Cyclic AMP increases cell surface expression of functional Na,K-ATPase units in mammalian cortical collecting duct principal cells.
Cyclic AMP (cAMP) stimulates the transport of Na(+) and Na,K-ATPase activity in the renal cortical collecting duct (CCD). The aim of this study was to investigate the mechanism whereby cAMP stimulates the Na,K-ATPase activity in microdissected rat CCDs and cultured mouse mpkCCD(c14) collecting duct cells. db-cAMP (10(-3) M) stimulated by 2-fold the activity of Na,K-ATPase from rat CCDs as well as the ouabain-sensitive component of (86)Rb(+) uptake by rat CCDs (1.7-fold) and cultured mouse CCD cells (1.5-fold). Pretreatment of rat CCDs with saponin increased the total Na,K-ATPase activity without further stimulation by db-cAMP. Western blotting performed after a biotinylation procedure revealed that db-cAMP increased the amount of Na,K-ATPase at the cell surface in both intact rat CCDs (1.7-fold) and cultured cells (1.3-fold), and that this increase was not related to changes in Na,K-ATPase internalization. Brefeldin A and low temperature (20 degrees C) prevented both the db-cAMP-dependent increase in cell surface expression and activity of Na,K-ATPase in both intact rat CCDs and cultured cells. Pretreatment with the intracellular Ca(2+) chelator bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid also blunted the increment in cell surface expression and activity of Na,K-ATPase caused by db-cAMP. In conclusion, these results strongly suggest that the cAMP-dependent stimulation of Na,K-ATPase activity in CCD results from the translocation of active pump units from an intracellular compartment to the plasma membrane.
Gonin
S
S
Division de Néphrologie, Fondation pour Recherches Médicales, CH-1211 Genève 4, Switzerland.
Deschênes
G
G
Roger
F
F
Bens
M
M
Martin
P Y
PY
Carpentier
J L
JL
Vandewalle
A
A
Doucet
A
A
Féraille
E
E
D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D003994:Bucladesine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D066298:In Vitro Techniques; D007672:Kidney Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008322:Mammals; D051379:Mice; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D012503:Saponins / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000187:drug effects / Q000378:metabolism*; D013696:Temperature
D002614:Chelating Agents; D011500:Protein Synthesis Inhibitors; D012503:Saponins; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D020126:Brefeldin A; D004533:Egtazic Acid; D003994:Bucladesine; D000242:Cyclic AMP; D000254:Sodium-Potassium-Exchanging ATPase; D002118:Calcium