PH REGULATION IN VENTRICULAR CELLS

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The long-term goal of this research is to better understand the mechanisms that regulate intracellular pH (pHi) in adult mammalian heart muscle. In this project we focus on three main areas: 1) peptide modulation of sarcolemmal ion transporters involved in cardiac pHi control, 2) H+ diffusion within cardiac cells and 3) H+ diffusion between cardiac cells, through gap junctions. These topics are closely related since the pHi changes induced by the ion transporters, which are initially confined to the subsarcolemmal space, must eventually reach the cell interior and neighboring cells to exert their full physiologic effect. The peptide studies focus on angiotensin II (AngII), endothelin 1 (ET1) and atrial natriuretic peptide (ANP). These agents exert a wide range of potent effects on cardiac function under both normal and pathological conditions. We will examine peptide effects on HCO3-CI exchange and CI-OH exchange. These acid loading systems are critically important in mediating pHi recovery from intracellular alkalosis and the fall in pHi during metabolic acidosis. The ANP studies focus on identifying its effects on Na-H exchange, Na-HCO3 cotransport, HCO3-CI exchange and CI-OH exchange. The peptide experiments will be performed on ventricular and atrial myocytes, enzymatically isolated from adult rabbits. pHi will be measured in single cells using epifluoresence techniques and the H+ sensitive indicator, SNARF-1. The H+ diffusion studies are designed to characterize H+ diffusion within and between single cardiac cells under a variety of conditions which are likely to create regional pHj gradients. The experiments will be performed on both single myocytes and myocyte pairs (ventricular and Purkinje, rabbit) using confocal microscopy and SNARF-1 to measure pHi. The results of this project will yield new information concerning hormonal modulation of cardiac pHi control systems and the action of intracellular and intercellular H+ diffusion to dissipate regional pHi gradients.