The long-term goal of the proposed research is to understand the nature of the synaptic interactions that take place within the inner plexiform layer of the mammalian retina, and how these interactions contribute to the receptive field properties of ganglion cells. In this research proposal, neurotransmission between cholinergic (CH) amacrine cells and ganglion cells in the rabbit retina will be examined. The overall strategy is to stimulate CH amacrine cells either chemically (with local microinjecton of K+) or electrically (with a metal microelectrode) while monitoring the responses from ganglion cells.
The specific aims are: (1) to determine the magnitude and spatial extent of CH input to ganglion cells, (2) to determine if GABA is co-released with acetylcholine upon stimulation of CH cells, (3) to examine how the responses of ganglion cells to CH cell stimulation are influenced by light stimuli, and (4) to investigate possible interactions between CH cells. I have developed a technique by which CH amacrine cells and ganglion cells, labeled with fluorescent dyes, can be visualized under an epifluorescence microscope in the living, superfused rabbit retina. Under the microscope, a recording microelectrode will be positioned near a selected ganglion cell body and the stimulating probe positioned near an CH cell body. All manipulations will be done using a low-light-level video system to minimize photobleaching. Four experiments will be conducted, each addressing a specific aim. In the first experiment, ganglion cells will be recorded extracellularly and the effects of stimulation of neighboring CH cells will be evaluated. In the second experiment, intracellular recordings will be made from ganglion cells to examine possible release of GABA from CH cells. In this experiment, the effects of acetylcholine (that is co-released) will be blocked pharmacologically to unmask and GABA effect. In the third experiment, ganglion cells will be recorded extracellularly and light stimuli presented just moments before CH cell stimulation. Of particular interest is the possibility that light-induced inhibitory mechanisms may modulate the release of acetylcholine or postsynaptic effects thereof in directionally selective ganglion cells. In the fourth experiment (addresses the last specific aim), the effects of CH cell stimulation on the response of a ganglion cell to stimulation of second CH cell will be analyzed.
