The proposed research will determine the morphology, topographic distribution, transmitter candidates and synaptic relations of three distinct neuronal cell types that have been observed for the first time in an in vitro preparation of the cat's retina. During the previous project period it was demonstrated that after intraocular injections of dopamine and the indoleaminergic transmitter analogue 5,7-dihydroxytryptamine (5,7-DHT), an intense yellow-green fluorescence (resembling formaldehyde-induced histofluorescence) appears in distinct subpopulations of neurons in the inner plexiform layer and the ganglion cell layer of the living retina. Although the chemical basis for the in vitro fluorescence is not understood, its discovery permitted the identification of the fluorescing cells by intracellular injections of horseradish peroxidase (HRP) under direct microscopic control. Three distinct monoamine-accumulating (Ma) cell types were identified: the first an amacrine cell, the second a ganglion cell and the third a cone bipolar cell type. Morphological analysis of the HRP injected cells demonstrated that the processes of the three cell types costratify in a narrow stratum at the outer border of the inner plexiform layer. The proposed research will be divided into five related projects that exploit the in vitro fluorescence to further characterize these cell types. 1) Following intracellular injections of HRP, an analysis of the detailed morphology, stratification, and mosaic organization of the dendritic trees of each type by will be completed. 2) The topographic distribution of each type will be determined by a method that permits systematic sampling form the living retina. 3) Neurotransmitter candidates for the Ma cell types will be investigated by colocalizing immunoreactivity for tyrosine hydroxylase, serotonin or GABA with intracellular injections of the fluorescent dye Lucifer Yellow. 4) The central projections of the Ma ganglion cell type will be examined by colocalizing retrogradely transported fluorescent tracers that have been injected into the superior colliculus or the C-laminae of the dorsal lateral geniculate nucleus, with the in vitro fluorescence. 5) The hypothesis that the three Ma cell types are functionally linked will be tested; potential synaptic contacts between HRP-injected Ma cells identified by light microscopy will be subsequently analyzed by electron microscopy. Taken together, the first three projects will provide a detailed, light microscopic characterization of each of the cell types. The last two projects will test the hypothesis that they are in direct synaptic contact, and comprise the major intraretinal elements in a newly identified visual pathway that projects to the superior colliculus and/or the C-laminae of the dorsal lateral geniculate nucleus.
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