This proposal is aimed at determining whether glial cells in lamprey spinal cord revert to an earlier developmental type in response to injury. In most vertebrates, the main type of glial cell, the astrocyte, is filled with bundles of filaments composed of a protein called glial fibrillary acidic protein (GFAP). However, in animals, such as the sea lamprey, that show regeneration in the central nervous system, the glial filaments are composed of keratins, similar to the protein that makes skin tough, and are interconnected by a network of "spot weld" mechanical contacts called "desmosomes". Our laboratory is determining the reasons for this correlation. In the lamprey, three different keratin subunits combine in different proportions to form filaments in glial cells of brain and spinal cord. Near a spinal cord transection, the glial filaments become like those found in the brain. Since the brain develops later than the spinal cord, this may represent a developmentally more primitive glial cell. The goal of the present proposal is to determine whether this change in keratin composition truly represents a reversion to an earlier embryonic form and whether this comes about by a change in the relative rates of synthesis of the three keratin subunits. We will clone and sequence the gene for each of the three subunits and thus generate molecular probes with which to determine how actively each subunit is made. In situ hybridization will be used on brains and spinal cords from animals of different ages, in order to determine how the mix of keratin subunits expressed by glial cells changes during development. Then we will compare this pattern of change with the pattern seen in glial cells at the site of a spinal cord cut. If there is evidence from the composition of keratin filaments for a reversion of glial cells to an earlier developmental form, future work in the laboratory will be targeted toward discovering the molecules on the surface of the Glial cells that may be responsible for guiding nerve fibers during their regeneration and presumably also their early development.
