The primary objective of this proposal will be to examine the interaction of hormones in regulating NaC1 absorption by the rat cortical collecting tubule. The basis for the proposed study lies in the recently demonstrated synergism between the actions of ADH and mineralocorticoids in regulating NaC1 and KC1 transport in this nephron segment. ADH results in a rapid and sustained increase net sodium and chloride absorption due to enhancement of amiloride-sensitive Na+ entry across the luminal membrane. This effect is potentiated by prior treatment with mineralocorticoids, i.e., there is a greater than additive synergism between the two hormones. This result suggests a basis for the requirement for ADH in the development of hypertension in the DOC-salt model in which the two hormones might interact to cause salt retention. Recent evidence also indicates that bradykinin decrease NaC1 absorption in the rat by an effect on an electrogenic NaC1 cotransport mechanism. Additional information indicates that alpha2 adrenergic agents decrease the cAMP response to ADH. It seems possible that these two agents may be involved in modulating the response to ADH and mineralocorticoids, especially in phenomena such as mineralocorticoid escape. The methodology to be used in this study revolves primarily around the isolated perfused tubule technique as applied to the rat cortical collecting tubule. The primary parameters measured will be the net transport of Na+, K+ and Cl- using electron probe liquid droplet ultramicroanalysis, as well as the recording of transepithelial voltage. Electrophysiological techniques will be used to measure changes in luminal membrane conductance, and to quantify the amiloride-sensitive pathway. We expect that during the course of these experiments, we will also be able to develop fluorescence-labelled monoclonal antibodies to the amiloride-sensitive Na+ channel in the apical membrane of the rat cortical collecting tubule. This would allow us to explore the cell biology of the regulation of this channel.
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