Na+-H+ exchange activity in rat hepatocytes: role in regulation of intracellular pH

Abstract

Amiloride-sensitive Na+-H+ exchange has been identified in basolateral membrane vesicles from rat liver, but little is currently known about its regulation or its role in maintenance of resting intracellular pH (pHi) in intact hepatocytes. We have assessed Na+-H+ exchange activity in isolated or cultured rat hepatocytes in nominally HCO3- free solution under basal conditions and after intracellular acidification by an NH4Cl pulse by measuring 1) pHi, using the pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxy fluorescein, 2) net H+ efflux by pH-stat titration, and 3) amiloride-inhibitable 22Na uptake. Under resting conditions, Na+-H+ exchange did not contribute measurably to Na+ uptake and accounted for less than 20% of net H+ efflux. Hepatocyte pHi averaged 7.07 +/- 0.03, significantly above H+ electrochemical equilibrium (6.92 +/- 0.08) determined using an electrogenic proton ionophore. Transient removal of extracellular Na+ or exposure to amiloride reversibly lowered pHi by 0.09 +/- 0.01 and 0.12 +/- 0.03 pH units, respectively, within 5-10 min. After intracellular acidification by an NH4Cl pulse, Na+ uptake rate increased about twofold, the increase being entirely amiloride inhibitable. Net H+ efflux increased about threefold, and 70% of the increase was amiloride inhibitable. Recovery of pHi after an NH4Cl pulse was reversibly blocked by exposure to amiloride or removal of Na+. Na+-H+ exchange activity (calculated from the rate of change in pHi and intracellular buffering capacity) was inversely related to pHi and was estimated to approach zero at pHi 7.25-7.50.(ABSTRACT TRUNCATED AT 250 WORDS)