Functional roles of apical membrane Na+/H+ exchange in rat medullary thick ascending limb

The medullary thick ascending limb (MTAL) of the rat actively absorbs both HCO3- and ammonium. The roles of apical membrane Na+/H+ exchange in these processes and in determining steady-state intracellular pH (pH(i)) were examined in MTAL perfused in vitro with solutions containing 146 mM Na+ and 25 mM HCO3- (pH 7.4). Addition of 1 mM amiloride or 50 mu M ethylisopropylamiloride (EIPA) to the lumen decreased HCO3- absorption (J(HCO3)) from 10.6 +/- 0.5 to 2.3 +/- 0.3 pmol . min(-1). mm(-1) (P < 0.001) and pH(i) from 7.10 +/- 0.02 to 6.86 +/- 0.03 (P < 0.001). The combination of lumen Na+ replacement plus amiloride abolished J(HCO3). Chronic metabolic acidosis (CMA) caused a 32% increase in J(HCO3) that was inhibited by luminal amiloride. Addition of 4 mM NH4Cl to perfusate and bath markedly decreased pH(i) (from 7.10 to 6.70) but did not stimulate luminal H+ secretion as assessed by HCO3- absorption. With 4 mM NH4Cl in perfusate and bath, luminal addition of amiloride decreased pH(i) from 6.70 +/- 0.06 to 6.50 +/- 0.05 (P < 0.005) but had no effect on net ammonium absorption. These results demonstrate that 1) apical membrane Na+/H+ exchange mediates virtually all of HCO3- absorption and is an important determinant of steady-state pH(i) in the MTAL; 2) the adaptive increase in HCO3- absorption in CMA is mediated by an increase in apical membrane Na+/H+ exchange; 3)ammonium markedly acidifies the cells but does not stimulate luminal acidification, suggesting that pH(i) is not a predominant influence on apical Na+/H+ exchange activity and that H+ generated in the cells as the result of transcellular ammonium absorption is extruded across the basolateral membrane; and 4) apical membrane Na+/H+ exchange is not important for ammonium absorption.