This research proposal is concerned with a study of synaptic mechanisms in the outer plexiform layer of the vertebrate retina. Pre-and postsynaptic functions will be examined separately by using dissociated retinal neurons. 1) Presynaptic transmitter release by exocytosis will be quantified with the rapid freezing technique in isolated amphibian and mammalian rod photoreceptors. This approach will yield information on the rate of synaptic vesicle release in the dark and during the light response. Synaptic vesicle membrane recycling will be followed by pulse labeling with ultrastructural tracers followed by rapid freezing or conventional fixation. By combining these techniques, it will be possible to identify the organelles which initiate membrane retrieval and which give rise to new generations of synaptic vesicles. 2) Receptors to the putative transmitters acetylcholine and gamma-aminobutyric acid will be visualized by radiolabeled ligands with light and electron microscopic autoradiography. Using both first and second order isolated retinal neurons, it will be possible to localize both pre-and postsynaptic receptors to specific cell types. These experiments on synaptic function will answer general biological questions concerning secretion and cytoplasmic organelle function; they explore how visual information is communicated by photoreceptors to second order neurons; and finally, they test the possible pharmacological interactions in the outer plexiform layer. In the long term, it is hoped that they will provide a basis for the use of cell culture techniques in studies of photoreceptor cell biology and synaptic interactions between pharmacologically identified retinal cells. The continued analysis of the normal function of visual cells is essential in order to understand the alterations which occur in retinal and choroidal disease processes.
