This is a wide-ranging application from an established retinal neuroscientist. The main focus of the project will be to study the effects of redox reagents on the neurotransmitter receptors of rat retinal ganglion cells. In a past grant period the applicant's laboratory discovered (on rat retinal ganglion cells) that NMDA receptors have a modulation site that responds to oxidizing agents to down regulate receptor activity. During the last project period the applicant's laboratory found that GABA-A and glycine inhibitory receptors on rat retinal ganglion cells also possessed """"""""redox modulatory sites"""""""". Based on preliminary data, the applicant proposes to use patch-clamp recording, two-electrode voltage clamp, digital calcium imaging and molecular biological techniques to extensively characterize these sites. A major goal of the work will be to attempt to determine if modulation of these receptors via the redox site has potential therapeutic value in preventing or treating excitotoxic cell death.
The specific aims of the application are: (1) to characterize the effects of endogenous redox agents, including redox-related forms of nitric oxide, glutathione, and lipoic acid, on the NMDA responses of rat retinal ganglion cells; (2) to characterize the molecular basis of the NMDA receptor's redox modulatory sites(s) using recombinant NMDA receptor subunits expressed in Xenopus oocytes and to compare these responses with those obtained from native NMDA receptors on intact retinal ganglion cells; (3) to characterize a new NMDA receptor subunit that they have cloned from the rat retina using a PCR strategy based upon the structure of the redox modulatory site; (4) to begin to develop strategies to use the neuroprotective effects of the NMDA receptors' redox modulatory site to protect retinal ganglion cells from injury related to ischemia, trauma, or glaucoma; (5) to characterize redox modulatory sites on inhibitory neurotransmitter receptors of rat retinal ganglion cells (GABA-A and glycine receptors); and (6) to characterize the redox modulatory sites of inhibitory receptors further by using recombinant GABA-A and glycine receptors expressed in Xenopus oocytes.
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