title: ANP and CNP activate CFTR expressed in Xenopus laevis oocytes by direct activation of PKA.
abstract: BACKGROUND
  Acting through different receptors, natriuretic peptides (atrial natriuretic peptide [ANP], brain type natriuretic peptide [BNP] and C-type natriuretic peptide [CNP]) increase intracellular cGMP, which then stimulates different pathways that activate fluid secretion.
  
  
  OBJECTIVE
  We used two-electrode voltage clamping to define the dominant pathway that is employed when natriuretic peptides activate cystic fibrosis transmembrane conductance regulator (CFTR) in the Xenopus oocyte expression system. Natriuretic peptides could activate CFTR by 1) cGMP cross-activation of protein kinase A (PKA), 2) cGMP activation of cGMP-dependent protein kinase II, 3) cGMP inhibition of phosphodiesterase type III (PDE3), or 4) direct activation of CFTR.
  
  
  METHODS
  cRNA-microinjected Xenopus laevis oocytes were perfused with diverse compounds that examined these pathways of natriuretic peptide signaling.
  
  
  CONCLUSIONS
  ANP stimulated the shark CFTR (sCFTR)-mediated chloride conductance and this activation was inhibited by H-89, a specific inhibitor of PKA. After co-expression of the CNP receptor (NPR-B), sCFTR became stimulatable by CNP and was similarly inhibited by H-89, pointing to cross-activation of PKA. 8-pCPT-cGMP, a relatively cGKII-selective cGMP, failed to stimulate sCFTR. Another membrane-permeable and non-hydrolyzable analog of cGMP, 8-Br-cGMP, stimulated CFTR only at millimolar concentrations, consistent with cross-activation of PKA. The PDE inhibitors EHNA, rolipram, cilostamide, and amrinone did not significantly increase chloride conductance, arguing against a significant role for PDE2, PDE3 and PDE4 signaling in the oocyte. Sildenafil, a PDE5 inhibitor, caused a partial activation of sCFTR channels and this effect was again inhibited by H-89.
  
  
  CONCLUSIONS
  From these experiments we conclude that in the Xenopus oocyte system, natriuretic peptides, 8-Br-cGMP, and PDE5 inhibitors activate CFTR by cross-activation of PKA.
authors.affiliation: a Nephrology Division, Department of Internal Medicine , Yale University School of Medicine , New Haven , CT , USA .|b The Mount Desert Island Biological Laboratory , Salisbury Cove , ME , USA , and.
authors.forename: Klaus
authors.initials: K
authors.lastname: Stahl
authors.affiliation: a Nephrology Division, Department of Internal Medicine , Yale University School of Medicine , New Haven , CT , USA .|b The Mount Desert Island Biological Laboratory , Salisbury Cove , ME , USA , and.
authors.forename: Maximilian
authors.initials: M
authors.lastname: Stahl
authors.affiliation: b The Mount Desert Island Biological Laboratory , Salisbury Cove , ME , USA , and.|c Department of Gastroenterology & Hepatology , Erasmus University Medical Center , Rotterdam , The Netherlands.
authors.forename: Hugo R
authors.initials: HR
authors.lastname: de Jonge
authors.affiliation: a Nephrology Division, Department of Internal Medicine , Yale University School of Medicine , New Haven , CT , USA .|b The Mount Desert Island Biological Laboratory , Salisbury Cove , ME , USA , and.
authors.forename: John N
authors.initials: JN
authors.lastname: Forrest
mesh_terms: D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000378:metabolism*; D004789:Enzyme Activation; D020098:Natriuretic Peptide, C-Type / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism*; D014982:Xenopus laevis
chemical_list: D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D020098:Natriuretic Peptide, C-Type; D009320:Atrial Natriuretic Factor; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: CFTR chloride channel; natriuretic peptides; xenopus laevis oocytes