METABOLIC COST OF BAFILOMYCIN-SENSITIVE H+ PUMP IN INTACT DOG, RABBIT, AND HAMSTER PROXIMAL TUBULES

Bafilomycin A1 is a specific inhibitor ot the brush-border membrane-bo und H+-adenosinetriphosphatase (H+-ATPase) of the kidney cortex with n o effect on the mitochondrial ATP synthetase or on the basolateral Na-K+-ATPase activities. Bafilomycin A1 is thus a useful tool to estimat e the contribution of the activity of the H+-ATPase to the cellular AT P turnover in a suspension of proximal tubules containing largely S1 a nd S2 segments. In dog proximal tubules incubated under control condit ions, we found that 81% of the respiration is directly related to ATP synthesis, i.e., is sensitive to oligomycin (phosphorylative respirati on). Of this amount, 29% is inhibited by 5 X 10(-7) M bafilomycin A1 a lone and 90-95% by the combination of bafilomycin plus ouabain. These results indicate that the H+-ATPase activity is a significant energy-r equiring process in dog proximal tubules. If bafilomycin is added afte r a 5- to 7-min preincubation with 1 mM ouabain, then the bafilomycin- sensitive ATP turnover is larger, reaching 44% of total phosphorylatio n. This may suggest that the H+ pump is stimulated by the indirect inh ibition 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 extracell ular medium. In rabbit and hamster proximal tubules, the bafilomycin-s ensitive 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 sp ecies to species.