ATP-SENSITIVE NA+-H+ ANTIPORT IN TYPE-II ALVEOLAR EPITHELIAL-CELLS

Type II alveolar epithelial cells in suspension have been previously shown to possess a Na+-H+ antiporter that modulates recovery from an intracellular acid load in the nominal absence of HCO3- [E. Nord, S. Brown, and E. Crandall. Am. J. Physiol. 252 (Cell Physiol. 21): C490-C498, 1987]. Such a Na+-dependent mechanism has also been demonstrated in cultured type II cell monolayers (K. Sano et al. Biochim. Biophys. Acta 939: 449-458, 1988). It has recently been suggested that cultured type II cells possess a H+-ATPase that contributes to recovery from an intracellular acid load [R. Lubman, S. Danto, and E. Crandall. Am. J. Physiol. 257 (Lung Cell. Mol. Physiol. 1): L438-L445, 1989]. The present study was undertaken to investigate and characterize the mechanisms by which cultured type II cells recover from an intracellular acid load in the nominal absence of HCO3-. Cultured type II cell monolayers were loaded with the pH-sensitive probe 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein, and the characteristics of recovery from an imposed intracellular acid load were studied. Recovery of intracellular pH (pH(i)) was found to be strictly Na+-dependent and inhibited greater-than-or-equal-to 95% by 1 mM amiloride. Initial rate of recovery was highly sensitive to pH(i), with recovery rates varying inversely with increasing pH(i). An acidic extracellular pH (6.5) abolished pH(i) recovery. Treatment of type II cells with either the sulfhydryl reagent N-ethylmaleimide, a nonspecific sulfhydryl reagent, or 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole, a specific vacuolar H+-ATPase inhibitor at the concentration tested, resulted in marginal but not statistically significant decrements in pH(i) recovery. Intracellular ATP depletion, using KCN or replacement of glucose by a nonmetabolizable glucose analogue, reduced pH(i) recovery by 70-75% relative to control values. Sensitivity to ATP was apparent even under conditions that preserved the transmembrane Na+ gradient. Taken together, these data are most consistent with a single mechanism for pH(i) recovery in the absence of HCO3-. We interpret this mechanism to be an ATP-sensitive Na+-H+ antiporter that acts to reestablish pH(i) in type II alveolar epithelial cells.