Metabolic cost of bafilomycin-sensitive H+ pump in intact dog, rabbit, and hamster proximal tubules.

Bafilomycin A1 is a specific inhibitor of the brush-border membrane-bound H(+)-adenosinetriphosphatase (H(+)-ATPase) of the kidney cortex with no effect on the mitochondrial ATP synthetase or on the basolateral Na(+)-K(+)-ATPase activities. Bafilomycin A1 is thus a useful tool to estimate the contribution of the activity of the H(+)-ATPase to the cellular ATP turnover in a suspension of proximal tubules containing largely S1 and S2 segments. In dog proximal tubules incubated under control conditions, we found that 81% of the respiration is directly related to ATP synthesis, i.e., is sensitive to oligomycin (phosphorylative respiration). Of this amount, 29% is inhibited by 5 x 10(-7) M bafilomycin A1 alone and 90-95% by the combination of bafilomycin plus ouabain. These results indicate that the H(+)-ATPase activity is a significant energy-requiring process in dog proximal tubules. If bafilomycin is added after a 5- to 7-min preincubation with 1 mM ouabain, then the bafilomycin-sensitive ATP turnover is larger, reaching 44% of total phosphorylation. This may suggest that the H+ pump is stimulated by the indirect inhibition of the Na+/H+ exchanger produced by the exposure of tubules to ouabain. The contribution of the bafilomycin-sensitive H+ pump to the cell ATP turnover is also increased by acidification of the extracellular medium. In rabbit and hamster proximal tubules, the bafilomycin-sensitive ATP requirement involves only 5 and 10%, respectively, of the total ATP turnover. These results demonstrate that the metabolic cost of proton secretion by the membrane-bound H(+)-ATPase in suspensions of proximal tubules may be considerable but varies significantly from species to species.