Effect of metabolic or respiratory acidosis on rabbit renal medullary proton-ATPase

Distal urinary acidification is thought to be mediated by an H+-ATPase sensitive to N-ethylmaleimide and dicyclohexyl-carbodiimide. We have studied the effect of chronic metabolic acidosis (NH4Cl for 3 days) or respiratory acidosis (inhalation of 10% CO2 for 2 days) on the H+-ATPase of plasma membranes prepared from the medulla. The enzymatic assay for the H+-ATPase was performed in the presence of ouabain and oligomycin and in the absence of Ca. H+-transport activity was assessed by the quenching of acridine orange in the presence of ATP. The 15-25% sucrose gradient fraction was enriched 40-fold in enzymatic activity over the homogenate, and 8-fold in enzymatic activity and 4-fold in H+-transport activity over the fluffy fraction (38,000 X g). Metabolic acidosis (pH less than 7.31) or chronic hypercapnia (PCO2 greater than 66 mmHg; 1 mmHg = 133.3 Pa) was induced for 2-3 days. Both groups showed the same enrichment factor in enzymatic and H+-transport assays as the control rabbits. Enzymatic and H+-transport activities, however, were not different between animals with respiratory acidosis and controls. Kinetic studies failed to disclose an increase in Vmax (673 vs. 702 mumol/(mg protein.min] or a decrease in Km (0.43 vs. 0.48 mM) in chronic hypercapnia as compared with controls. Metabolic acidosis also failed to increase H+-ATPase activity. These data demonstrate that the H+-ATPase of renal medulla does not display the expected increase in activity during acidosis. The role of this H+-ATPase in the adaptation to acidosis remains to be determined.