Sch-28080 depletes intracellular ATP selectively in mIMCD-3 cells

Two H+-K+-ATPase isoforms are present in kidney: the gastric, highly sensit ive to Sch-28080, and the colonic, partially sensitive to ouabain. Upregula tion of Sch-28080-sensitive H+-K+-ATPase, or "gastric" H+-K+-ATPase, has be en demonstrated in hypokalemic rat inner medullary collecting duct cells (I MCDs). Nevertheless, only colonic H+-K+-ATPase mRNA and protein abundance i ncrease in this condition. This study was designed to determine whether Sch -28080 inhibits transporters other than the gastric H+-K+-ATPase. In the pr esence of bumetanide, Sch-28080 (200 mu M) and ouabain (2 mM) inhibited Rb- 86(+) uptake (>90%). That Rb-86(+) uptake was almost completely abolished b y Sch-28080 indicates an effect of this agent on the Na+-K+-ATPase. ATPase assays in membranes, or lysed cells, demonstrated sensitivity to ouabain bu t not Sch-28080. Thus the inhibitory effect of Sch-28080 was dependent on c ell integrity. Rb-86(+)-uptake studies without bumetanide demonstrated that ouabain inhibited activity by only 50%. Addition of Sch-28080 (200 mM) blo cked all residual activity. Intracellular ATP declined after Sch-28080 (200 mM) but recovered after removal of this agent. In conclusion, high concent rations of Sch-28080 inhibit K+-ATPase activity in mouse IMCD-3 (mIMCD-3) c ells as a result of ATP depletion.