The primary goal is to advance understanding of the functional and structural relationships in the mammalian renal inner medulla by determining the three-dimensional structural organization of the descending and ascending thin limbs of Henle' s loops, collecting ducts, and vasa recta in the inner medulla and the relationship of this structure to integrated function, including the fluxes of urea, inorganic ions, and water, and, thus, development of the inner medullary osmotic gradient. Observations that: 1) many inner medullary thin limbs contain segments of cells with the appearance and function of cells in pure ascending limbs intermixed with segments of cells with the appearance and function of cells in pure descending limbs; 2) the lower 60% of all pure descending thin limbs with bends below the first millimeter of the inner medulla appear to lack water and chloride channels; 3) the inner medullary long descending thin limbs tend to be arranged in a ring-like arrangement around collecting ducts; and 4) the prebend segments are all of similar length and begin abruptly with the appearance of chloride channels suggest that the architectural arrangement of these segments and their transporters and channels may be critical to the development of inner medullary ionic and osmotic gradients. An initial mathematical model supports the latter concept. The planned studies will involve primarily Munich-Wistar rats and take advantage of major developments in techniques for three-dimensional functional reconstruction of tubular structures in the inner medulla and of techniques for studying functions of segments of thin limbs by perfusion in vitro. They will involve 1) complete three-dimensional, quantitative reconstruction of all inner medullary thin limbs of Henle's loops, collecting ducts, and vasa recta from serial sections, using immunocytochemical markers of physiological function to identify tubule segments and tubular sites of transport functions; 2) direct measurements, by in vitro microperfusions, of the urea, NaC1, and water permeabilities of specific thin limb segments defined in the reconstructions; and 3) integration of the function of the inner medullary thin limbs, vasa recta, and collecting ducts in the process of producing a concentrated urine via mathematical modeling based on information obtained from the functional reconstructions and the in vitro microperfusions. ? ?
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