Ethanol Influences on Bax Translocation, Mitochondrial Membrane Potential, and Reactive Oxygen Species Generation Are Modulated by Vitamin E and Brain-Derived Neurotrophic Factor

Background: This study investigated ethanol influences on intracellular events that predispose developing neurons toward apoptosis and the capacity of the antioxidant alpha-tocopherol (vitamin E) and the neurotrophin brain-derived neurotrophic factor (BDNF) to modulate these effects. Assessments were made of the following: (i) ethanol-induced translocation of the pro-apoptotic Bax protein to the mitochondrial membrane, a key upstream event in the initiation of apoptotic cell death; (ii) disruption of the mitochondrial membrane potential (MMP) as a result of ethanol exposure, an important process in triggering the apoptotic cascade; and (iii) generation of damaging reactive oxygen species (ROS) as a function of ethanol exposure. Methods: These interactions were investigated in cultured postnatal day 8 neonatal rat cerebellar granule cells, a population vulnerable to developmental ethanol exposure in vivo and in vitro. Bax mitochondrial translocation was analyzed via subcellular fractionation followed by Western blot, and mitochondrial membrane integrity was determined using the lipophilic dye, JC-1, that exhibits potential-dependent accumulation in the mitochondrial membrane as a function of the MMP. Results: Brief ethanol exposure in these preparations precipitated Bax translocation, but both vitamin E and BDNF reduced this effect to control levels. Ethanol treatment also resulted in a disturbance of the MMP, and this effect was blunted by the antioxidant and the neurotrophin. ROS generation was enhanced by a short ethanol exposure in these cells, but the production of these harmful free radicals was diminished to control levels by cotreatment with either vitamin E or BDNF. Conclusions: These results indicate that both antioxidants and neurotrophic factors have the potential to ameliorate ethanol neurotoxicity and suggest possible interventions that could be implemented in preventing or lessening the severity of the damaging effects of ethanol in the developing central nervous system seen in the fetal alcohol syndrome (FAS).