The long-term goal of this program is to understand visual processing in the mammalian inner retina by defining its transmitter systems, cellular morphology and circuitry relationships. A major focus of these studies is on the inhibitory peptide, somatostatin (SRIF) and its G(i/o)-protein coupled receptors (sst1-sst5) as a model system for peptide actions in the retina. Past studies have established that SRIF immunoreactivity is localized to sparsely occurring, wide-field amacrine cells and there is a differential expression of sst receptors to multiple retinal cell populations. Preliminary studies show light-evoked stimulation of SRIF mRNA expression, and an unexpected complexity in the organization of sst receptors with discrete bipolar and ganglion cell types expressing different sst receptors. Other studies show SRIF inhibition of K+-depolarization-evoked [Ca 2+] increase of photoreceptor, bipolar and ganglion cells. These findings suggest that SRIF has a paracrine mode of action on multiple cell populations and, therefore, a broad modulatory influence on the processing of visual information. Proposed studies will test the hypotheses that SRIF levels in the retina are increased by light, and that SRIF exerts its effects at both the cellular and circuitry levels by acting at pre- and postsynaptic sites via distinct sst receptors expressed by multiple cell populations.
Specific Aim 1 will evaluate A) diurnal, circadian, and light-evoked influences on SRIF synthesis and content, and B) SRIF immunoreactive amacrine cell organization and circuitry relationships by establishing its synaptic inputs using real-time RT-PCR, radioimmunoassay and immunohistochemistry with confocal and pre-embedding electron microscopy.
Specific Aim 2 will evaluate the neuronal targets of SRIF by A) characterizing the cellular organization of sst1 and sst4 immunoreactive ganglion cells, and B) determining their bipolar and amacrine cell synaptic inputs, and central projections using immunohistochemistry and intracellular labeling with confocal and pre-embedding electron microscopy.
Specific Aim 3 will A) evaluate the action of SRIF, and sst agonists and antagonists, and B) characterize the intracellular transduction pathways that mediate SRIF's inhibition of a depolarization-induced [Ca2+]i increase in sst1 and sst4 immunoreactive ganglion cells using acutely dissociated and cultured ganglion cells with fluorometric [Ca2+] imaging techniques. Experimental studies will use rats, and wild-type and a CFP-fluorescently labeled ganglion cell transgenic mouse to identify ganglion cells. The proposed studies will provide new information about the role of peptides in the modulation of retinal cells and circuitry in the inner retina, thus providing the basis for a better understanding of visual image processing in the retina. These objectives are consistent with the health-related goals of the National Eye Institute to develop more effective treatments and ultimately to prevent retinal diseases, such as diabetic retinopathy and macular degeneration.
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