Although for many years it has been proposed that megadoses of pyridoxine protect from ethanol toxicity, this issue remains unclear. In the present report the interaction between ethanol and pyridoxine was tested. Pyridoxine was administered intramuscularly or intracerebroventricularly to rats. Intramuscular administration of 187.2 mg/kg of pyridoxine displaced the ethanol-lethality dose curve significantly toward the right (p < 0.005) and increased the LD50 of ethanol from 4.46 to 5.19 g/kg (p < 0.005). Intracerebroventricular administration of pyridoxine (1.1 mg) completely suppressed the mortality due to a LD100 of ethanol and the effect was dose dependent. We conclude that pyridoxine is an effective treatment for ethanol intoxication. The results are discussed in terms of an interaction of ethanol and pyridoxine on the GABAergic system.