The effects of 7-nitroindazole (7NI, a preferential neuronal nitric oxide synthase inhibitor) on the anticonvulsant activity of four second-generation antiepileptic drugs (AEDs: felbamate [FBM], lamotrigine [LTG], oxcarbazepine [OXC] and topiramate [TPM]) were studied in the mouse maximal electroshock-induced seizure (MES) model. Moreover, the influence of 7NI on the acute neurotoxic (adverse-effect) profiles of the studied AEDs, with regard to motor coordination, was determined in the chimney test in mice. Results indicate that 7NI (50 mg/kg; i.p.) significantly potentiated the anticonvulsant activity of OXC, but not that of FBM, LTG and TPM against MES-induced seizures and, simultaneously, it enhanced the acute neurotoxic effects of TPM, but not those of FBM, LTG and OXC in the chimney test in mice. 7NI at the lower dose of 25 mg/kg had no effect on the antiseizure activity and acute neurotoxic profiles of all investigated AEDs. Pharmacokinetic evaluation of interactions between 7NI and LTG, OXC and TPM against MES-induced seizures revealed no significant changes in free (non-protein bound) plasma AED concentrations following 7NI administration. Moreover, none of the examined combinations of 7NI with AEDs from the MES test were associated with long-term memory impairment in mice subjected to the step-through passive avoidance task. Based on our preclinical study, it can be concluded that only the combination of 7NI with OXC was beneficial, when considering its both anticonvulsant and acute neurotoxic effects. Moreover, the lack of impairment of long-term memory and no pharmacokinetic interactions in plasma of experimental animals make the combination of 7NI with OXC worthy of consideration for the treatment of patients with refractory epilepsy. The other combinations tested between 7NI and LTG, FBM and TPM were neutral, when considering their both anticonvulsant effects and acute neurotoxic profiles, therefore, no useful recommendation can be made for their clinical application.