Ouabain Regulates CFTR-Mediated Anion Secretion and Na,K-ATPase Transport in ADPKD Cells.
Cyst enlargement in autosomal dominant polycystic kidney disease (ADPKD) requires the transepithelial secretion of fluid into the cyst lumen. We previously showed that physiological amounts of ouabain enhance cAMP-dependent fluid secretion and cyst growth of human ADPKD cyst epithelial cells in culture and formation of cyst-like dilations in metanephric kidneys from Pkd1 mutant mice. Here, we investigated the mechanisms by which ouabain promotes cAMP-dependent fluid secretion and cystogenesis. Ouabain (3 nM) enhanced cAMP-induced cyst-like dilations in embryonic kidneys from Pkd1 (m1Bei) mice, but had no effect on metanephroi from Pkd1 (m1Bei) mice that lack expression of the cystic fibrosis transmembrane conductance regulator (CFTR). Similarly, ouabain stimulation of cAMP-induced fluid secretion and in vitro cyst growth of ADPKD cells were abrogated by CFTR inhibition, showing that CFTR is required for ouabain effects on ADPKD fluid secretion. Moreover, ouabain directly enhanced the cAMP-dependent Cl(-) efflux mediated by CFTR in ADPKD monolayers. Ouabain increased the trafficking of CFTR to the plasma membrane and up-regulated the expression of the CFTR activator PDZK1. Finally, ouabain decreased plasma membrane expression and activity of the Na,K-ATPase in ADPKD cells. Altogether, these results show that ouabain enhances net fluid secretion and cyst formation by activating apical anion secretion via CFTR and decreasing basolateral Na(+) transport via Na,K-ATPase. These results provide new information on the mechanisms by which ouabain affects ADPKD cells and further highlight the importance of ouabain as a non-genomic stimulator of cystogenesis in ADPKD.
Department of Molecular and Integrative Physiology, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS, 66160, USA.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA.
Kyle
K
Jansson
Department of Molecular and Integrative Physiology, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS, 66160, USA.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA.
Jessica
J
Venugopal
Department of Molecular and Integrative Physiology, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS, 66160, USA.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA.
Gladis
G
Sánchez
Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, KS, USA.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA.
Brenda S
BS
Magenheimer
Department of Medicine, University of Kansas Medical Center, Kansas City, KS, USA.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA.
Gail A
GA
Reif
Department of Molecular and Integrative Physiology, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS, 66160, USA.|Department of Medicine, University of Kansas Medical Center, Kansas City, KS, USA.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA.
Darren P
DP
Wallace
Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, KS, USA.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA.
James P
JP
Calvet
Department of Molecular and Integrative Physiology, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS, 66160, USA. gblanco@kumc.edu.|The Kidney Institute, University of Kansas Medical Center, Kansas City, KS, USA. gblanco@kumc.edu.
Gustavo
G
Blanco
D000818:Animals; D000838:Anions / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism; D005576:Colforsin / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004195:Disease Models, Animal; D055786:Gene Knockout Techniques; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins; D051379:Mice; D018345:Mice, Knockout; D009924:Organ Culture Techniques; D010042:Ouabain / Q000494:pharmacology*; D016891:Polycystic Kidney, Autosomal Dominant / Q000235:genetics / Q000378:metabolism*; D061251:Primary Cell Culture; D028021:Sodium-Potassium-Chloride Symporters / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D000838:Anions; D002712:Chlorides; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C514896:PDZK1 protein, mouse; D028021:Sodium-Potassium-Chloride Symporters; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase
Cystogenesis; Fluid secretion; Polycystic kidney disease; Signaling