The axons of retinal ganglion cells project to the central visual nuclei in the brain in a topographic pattern, such that the two dimensional pattern of the ganglion cells in the retina is approximately recreated in the pattern of their terminals in the visual nuclei. Alteration of this pattern results in the ineffective processing of visual information and an inability to respond to visual stimuli in a meaningful manner. The mechanism responsible for development of the topographic pattern of connections is unknown. One hypothesis suggests that axons carry specific molecules that act as positional labels. The interaction between positionally labeled retinal axons and tectal cells with complimentary labels could determine the pattern in which connections develop. The goal of this project is to identify differences between axons growing from the nasal and temporal sides of the developing chick retina that might reflect positional labels. The initial steps of the project will concentrate on characterization of anatomical and molecular differences between the two groups of axons. The long term goal is to understand functional differences that account for development of the specific pattern of axon growth and synapse formation exhibited by retinal axons. The project is divided into four specific aims. First, TRAP, a molecule expressed on most temporal but few nasal retinal axons, will be characterized in more detail. This will involve cloning the gene for TRAP from a cDNA library, sequencing the gene, and developing antibodies to the protein for use in analyzing the function of TRAP. Second, three complementary approaches will be used to identify other molecules expressed asymmetrically between the nasal and temporal sides of the developing retina. These approaches include the use of monoclonal antibodies, subtractive cDNA hybridization, and lectin blots. As molecules are discovered, they will be characterized as described for TRAP. Third, anatomical differences in the pattern of growth between nasal and temporal retinal axons will be identified. This information will be used to develop in vivo assays to study the function of side specific molecules, and may also supply insights into functional differences between nasal and temporal axons. Fourth, the function of molecules asymmetrically expressed in the developing retina will be examined. This will involve perturbing the function of these molecules with antibodies in vivo and in vitro. Preliminary studies will also evaluate techniques for introducing genes to alter the expression of specific molecules. This project will have a significant impact on the investigators future ability to reconnect retinal axons with the brain in a useful manner in order to cure blindness.
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