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The amiloride-sensitive epithelial sodium channel (ENaC) is usually found in the apical membrane of epithelial cells but has also recently been described in vascular endothelium. Because little is known about the regulation and cell surface density of ENaC, we studied the influence of aldosterone, spironolactone, and amiloride on its abundance in the plasma membrane of human endothelial cells. Three different methods were applied, single ENaC molecule detection in the plasma membrane, quantification by Western blotting, and cell surface imaging using atomic force microscopy. We found that aldosterone increases the surface expression of ENaC molecules by 36% and the total cellular amount by 91%. The aldosterone receptor antagonist spironolactone prevents these effects completely. Acute application of amiloride to aldosterone-pretreated cells led to a decline of intracellular ENaC by 84%. We conclude that, in vascular endothelium, aldosterone induces ENaC expression and insertion into the plasma membrane. Upon functional blocking with amiloride, the channel disappears from the cell surface and from intracellular pools, indicating either rapid degradation and/or membrane pinch-off. This opens new perspectives in the regulation of ENaC expressed in the vascular endothelium.

Original publication

DOI

10.1007/s00424-007-0341-0

Type

Journal article

Journal

Pflugers Archiv : European journal of physiology

Publication Date

02/2008

Volume

455

Pages

849 - 857

Addresses

Institute of Animal Physiology, University of Muenster, Hindenburgplatz 55, 48143 Muenster, Germany. kusche@uni-muenster.de

Keywords

Umbilical Veins, Cells, Cultured, Cell Membrane, Endothelial Cells, Humans, Spironolactone, Amiloride, Aldosterone, Diuretics, Microscopy, Atomic Force, Fluorescent Antibody Technique, Blotting, Western, Cell Membrane Permeability, Epithelial Sodium Channels