Development of the brain procedes by a complex and regulated series of morphogenetic steps, disturbance of which results in injury ranging from retardation to gross malformations. The long-term goal of these studies is to better understand the mechanisms by which toxic compounds interfere with brain development. To study these processes, advantage will be taken of the well-recognized developmental neurotoxicity of methylmercury (MeHg). MeHg arrests neuronal migration and inhibits maturation of post-migratory neurons. The hypotheses to be tested are: 1) MeHg damages the microtubular cytoskeleton of neurons and glia, thereby disturbing neural-glial interactions required for neuronal migration and maturation, and 2) MeHg independently perturbs the expression and/or function of cell adhesion molecules (CAMs) essential for cell migration, aggregation, and differentiation. These hypotheses will be tested using an embryonal carcinoma (EC) cell culture model of differentiating neuroectoderm, murine cerebellar slices in culture, and mice treated with MeHg in vivo. The relative sensitivities to MeHg of microtubules undergoing post-transnational modifications will be studied using indirect immunofluorescence microscopy. Data from the culture system will be compared with MeHg-induced microtubule injury in murine cerebellar explants and in cerebella of postnatally-exposed mice. The relationships between microtubule disassembly, neuronal migration, and neurite formation will be established. In parallel studies, the effects of MeHg on the appearance and function of major CAMs will be assessed in differentiating EC cells, cerebellar explants, and in vivo. The dose-dependent effects of MeHg on cell migration in the EC and cerebellar explant systems will be assessed by time-lapse photomicroscopy, and correlated with microtubule and CAM staining. CAMs will be identified in cultures and in cerebellum sections by immunofluorescence microscopy and by SDS-PAGE. These studies will: 1) determine the relative sensitivities of different classes of interphase microtubules in neurons and astroglia to MeHg-induced disassembly, 2) correlate the pattern of microtubule damage and reversibility with effects on neuron migration and maturation, 3) determine whether function of neuronal and/or glial CAMs is impaired by MeHg, and 4) determine whether CAM involvement contributes to MeHg-induced brain injury.
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