Localization of amiloride-sensitive sodium channels in A6 cells by atomic force microscopy.
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.
Smith
P R
PR
Department of Physiology, Medical College of Pennsylvania and Hahnemann University, Philadelphia 19129, USA.
Bradford
A L
AL
Schneider
S
S
Benos
D J
DJ
Geibel
J P
JP
D000584:Amiloride / Q000494:pharmacology*; D000818:Animals; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000648:ultrastructure; D007150:Immunohistochemistry; D007668:Kidney / Q000166:cytology / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D011817:Rabbits; D015222:Sodium Channels / Q000187:drug effects* / Q000378:metabolism*; D014018:Tissue Distribution; D014982:Xenopus laevis
D015222:Sodium Channels; D000584:Amiloride