Objective Ethanol has long been demonstrated to trigger cell apoptosis in the central nervous system.The over-production of reactive oxygen species (ROS) is considered as one of the most important mechanisms involving in the apoptosis caused by ethanol.Ginkgolide B(GB),which was widely used as a monomer of traditional Chinese medicine, was reported to scavenge free radicals in endothelial cells and smooth muscle cells.However, whether GB can prevent ethanol-induced neuro-cytotoxicity is still unknown.The aim of this study was to investigate effects of GB on ethanol-induced cytotoxicity, oxidative stress and apoptosis, and to explore potential protective molecular mechanism of GB.Methods Effects of GB on ethanol-induced cytotoxicity were detected by MTT and LDH assays.Cell death and apoptosis were detected by flow cytometry.Caspase-3 activity and ROS were determined by fluorophotometer.Lipid peroxidation was also detected.Expression of NOX2 and p47phox was detected by real-time reverse transcription polymerase chain reaction and Western blot analysis.NADPH oxidase activity was measured by SOD-inhibitable cytochrome c reduction.Results (1) When PC12 cells were pretreated with different concentration of GB, cell viability was significantly increased and LDH leakage was decreased in a dose-dependent manner (P < 0.05, as compared with control).(2) A significant increase in caspase-3 activity was found in PC12 cells incubated with ethanol for 24 h, and pretreatment of GB decreased caspase-3 activity (P < 0.05).(3) MDA level in GB-pretreated cells was significantly lower than that treated with ethanol alone (P < 0.05).(4) By RT-PCR and Western blot analysis, we found that ethanol upregulated expression of ROS-generating NOX family members including NOX2 and p47phox.Moreover, GB attenuated NADPH activities induced by ethanol.(5) Ethanol-induced production of ROS was significantly decreased in cells treated with apocynin.Conclusion Ginkgolide B attenuates ethanol-induced neurotoxicology through regulating NADPH oxidases.