MOLECULAR DEVELOPMENT OF CENTRAL RETINAL PATHWAYS

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The long term objective of my research is to understand at a molecular level the mechanisms governing the formation of precise neuronal connections in the mammalian visual system. The experiments proposed here pursue this goal by investigating how the functionally important retinal ganglion cell axon pathways at the optic chiasm are formed during embryonic development. An understanding of the cell surface molecules involved in retinal axon guidance will enable us to devise strategies to control and direct the growth of retinal axons, and is thus fundamental to efforts attempting to re-establish visual connections following injury and disease. Furthermore, since cell surface interactions are not only important for retinal axon guidance, but very likely also govern the patterns of connectivity among other retinal neurons, an understanding of how cell surface molecules direct these developmental events is central to strategies using tissue transplantation to restore visual neuronal connections. The studies proposed here form an integrated set of experiments in which we use both in vitro and in vivo approaches to analyze the function of CD44 and L1, two cell surface molecules prominently expressed on embryonic retinal axons and neurons at the future region of the optic chiasm. We test the hypotheses that L1 and CD44 acting together, mediate interactions between ingrowing retinal axons and embryonic chiasm neurons and govern the formation of the "X" shaped retinal pathway intersection known as the optic chiasm. Secondly, we also examine whether CD44, which is prominently expressed at the midline, is involved, either directly or in conjunction with other cell surface molecules, in the specific routing of retinal axons into the ipsilateral and contralateral optic tracts. This hypothesis is based on the fact that CD44 regulates cell surface ligand-receptor interactions and is uniquely placed at the midline to influence the decision of axons whether to cross the midline into the contralateral optic tract or avoid crossing and project ipsilaterally. Experimentally, the molecular basis of CD44 and L1 action and their role in retinal pathway development in vivo is studied using three approaches. One, is direct perturbation of CD44 and L1 function in vivo by injections of blocking reagents into living mouse embryos and examination of the effects on retinal pathway formation. Second, is interference of CD44 and L1 function in embryonic visual system preparations and analysis of effects on growth cone behavior using time-lapse videomicroscopy. Third, is the use of cell lines expressing truncated forms of CD44 and L1 to define the functional domains mediating their actions on retinal axon growth. These studies address major gaps in our understanding of visual pathway development by testing defined surface molecules for functional importance in vivo and by dissecting the mechanisms to learn the molecular basis for central visual pathways development.