FETAL ALCOHOL EXPOSURE, NMDA RECEPTORS AND NEUROSTEROIDS

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The brain is severely affected by prenatal alcohol exposure resulting in neurobehavioral deficits that have devastating effects later in life. Therefore, it is important to develop therapeutic measures that minimize these noxious effects of fetal alcohol exposure. Understanding the cellular and molecular mechanism of the prenatal effects of ethanol is a prerequisite to developing such treatments. Among the neuronal proteins affected by in utero ethanol exposure are the NMDA receptors (NMDA- Rs), which play important roles in neuronal development, plasticity and death. A number of studies indicate that prenatal ethanol exposure affects NMDA-Rs; however, the mechanism of this effect of ethanol has not yet been elucidated. Our hypothesis is that prenatal ethanol exposure alters the actions of neurosteroids, which are endogenous modulators of NMDA-Rs. The research design includes exposure of rats during pregnancy to a diet that results in blood alcohol concentrations near the legal intoxication limit in humans (approximately 0.08 g/dl). Aim #1 is to measure the effects of fetal ethanol exposure on the regulation of NMDA-Rs by different classes of neurosteroids. We will use electrophysiological techniques to study the actions of neurosteroids that have been shown to either potentiate or inhibit NMDA-R function. These studies will be performed in cultured hippocampal neurons from neonatal rats as well as hippocampal slices from adult rats. Aim #2 is to determine the mechanism of the prenatal ethanol-induced alterations o the modulatory actions of neurosteroids on NMDA-Rs. Subunit composition and protein phosphorylation are factors that could regulate the sensitivity of NMDA-Rs to neurosteroids. We will use Western blot, radioimmunohistochemistry and single-cell RT-PCR techniques to assess subunit composition. We will test the effect of inhibitors and/or activators o protein kinases and/or phosphatases on neurosteroid sensitivity of NMDA-Rs in cultured neurons and hippocampal slices. Aim #3 is to determine the effect of fetal alcohol exposure on the hippocampal levels of neurosteroids that regulate NMDA-R function. We will use radioimmunoassays to determine levels in hippocampi from embryos, and neonates as well as adult rats. Importantly, studies have shown that prenatal stress produced by fetal alcohol exposure results in a reduction in the responsiveness to the neurobehavioral effects of neurosteroids later in life. The experiments that we propose to do could increase our understanding of the mechanism of this ethanol-induced decrease in sensitivity to neurosteroids. Moreover, these experiments will increase our knowledge of the roles played by hippocampal NMDA-Rs in the pathogenesis of alcohol-related birth defects.