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Ouabain signaling requires angiotensin II type 1 receptor activation.
Author(s) -
Conner Clayton,
Duplessis Madalyn,
Lederer Eleanor,
Khundmiri Syed
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.845.31
Subject(s) - ouabain , angiotensin ii , endocrinology , medicine , chemistry , receptor , biology , sodium , biochemistry , organic chemistry
Low dose ouabain stimulates sodium transport in proximal tubules. Patients with congestive heart failure and CKD show significantly elevated concentrations (pM range) of endogenous ouabain (EO), and exogenous ouabain increases blood pressure in rats. IP of ouabain bound proteins from proximal tubule cells showed association with the angiotensin II type 1 receptor (AT1R), suggesting a direct pathway linking EO and RAS in mediating salt sensitive hypertension. We hypothesize that low dose ouabain signaling is dependent on AT1R activation. We treated human (HKC11), mouse (mRPT) and porcine (LLC‐PK1) kidney cells with pathophysiological concentrations of ouabain and performed confocal, Total Internal Reflection Fluorescence (TIRF), and Forster's Resonance Electron Transfer (FRET) microscopy to examine associations between AT1R, ouabain, and Na‐K ATPase a1 subunit (NKA). TIRF and FRET microscopy showed that ouabain binds directly to NKA but not to AT1R. NKA and AT1R associated directly with each other and treatment with ouabain increased association between NKA and AT1R. Ouabain increased phosphorylation of signaling proteins (Src, EGFR, and ERK), and increased activity ( 86 Rb uptake), tyrosine phosphorylation (IP), and surface expression (biotinylation) of NKA in cells expressing AT1R but not in cells that lack AT1R expression. Treatment with candesartan, an AT1R antagonist, prevented the ouabain‐stimulated effects on signaling and NKA. These results suggest that pathophysiological concentrations of ouabain may stimulate renal salt reabsorption through an AT1R‐dependent pathway.