cAMP-dependent protein kinase mediates hydrosmotic effect of vasopressin in collecting duct.
The role of adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase A (PKA) in mediating the hydrosmotic effect of vasopressin in in vitro microperfused rabbit cortical collecting ducts (CCDs) was examined. We measured PKA substrate phosphorylation and water permeability [hydraulic conductivity (Lp) = 10(-7) cm.atm-1.s-1], stimulated by substituted cAMP analogues selective for a unique cAMP binding site (site A or B) on PKA regulatory subunit (R). Synergy between site A- and site B-selective analogues suggests involvement of PKA, because both sites must be occupied for R to dissociate from the catalytic subunit (C), allowing phosphorylation to proceed. As single agents, the site B-selective analogues 8-(4-chlorophenylthio)-cAMP (8-CPT) and 8-thiomethyl-cAMP (8-SCH3) were at least two orders of magnitude more potent than the site A-selective analogues N6-monobutyryl-cAMP (N6-mono) or N6-benzoyl-cAMP (N6-benz). Combinations of subthreshold concentrations of two site A analogues (N6-mono+N6-benz) or two site B-selective analogues (8-CPT + 8-SCH3) failed to significantly increase protein phosphorylation or water permeability. In contrast, combination of a site A plus site B analogue synergistically stimulated both protein phosphorylation and Lp. Rp-cAMPS, an inhibitor of cAMP binding to PKA, reduced both vasopressin (41% inhibition)- and cAMP (56% inhibition)-stimulated water permeability. H-89 (50 microM), an inhibitor of PKA kinase activity, also blocked cAMP-stimulated water permeability (90% inhibition). These findings suggest that vasopressin-induced water permeability in the rabbit CCD is mediated by PKA.
Snyder
H M
HM
Department of Veterans Affairs Medical Center, Nashville, Tennessee.
Noland
T D
TD
Breyer
M D
MD
D000818:Animals; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D007546:Isoquinolines / Q000494:pharmacology; D007685:Kidney Tubules, Collecting / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D009995:Osmosis; D010539:Permeability; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000502:physiology*; D011817:Rabbits; D013237:Stereoisomerism; D013449:Sulfonamides; D013806:Theophylline / Q000031:analogs & derivatives / Q000494:pharmacology; D014667:Vasopressins / Q000494:pharmacology*; D014867:Water / Q000378:metabolism
D007546:Isoquinolines; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; D014867:Water; D014667:Vasopressins; C053907:8-cyclopentyl-1,3-dimethylxanthine; D013806:Theophylline; D000242:Cyclic AMP; D011494:Protein Kinases; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide