Alcohol is a neuroteratogen; alcohol consumption during pregnancy may cause Fetal Alcohol Spectrum Disorders (FASD), among which, fetal alcohol syndrome (FAS) is the most severe form. The depletion of neurons in the developing CNS is the most deteriorating effect of ethanol. The loss of CNS neurons may underlie many of the behavioral deficits observed in FASD. Glycogen synthase kinase 3 (GSK3), a serine/threonine kinase, is an important mediator of neuron degeneration. We have demonstrated that ethanol activates GSK3, and the activation of GSK3 leads to neuronal death. Oxidative stress is also considered an important contributor to ethanol-induced neurotoxicity. We have purified a potent antioxidant from blackberries, cyanidin-3-glucoside (C3G). Our study indicates that C3G inhibits GSK3 activity and alleviates oxidative stress in cultured neuronal cells. In addition, C3G protects neuronal cells against ethanol-induced cell death. C3G-mediated neuroprotection is much more potent than other antioxidants and GSK3 inhibitors. C3G can cross the blood brain barrier (BBB) and distribute in the brain. C3G diminishes ethanol-induced activation of caspase-3 and Bax in the developing brain. Pharmacologically relevant concentrations of C3G are achievable through oral administration or intravenous (iv) injection in animals, and no adverse effect is observed. These findings suggest that C3G is a promising neuroprotective agent that may ameliorate/prevent ethanol-induced neuronal damage. We hypothesize that C3G's potent protection against ethanol-induced neuronal loss is mediated by the combined action of its antioxidant property and inhibition of pro-apoptotic signaling, GSK3/Bax pathways. To test the hypothesis, we will (1) investigate the antioxidant property of C3G and its metabolites, and their effects on GSK3 activity; (2) determine whether C3G protection against ethanol- induced neuronal loss is mediated by the combined action of its antioxidant property and the inhibition of GSK3; (3) determine whether C3G ameliorates ethanol-induced behavioral deficits. C3G is a potent natural antioxidant and has diverse potential benefits for human health. It is a promising neuroprotective agent against ethanol toxicity due to its dual functions as an antioxidant and a GSK3 inhibitor. As a unit, the proposed experiments will elucidate the novel role of C3G, and provide an important basis for future clinical trials to evaluate the feasibility of C3G to treat ethanol neurotoxicity. Our study will potentially offer a new therapeutic strategy. PUBLIC HEALTH RELEVANCE: Alcohol consumption during pregnancy may cause Fetal Alcohol Spectrum Disorders (FASD), among which, fetal alcohol syndrome (FAS) is the most severe form. The developing central nervous system (CNS) is particularly sensitive to ethanol. The loss of CNS neurons may underlie many of the behavioral deficits observed in FASD. Both glycogen synthase kinase 3 (GSK3) and oxidative stress contributes to ethanol neurotoxicity. We have purified a potent antioxidant from blackberries, cyanidin-3-glucoside (C3G) which can also inhibit GSK3 and reduce oxidative stress. The current study is to determine whether C3G ameliorates ethanol-induced neuronal loss as well as behavioral deficits. The proposal will potentially develop a new therapeutic strategy.