Ethanol-induced neuro-developmental abnormalities are associated with impaired insulin and IGF signaling, and increased oxidative stress in CNS neurons. We examined the roles of ethanol and its principal toxic metabolite, acetaldehyde, as mediators of impaired insulin/IGF signaling and oxidative injury in immature cerebellar neurons. Cultures were exposed to 3.5？mM acetaldehyde or 50？mM ethanol ± 4-methylpyrazole (4-MP), an inhibitor of ethanol metabolism, and viability, mitochondrial function, oxidative stress, DNA damage, and insulin responsiveness were measured 48 hours later. Acetaldehyde or ethanol increased neuronal death and levels of 8-OHdG and 4-HNE, and reduced mitochondrial function. Ethanol inhibited insulin responsiveness, whereas acetaldehyde did not. 4-MP abated ethanol-induced oxidative stress and mitochondrial dysfunction, but failed to restore insulin responsiveness. Furthermore, alcohol and aldehyde metabolizing enzyme genes were inhibited by prenatal ethanol exposure; this effect was mediated by acetaldehyde and not ethanol + 4MP. These findings suggest that brain insulin resistance in prenatal alcohol exposure is caused by direct effects of ethanol, whereas oxidative stress induced neuronal injury is likely mediated by ethanol and its toxic metabolites. Moreover, the adverse effects of prenatal ethanol exposure on brain development may be exacerbated by down-regulation of genes needed for metabolism and detoxification of alcohol in the brain. 1. Introduction Prenatal alcohol exposure causes fetal alcohol spectrum disorder (FASD), which is associated with multiple and varied developmental abnormalities in the brain and results in sustained deficits in cognitive and motor functions [1–6]. Ethanol exerts its neurotoxic and teratogenic effects [7, 8] by promoting oxidative stress and impairing insulin and insulin-like growth factor (IGF) signaling in the developing brain [9, 10]. Whether these effects are mediated by direct toxic effects of ethanol or its principal metabolite, acetaldehyde, has not yet been determined. Ethanol has broad inhibitory effects on insulin and IGF signaling in the developing brain and immature neurons. For example, ethanol impairs ligand-receptor binding, tyrosine phosphorylation and activation of receptor tyrosine kinases, transmission of signals through insulin receptor substrate (IRS) proteins, and downstream activation of phosphatidylinositol-3 kinase (PI3 kinase)-Akt, p21ras, and mitogen-activated protein kinase kinase (MAPKK) [11]. Consequences include reduced neuronal proliferation, survival, migration,
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