Heavy prenatal alcohol exposure can cause fetal alcohol syndrome (FAS) characterized by growthdeficits, facial dysmorphology, brain damage, and neurobehavioral dysfunction. These abnormalities varywidely across a range now considered to reflect a fetal alcohol spectrum disorder (FASD). Variation in FASDis linked to individual differences in patterns of drinking during pregnancy together with genetic and biologicalfactors influencing vulnerability. Little is understood about the molecular mechanisms of FASD pathogenesis,or about genetic differences that contribute to the diverse FASD presentations. Our long-term objectives areto identify alcohol effects on gene expression that can link mechanistically to abnormal brain development,and to identify genetic factors that influence susceptibility to those pathogenesis mechanisms. We have usedembryonic cultures to expose C56BL/6 (B6) and DBA/2 (D2) inbred mice to alcohol during early neurulation.The B6 and D2 embryos showed different patterns of developmental delays and structural defects after 44hours of exposure to high alcohol concentrations (300-400 mg/dl). In the B6 embryos, DMA microarrayexpression profiling, followed by hypothesis-driven bioinformatics analysis of differentially expressed genes,revealed over-representation of genes in gene sets regulating specific functions. Prominent down regulationof many genes critical for neural specification, neural typing, and neural patterning was evident, andconfirmed for several. Our goal now is to generate more detailed temporal and spatial analyses of thedisrupted expression of gene cohorts and their regulators, to identify correlations with dysmorphology anddifferential outcomes in the two inbred strains.
Aim 1 will identify gene expression changes in B6 and D2embryos after either 4 or 20 hrs of alcohol exposure, using DNA microarray and hypothesis-drivenbioinformatics analysis to identify effects in gene sets in specific functional pathways. Patterns ofdysmorphology will be compared with localization of changes seen with in situ hybridization of selectedgenes.
Aim 2 will assess the relevance of observed down regulation of selected mouse genes to vertebrateneural development, by using novel zebrafish morpholino knockdown technology to carry out rapid screeningof loss-of-function effects of orthologous genes.
Aim 3 will use informatics to determine whether alcoholeffects may be due to actions on functions of specific transcription factors (TF), by identifying TFs predictedfrom TF binding sites common to alcohol-affected gene cohorts in the mouse models. These studies willprovide insights into prenatal alcohol effects on molecular regulators of embryonic brain development, alongwith genetic factors that influence the type or extent of damage associated with FAS. This knowledge willadd important information for the identification, intervention, or treatment in pregnancies at-risk for FASD.
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