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MEMBRANE TRANSPORTERS, ION CHANNELS, AND PUMPS

Regulation of apical NHE3 trafficking by ouabain-induced activation of the basolateral Na⁺-K⁺-ATPase receptor complex

Departments of ¹Medicine and ²Pharmacology, University of Toledo College of Medicine, Toledo, Ohio

Submitted 10 October 2007 ; accepted in final form 6 December 2007

The long-term effects of ouabain on transepithelial Na⁺ transport involve transcriptional downregulation of apical Na⁺/H⁺ exchanger isoform 3 (NHE3). The aim of this study was to determine whether ouabain could acutely regulate NHE3 via a posttranscriptional mechanism in LLC-PK1 cells. We observed that the basolateral, but not apical, application of ouabain for 1 h significantly reduced transepithelial Na⁺ transport. This effect was not due to changes in the integrity of tight junctions or increases in the intracellular Na⁺ concentration. Ouabain regulated the trafficking of NHE3 and subsequently inhibited its activity, a process independent of intracellular Na⁺ concentration. Ouabain-induced NHE3 trafficking was abolished by either cholesterol depletion or Src inhibition. Moreover, ouabain increased the intracellular Ca²⁺ concentration. Pretreatment of cells with the intracellular Ca²⁺ chelator BAPTA-AM blocked ouabain-induced trafficking of NHE3. Also, blockade of Na⁺-K⁺-ATPase endocytosis by a phosphatidylinositol 3-kinase inhibitor was equally effective in attenuating ouabain-induced NHE3 trafficking. These data indicate that ouabain acutely stimulates NHE3 trafficking by activating the basolateral Na⁺-K⁺-ATPase signaling complex. Taken together with our previous observations, we propose that ouabain can simultaneously regulate basolateral Na⁺-K⁺-ATPase and apical NHE3, leading to inhibition of transepithelial Na⁺ transport. This mechanism may be relevant to proximal tubular Na⁺ handling during conditions associated with increases in circulating endogenous cardiotonic steroids.

kidney; sodium; sodium/hydrogen exchanger isoform 3; trafficking; c-Src; phosphatidylinositol 3-kinase

This article has been cited by other articles:

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