Retinal Ganglion Cell Degeneration and Regeneration in Isl3-Gfp Transgenic Xenopus Laevis

Anthony Diaz-Santana1, Nicholas Marsh-Armstrong2, Ileana Soto-Reyes2, and Shih-Jung Pan2. (1) Chemical Engineering, University of Puerto Rico, Mayaguez, PR 00681, (2) Neuroscience, Johns Hopkins University, School of Medicine, Baltimore, MD 21205

Glaucoma is the leading neurodegenerative cause of blindness and the second leading cause of blindness worldwide. Elevated pressure in the eye is a risk factor, but even people with normal pressure can lose vision due to glaucoma. Glaucoma affects retinal ganglion cells. Chemicals that promote retinal ganglion cell regeneration may hold promise in the treatment of glaucoma. Using transgenic Xenopus laevis in which retinal ganglion cells are labeled by a transgene, I have developed and tested a novel in vivo assay for the testing of such chemicals. Male Isl3:GFP transgenic Xenopus laevis and female Wild Type (WT) Xenopus laevis' progeny were grown and screened for high GFP expression and albinism. Axotomy was performed using two glass needles, of 50 and 100 ?m outer diameter, beveled to a 20 angle, with the aid of micro manipulators. Injections of 2.6nL of brain derived neurotrophic factor (BDNF), Forskolin, and BDNF with Forskolin were made using glass needles as well. Imaging of live animals at injury site and optic tectum was performed at specific time points of the intervention (including axotomy and injection). Innervation values were obtained by measuring pixel intensities in selected regions of interest using custom scripts in IPLab Spectrum 4.0 software. Results show that BDNF may promote retinal ganglion cell regeneration in Xenopus laevis. Provided that these results reproduce in a larger population sample, BDNF will be used as a positive control in future drug screening studies. Isl3-GFP Transgenic Xenopus laevis shows promise as an in vivo assay to screen drugs that promote retinal ganglion cell axonal regeneration.