group. L-Arginine did not reverse the anticonvulsant effect of 7-NI in the penicillin and pyridoxine administered groups. The results of present study indicate that the inhibitory effect on the anticonvulsant activity of pyridoxine against penicillin-induced epileptiform activity was produced by L-NAME, not by 7-NI, and is probably not related to the decrease of NOS activity in the brain. (c) 2007 Elsevier B.V. All rights reserved.