H+-ATPASES OF RENAL CORTICAL AND MEDULLARY ENDOSOMES ARE DIFFERENTIALLY SENSITIVE TO SCH-28080 AND OMEPRAZOLE

Adenosinetriphosphatase (ATPase) activity stimulated by K+ and inhibited by Sch-28080 (SCH), omeprazole (OME), and vanadate has been measured in microsomes from mammalian renal medulla and attributed to a kidney isoform of the H+-K+-ATPase. To determine whether the H+-K+-ATPase inhibitors could also inhibit the vacuolar (V)-type H+-adenosinetriphosphatase (H+-ATPase, i.e., HC pump) in mammalian intracellular vesicles, we examined their effects on bafilomycin-sensitive acidification in renal cortical vesicles (CEV) and medullary endocytic vesicles (MEV). Rats were injected with fluorescein isothiocya nate-labeled dextran, and labeled endosomes were enriched from kidney tissue homogenates by differential and Percoll density gradient centrifugation. In the CEV, the V-type H+ pump was inhibited 25% by SCH and 30% by OME (100 mu M each). Whereas the inhibition by OME was concentration and time dependent, the inhibition by SCH was only concentration dependent. Inhibition by these compounds was similar in the presence of 50 mM K+ (in = out) and in the complete absence of K+, thus ruling out a significant involvement of H+-K+-ATPase-mediated acidification. Inhibition, however, was not observed with 10 mu M SCH and OME. The sensitivity of the V-type H+ pump to 100 mu M SCH and OME in CEV was confirmed by the comparable inhibitions of intravesicular acidification observed in acridine orange fluorescence quench studies and by inhibition of P-i liberation in an ATPase assay. We also found that the V-type H+ pump in isolated rat liver endosomes is sensitive to 100 mu M SCH and OME to a similar degree. In the MEV, acidification was only weakly affected by 100 mu M SCH and OME, thus suggesting that H+-ATPases in endosomes from cortical and medullary tubules are different, possibly due to a previously described selective expression of subunit isoforms. Our finding indicates the importance of using low concentrations (<10 mu M) of OME and SCH in studies of H+-K+-ATPase in nongastric tissues to avoid misinterpretation of the data due to nonspecific inhibition of V-type H+-ATPases.