The overall objective of this proposal is to establish meaningful structural-functional relationships in the mammalian nephron. Three major lines of inquiry will be pursued, two of which represent natural progressions of ongoing research that will be expanded and intensified through the development of new methodologies and new collaborative relationships. The three areas of inquiry include examination of heterophagy and autophagy in the renal tubule, delineation of the sites and mechanisms of proton and bicarbonate transport in the collecting duct, and continued morphological exploration of the kidney. More specifically: 1. The processes of autophagy and heterophagy will be evaluated in the mammalian proximal tubule to better characterize the manner in which subcellular organelles, proteins and related molecular substances undergo degradation. Special emphasis will be placed on the measurement of cathepsin B mRNA expression as a means of obtaining a better understanding of the mechanism that controls cathepsin B levels in the autophagic vacuole-lysosomal compartment. 2. The sites and subcellular events involved in proton and bicarbonate transport will be evaluated along the mammalian collecting duct by examining the structure and function of the principal cells and the intercalated cells. Heavy reliance will be placed on identification of transport proteins and ion channels through the use of highly specific antibody probes and immunocytochemical and photoaffinity labelling techniques. 3. The distribution of the sympathetic nerve fibers in the mammalian kidney and the relationship between neuroeffector junctions and alpha- and beta-adrenoceptors on tubules and vasculature will be examined. Special emphasis will be placed on the identification of alpha- and beta-adrenoceptors throughout the renal parenchyma at the ultrastructural level with the use of specific antibodies directed against the receptors and by autoradiographic localization of radiolabelled adrenergic antagonists using photoaffinity labelling. These investigations will require the application of a variety of physiologic, biochemical and morphologic techniques including light microscopy, transmission and scanning electron microscopy, light and electron microscopic autoradiography, in vitro microperfusion of isolated kidney tubules, ultramicroenzyme analysis and immunocytochemistry.
Showing the most recent 10 out of 20 publications
