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Am J Physiol Cell Physiol 273: C1458-C1465, 1997;
0363-6143/97 $5.00
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Vol. 273, Issue 5, C1458-C1465, November 1997

Receptor-mediated inhibition of renal Na+-K+-ATPase is associated with endocytosis of its alpha - and beta -subunits

Alexander V. Chibalin, Adrian I. Katz, Per-Olof Berggren, and Alejandro M. Bertorello

Department of Molecular Medicine, Karolinska Institutet, Rolf Luft Center for Diabetes Research, Karolinska Hospital, 171 76 Stockholm, Sweden; and Department of Medicine, The University of Chicago, Chicago, Illinois 60637

The mechanisms involved in receptor-mediated inhibition of Na+-K+-ATPase remain poorly understood. In this study, we evaluate whether inhibition of proximal tubule Na+-K+-ATPase activity by dopamine is linked to its removal from the plasma membrane and internalization into defined intracellular compartments. Clathrin-coated vesicles were isolated by sucrose gradient centrifugation and negative lectin selection, and early and late endosomes were separated on a flotation gradient. Inhibition of Na+-K+-ATPase activity by dopamine, in contrast to its inhibition by ouabain, was accompanied by a sequential increase in the abundance of the alpha -subunit in clathrin-coated vesicles (1 min), early endosomes (2.5 min), and late endosomes (5 min), suggesting its stepwise translocation between these organelles. A similar pattern was found for the beta -subunit. The increased incorporation of both subunits in all compartments was blocked by calphostin C. The results demonstrate that the dopamine-induced decrease in Na+-K+-ATPase activity in proximal tubules is associated with internalization of its alpha - and beta -subunits into early and late endosomes via a clathrin-dependent pathway and that this process is protein kinase C dependent. The presence of Na+-K+-ATPase subunits in endosomes suggests that these compartments may constitute normal traffic reservoirs during pump degradation and/or synthesis.

dopamine; proximal tubules; clathrin vesicles; endosomes; protein kinase C; actin-microtubule cytoskeleton


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