In the present study, the possible protective effect of the pyridoxine against glutamate-induced neurotoxicity in cerebellar granular cell culture of rat pups is investigated for its therapeutic potential. Glutamate (10(-7) M) was administered to cerebellar granular cell cultures that were prepared from one-day-old Sprague-Dawley rats. The neuroprotective effect of pyridoxine was examined. Pyridoxine at the doses of 10(-8), 10(-7), 10(-6), and 10(-5) M was introduced into the culture flasks before inclusion of glutamate. Pyridoxine at the doses of 10(-8) M and 10(-7) M significantly reduced glutamate cytotoxicity. A 10(-7) M dose of pyridoxine proved to be more effective than a 10(-8) M dose. The present study demonstrates that pyridoxine may protect glutamate-induced neurotoxicity. Neuroprotective effect of pyridoxine, at least in part, may result from its anti-glutamatergic activity. Pyridoxine merits further investigation as a therapeutic option in hypoxic-ischemic brain injury.