Much of what we know about the neural retina is constrained by our available methods. For example, our anatomical catalogues of retinal cell types, the distribution within them of the neurotransmitter glutamate, and their expression of the glutamate receptor subunits are based entirely on images of dead tissue prepared for histology. Our physiology/pharmacology of the actions of glutamate in the retina is limited by the difficulty of reliably accessing particular cell types and is complicated by the action of receptor antagonists at more than one synapse. The idea of this proposal is that it should be possible to express flourescent proteins in transgenic mice to label defined cell types for studies of living retina, to follow and measure glutamate receptor subunits in retinal cells, and to determine the consequences of expressing mutant glutamate receptor subunits that act as dominant- negative elements.
Specific aim one is to determine whether it is possible to express and detect a reporter enzyme fused to the jellyfish Green Fluorescent Protein (GFP) in the living retinae of transgenic mice.
Specific aim two is to use GFP-tagged glutamate receptor subunits to ask: can we localize, measure, and follow fluorescent receptor subunits in retinal neurons.
Specific aim three will test the idea that mutant receptor subunits, which act as dominant-negative elements, can be used to eliminate defined sets of glutamate receptors from a retinal cell type. The significance of the proposed work is that it would, if successful, provide a new means of studying the dynamic properties of retinal neurons as well as testing hypothesis of synaptic circuitry and function.
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