Localization of amiloride-sensitive sodium channels in A6 cells by atomic force microscopy

Author:

Smith P. R.1,Bradford A. L.1,Schneider S.1,Benos D. J.1,Geibel J. P.1

Affiliation:

1. Department of Physiology, Medical College of Pennsylvania andHahnemann University, Philadelphia 19129, USA.

Abstract

Atomic force microscopy (AFM) was used for high-resolution imaging of the apical distribution of epithelial Na+ channels in A6 renal epithelial cells. A6 cells grown on coverslips were labeled with antibodies generated against an amiloride-sensitive epithelial Na+ channel complex purified from bovine renal medulla that had been conjugated to 8-nm colloidal gold particles before preparation for AFM. AFM revealed that there was a marked increase in the height of the microvilli in cells labeled with the anti-epithelial Na+ channel antibodies compared with unlabeled cells or cells labeled with rabbit nonimmune immunoglobulin G conjugated to colloidal gold particles. We interpret this apparent increase in microvillar height to be due to anti-epithelial Na+ channel antibody binding to the apical microvilli. These data demonstrate that epithelial Na+ channels are restricted to the apical microvilli in Na+-transporting renal epithelial cells. Furthermore, they demonstrate the applicability of using AFM for high-resolution imaging of the cell surface distribution of epithelial ion channels.

Publisher

American Physiological Society

Subject

Cell Biology,Physiology

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