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Am J Physiol Renal Physiol 280: F1093-F1106, 2001;
0363-6127/01 $5.00
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Vol. 280, Issue 6, F1093-F1106, June 2001

Immunocytochemical and immunoelectron microscopic localization of alpha -, beta -, and gamma -ENaC in rat kidney

Henrik Hager1, Tae-Hwan Kwon1,2, Anna K. Vinnikova3, Shyama Masilamani4, Heddwen L. Brooks4, Jørgen Frøkiaer5, Mark A. Knepper4, and Søren Nielsen1

1 Department of Cell Biology, Institute of Anatomy, University of Aarhus, DK-8000 Aarhus C; 2 Department of Physiology, School of Medicine, Dongguk University, Kyungju 780-714, Korea; 3 Research Service, McGuire Veterans Affairs Medical Center, Richmond, Virginia 23249; 4 Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892; and 5 Department of Clinical Physiology, Aarhus University Hospital and Institute of Experimental Clinical Research, DK-8200 Aarhus N, Denmark

Epithelial sodium channel (ENaC) subunit (alpha , beta , and gamma ) mRNA and protein have been localized to the principal cells of the connecting tubule (CNT), cortical collecting duct (CCD), and outer medullary collecting duct (OMCD) in rat kidney. However, the subcellular localization of ENaC subunits in the principal cells of these cells is undefined. The cellular and subcellular localization of ENaC subunits in rat kidney was therefore examined. Immunocytochemistry demonstrated the presence of all three subunits in principal cells of the CNT, CCD, OMCD, and IMCD. In cortex and outer medulla, confocal microscopy demonstrated a difference in the subcellular localization of subunits. alpha -ENaC was localized mainly in a zone in the apical domains, whereas beta - and gamma -ENaC were found throughout the cytoplasm. Immunoelectron microscopy confirmed the presence of ENaC subunits in both the apical plasma membrane and intracellular vesicles. In contrast to the labeling pattern seen in cortex, alpha -ENaC labeling in IMCD cells was distributed throughout the cytoplasm. In the urothelium covering pelvis, ureters, and bladder, immunoperoxidase and confocal microscopy revealed differences the presence of all ENaC subunits. As seen in CCD, alpha -ENaC was present in a narrow zone near the apical plasma membrane, whereas beta - and gamma -ENaC were dispersed throughout the cytoplasm. In conclusion, all three subunits of ENaC are expressed throughout the collecting duct (CD), including the IMCD as well as in the urothelium. The intracellular vesicular pool in CD principal cells suggests ENaC trafficking as a potential mechanism for the regulation of Na+ reabsorption.

aldosterone; collecting duct; urothelium; epithelial sodium channel; intracellular trafficking; sodium transport


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