This research is aimed at understanding the basis for the conductance of the crystalline lens, how this conductance is controlled, and how it is altered by the process of cataractogenesis. The long-term aim is to develop ways to therapeutically control conductance so as to slow the progress of cataract formation. In this proposal, the distribution of lens gap junctions will be mapped by intracellular dye injections through microelectrodes followed by an image analyzer based quantitation of the dye spread at many lens locations. Direct measurements of the conductance, voltage dependence, Ca dependence, pH dependence, and the calmodulin dependence of the gap junctions connecting the two cells are planned using a 2-cell voltage clamp technique. Epithelial- epithelial, epithelial-fiber, and fiber-fiber junctions will be characterized. These results will be verified with measurements of the diffusion of fluorescent substances between the cells. The electrical and dye coupling as well as transepithelial cleft resistance from capsule-epithelium monolayers will be measured. Spatial differences in the diffusion in the extracellular space near the lens surface will be determined. The properties and distribution of ionic channels in lens membranes will be characterized using patch clamp techniques and noise analysis. Blockers for the channels will be found. All of these methodologies will be applied to normal rat lenses and to rat lenses early in formation of galactose cataracts to identify conductance mechanisms involved in the cataractogenesis. We will attempt to alter the cataract process through the use of channel blocking drugs.
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