ETHANOL AND L1 MEDIATED NEURITE OUTGROWTH

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The ultimate goal of this work is to identify the role of L1, a neural adhesion molecule, in alcohol related neurodevelopmental disorder. Patients with the most severe form of this disorder, fetal alcohol syndrome, possess neuroanatomic features which are strikingly familiar to those with genetic defects in L1. These observations suggest that L1 plays a role in the pathogenesis of alcohol related neurodevelopmental disorder. Our preliminary results show that ethanol inhibits L1 mediated neurite outgrowth at concentrations comparable to social drinking in rat postnatal day 6 cerebellar granule neurons. L1 is a developmentally regulated cell surface glycoprotein which is critical for proper neural migration, axon guidance and axon-fascicle formation through binding to itself or other molecules at the cell surface. Binding of L1 to itself is followed by cascades of signaling events critical for neurite outgrowth. These cascades can be divided into two pathways: A pathway common to several cell adhesion molecules involving activation of the fibroblast growth factor receptor and subsequent release of arachidonic acid, and pathways unique to L1 with phosphorylation of L1 on the cytoplasmic domain. Our hypothesis is that ethanol disrupts central nervous system development by altering those L1 mediated signaling cascades which lead to neurite outgrowth. This hypothesis will be tested in rat cerebellar granule cells and in a rat model of ARND. Using assays of neurite outgrowth, immunoprecipitation and Western blot, the ethanol sensitivity of the common pathway will be tested by determining the effect of ethanol on: 1) neurite outgrowth stimulated by other cell adhesion molecules, 2) levels of phosphotyrosine modified proteins, and 3) phosphorylation of the fibroblast growth factor receptor. For the L1 unique pathways, in vitro kinase assays, metabolic labeling, and immunocytochemistry will be used to determine the effects of ethanol on: 1) serine kinases associated with L1, 2) the serine and tyrosine phosphorylation of LI, 3) the binding of L1 to ankyrin, and 4) the cellular distribution of L1. In vitro experiments will be correlated to in vivo experiments. These experiments will provide important information on the underlying mechanism of ethanol's inhibitory effect on L1 mediated neurite outgrowth.