It is the overall objective of the present proposal to investigate the cellular representation of the proximal-tubule acidification response to changes in PaCO2 in the rabbit and the dog utilizing membrane-vesicle methodologies. Our understanding of the cellular mechanisms of basal proximal acidification remains incomplete. Moreover, the mechanisms that mediate the adaptation of proximal acidification to respiratory disorders are largely unknown. Therefore, the specific aims of the present proposal are: 1) To further our knowledge on the multitude of the cellular systems of proximal acidification and to characterize their functional expression under normal acid-base conditions. 2) To explore the cellular mechanisms and the time-course of the adaptation of proximal acidification to acute and chronic changes in PaCO2. Changes in PaCO2 induce prompt, directional changes in proximal-tubule acidification. When the alterations in PaCO2 are sustained, additional, much larger, and slowly-evolving changes in the rate of bicarbonate reabsorption ensue that require several days for completion. Given the slowly-evolving nature of the changes in acidification, it is hypothesized that the renal response reflects graded adaptation and/or sequential recruitment of several cellular systems of acidification. It is further hypothesized that these adaptations occur in a coordinated fashion aiming at the maintenance of the intracellular milieu despite brisk changes in transcellular ionic fluxes. Cellular systems to be investigated include Na+-H+ and pH-dependent C1- transporters in brush-border vesicles (BBMV), H+ ATPase in cortical endosomes and BBMV, Na+-HCO3- cotransport in basolateral vesicles (BLMV), and Na+-dependent glutamine and phosphate uptake in BBMV and BLMV. Characterization of these systems in the basal state will be followed by studies during respiratory disorders. Hypercapnia and hypocapnia will be induced in unanesthetized animals within a large environment chamber. The adaptation of the above systems of proximal acidification will be examined at intervals of 1 hour, 1 day, and 5 days from induction of the respiratory disturbance. Preliminary results in support of widespread adaptive changes in the activity of the cellular systems of proximal acidification following 4 days of hypercapnia or 5 days of hypocapnia have been obtained; notably, BLMV Na+-HCO3- cotransport was not altered after only 1 hour of hypercapnia, nor had adaptation occurred in the BBMV Na+-H+ exchanger after 1 day of hypercapnia. These studies may provide important new insights into the mechanism and regulation of proximal acidification under basal conditions as well as during acute and chronic changes in PaCO2.
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