Cellular and molecular mechanisms of retinal bipolar cell regeneration in zebrafish

Human retinal degenerative diseases are characterized by slow progressive loss of retinal cells which induces reactive gliosis in Muller glia cells. In mammalian systems, this results in scar tissue formation which exacerbates loss of vision. Similar initial responses are observed following injury in highly regenerative species, such as zebrafish. However, Muller glia cells in these systems are capable of regenerating a functional retina. We are interested in determining how the regenerative potential of Muller glia cells is triggered and controlled. Thus, we are studying the cellular and molecular mechanisms governing how zebrafish regenerate specific retinal cell types. Ultimately, we seek to identify factors that could be harnessed to redirect mammalian Muller glia cells into regenerative pathways. Such insights could aid the development of regenerative therapies for degenerative diseases. Despite the relevance to disease, little is known about how the retina responds to loss of discrete cell types. Here, we focused on characterizing this paradigm to study four principle aspects of the regenerative process: 1) endogenous stem cell activation, 2) stem and progenitor cell proliferation, 3) progenitor cell differentiation, and 4) functional recovery. By studying all four of these aspects in relation to each other we were able to reveal fundamental insights into how retinal regeneration is governed. Specifically in Aim 1, we used transgenic and pharmacological techniques to induce ablation of nyx-expressing retinal bipolar cell subtypes and asked whether the lost cells were subsequently regenerated in zebrafish larvae. We then sought to identify potential stem cell sources. In Aim 2, lineage tracing of retinal stem cell populations was used to ask whether the extent of bipolar cell loss altered the specificity of the regenerative response. In Aim 3, we manipulated the Wnt pathway to investigate the role of Wnt signaling in bipolar cell regeneration. Finally, in Aim 4 we used visual behavior assays to determine if functional deficits attend the loss of nyx-expressing bipolar cells and, if so, whether functional recovery was evident following their regeneration. In particular, our observations demonstrating opposing roles of the Wnt pathway in regeneration have implications regarding the development of age-appropriate and/or cell-specific regenerative therapies.;INDEX WORDS: Regeneration, Retina, Stem Cell, Muller glia, Bipolar, Wnt, beta-catenin.