Ouabain stimulates Na-K-ATPase through a sodium/hydrogen exchanger-1 (NHE-1)-dependent mechanism in human kidney proximal tubule cells

Recent investigations demonstrate increased Na/H exchanger-1 (NHE-1) activity and plasma levels of ouabain-like factor in spontaneously hypertensive rats. At nanomolar concentrations, ouabain increases Na-K-ATPase activity, induces cell proliferation, and activates complex signaling cascades. We hypothesize that the activity of NHE-1 and Na-K-ATPase are interdependent. To test whether treatment with picomolar ouabain regulates Na-K-ATPase through an NHE-1-dependent mechanism, we examined the role of NHE-1 in ouabain-mediated stimulation of Na-K-ATPase in kidney proximal tubule cell lines [opossum kidney (OK), HK-2, HKC-5, and HKC-11] and rat kidney basolateral membranes. Ouabain stimulated Na-K-ATPase activity and tyrosine phosphorylation in cells that express NHE-1 (OK, HKC-5, and HKC-11) but not in HK-2 cells that express very low levels of NHE-1. Inhibition of NHE-1 with 5 [micro]M EIPA, a NHE-1specific inhibitor, prevented ouabain-mediated stimulation of S6Rb uptake and Na-K-ATPase phosphorylation in OK, HKC-5, and HKC- 11 cells. Expression of wild-type NHE- 1 in HK2 cells restored regulation of Na-K-ATPase by picomolar ouahain. Treatment with picomolar ouabain increased membrane expression of Na-K-ATPase and enhanced NHE-1-Na-K-ATPase [alpha]-subunit association. Treatment with ouabain (1 [micro]g x kg body [wt.sup.-1] x [day.sup.-1]) increased Na-K-ATPase activity, expression, phosphorylation, and association with NHE-1 increased in rat kidney cortical basolateral membranes. Eight days' treatment with ouabain (1 [micro]g x kg body [wt.sup.-1] x [day.sup.-1]) resulted in increased blood pressure in these rats. These results suggest that the association of NHE-1 with Na-K-ATPase is critical for ouabain-mediated regulation of Na-K-ATPase and that these effects may play a role in cardioglycoside-stimulated hypertension. cardioglycosides; phosphorylation doi: 10.1152/ajprenal.00581.2009.