Strategies for Retinal Regeneration in Vertebrates: from Fish to Human

Abstract

We are studying various aspects of retinal regeneration in zebrafish. We developed a light-induced model for retinal degeneration that results in massive rod and cone cell death in the dorsal and central retina and limited rod cell death in the ventral retina. Within four weeks, the rods and cones regenerated from a population of inner nuclear layer (INL) stem cells in the dorsal and central retina and outer nuclear layer (ONL) rod precursor cells in the ventral retina. A microarray experiment, which examined gene expression at five different key times during light damage and subsequent regeneration, identified several clusters of genes that may play various roles in photoreceptor regeneration. We will use this microarray set to identify the molecules that signal the proliferation response to the INL and ONL cells. To examine the breadth of retinal neurons that can be regenerated, we characterized the effects of intraocular ouabain injection. At low ouabain concentrations, damage was primarily restricted to the INL and ganglion cell layers, while higher ouabain concentrations caused significant cell death in all the nuclear layers. Both types of ouabain-damaged retinas regenerated a nearly wild-type laminated retina, which involved a proliferative cell population residing throughout the retina that may correspond to the stem cells within both the INL and the circumferential germinal zone. Regeneration of the retina also restored the vision-dependent behavioral response. Data will be presented on the degeneration and regeneration responses of the ouabain-treated retina and discussed in relation to the light-damaged retina. The Muller glia, which are neuronal support cells, are a source of regenerated neurons in a variety of organisms and degeneration models. In the light-damaged zebrafish retina, the Muller glia expressed a variety of neuronal precursor genes. These changes in Muller cell gene expression were restricted to the central and dorsal retina, suggesting they are related to either the level of photoreceptor cell damage or the INL cell proliferation. In the ouabain-damaged retina, the Muller glia expressed a different set of genes, which suggests they respond differently to the light and ouabain damage. We will discuss the complexity of the Muller glial cell population and their relative contributions during the regeneration of the light-damaged and ouabain-treated retinas. Finally, to test potential signaling molecules involved in various steps of regeneration (inducing stem cell proliferation, activating Muller glial cells, and identifying the damaged neuronal cell population), we are generating transgenic zebrafish lines that express reporter transgenes at these various steps. Our efforts at isolating these transgenic lines and using them to characterize potential signaling molecules will be discussed.