title: ENaC activity is increased in isolated, split-open cortical collecting ducts from protein kinase Cα knockout mice.
abstract: The epithelial Na channel (ENaC) is negatively regulated by protein kinase C (PKC) as shown using PKC activators in a cell culture model. To determine whether PKCα influences ENaC activity in vivo, we examined the regulation of ENaC in renal tubules from PKCα⁻/⁻ mice. Cortical collecting ducts were dissected and split open, and the exposed principal cells were subjected to cell-attached patch clamp. In the absence of PKCα, the open probability (P₀) of ENaC was increased three-fold vs. wild-type SV129 mice (0.52 ± 0.04 vs. 0.17 ± 0.02). The number of channels per patch was also increased. Using confocal microscopy, we observed an increase in membrane localization of α-, β-, and γ-subunits of ENaC in principal cells in the cortical collecting ducts of PKCα⁻/⁻ mice compared with wild-type mice. To confirm this increase, one kidney from each animal was perfused with biotin, and membrane protein was pulled down with streptavidin. The nonbiotinylated kidney was used to assess total protein. While total ENaC protein did not change in PKCα⁻/⁻ mice, membrane localization of all the ENaC subunits was increased. The increase in membrane ENaC could be explained by the observation that ERK1/2 phosphorylation was decreased in the knockout mice. These results imply a reduction in ENaC membrane accumulation and P₀ by PKCα in vivo. The PKC-mediated increase in ENaC activity was associated with an increase in blood pressure in knockout mice fed a high-salt diet.
authors: Emory Univ. School of Medicine, Dept. of Physiology, Whitehead Biomedical Research Bldg., 615 Michael St., Atlanta, GA 30322. deaton@emory.edu.
authors: Hui-Fang
authors: HF
authors: Bao
authors: Tiffany L
authors: TL
authors: Thai
authors: Qiang
authors: Q
authors: Yue
authors: He-Ping
authors: HP
authors: Ma
authors: Amity F
authors: AF
authors: Eaton
authors: Hui
authors: H
authors: Cai
authors: Janet D
authors: JD
authors: Klein
authors: Jeff M
authors: JM
authors: Sands
authors: Douglas C
authors: DC
authors: Eaton
mesh_terms: D000450:Aldosterone / Q000097:blood; D000818:Animals; D051399:Aquaporin 2 / Q000378:metabolism; D001794:Blood Pressure / Q000187:drug effects; D053503:Epithelial Sodium Channels / Q000378:metabolism*; D007685:Kidney Tubules, Collecting / Q000166:cytology* / Q000502:physiology; D051379:Mice; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D051571:Protein Kinase C-alpha / Q000172:deficiency* / Q000378:metabolism; D017673:Sodium Chloride, Dietary / Q000008:administration & dosage
chemical_list: C496025:Aqp2 protein, rat; D051399:Aquaporin 2; D053503:Epithelial Sodium Channels; D017673:Sodium Chloride, Dietary; D000450:Aldosterone; D051571:Protein Kinase C-alpha; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3
keywords: ENaC; hypertension; knockout mice; protein kinase Cα; renal tubules; single channels