The overall goal of this Program Project is to understand the mechanisms underlying renal fluid and electrolyte homeostasis. To this end, we will use a broad spectrum of techniques to address a continuum of problems ranging from the cloning and characterization of individual transporters to the contribution of these transporters to integrated renal function at the level of the intact tubule and organ. Our strategy to pursue these themes successfully will include close collaboration on interrelated research projects; sharing of expertise, concepts and techniques by program investigators; and joint use of core facilities. The proposed research projects comprise a broad range of experimental preparations including transport proteins, isolated membrane vesicles, tissue culture cells, Xenopus oocytes, isolated individual kidney cells, single tubules, and whole kidney in vivo. We shall use a wide range of methods including molecular cloning and mutagenesis, functional cDNA expression, generation of transgenic mice, immunocytochemistry, confocal microscopy, fluorometric assays of cell ion activities, whole cell and patch-clamp techniques, in vivo and in vitro perfusion of defined tubule segments, and clearance studies. The introduction into the program of novel approaches and new techniques is also an important feature of the present proposal. A key element in this regard is the adaptation of clearance, micropuncture and microperfusion techniques (proximal and distal tubules) to the study of renal function in transgenic and knockout mice.
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