beta3-adrenoceptor control the cystic fibrosis transmembrane conductance regulator through a cAMP/protein kinase A-independent pathway.
In human cardiac myocytes, we have previously identified a functional beta3-adrenoceptor in which stimulation reduces action potential duration. Surprisingly, in cardiac biopsies obtained from cystic fibrosis patients, beta3-adrenoceptor agonists produced no effects on action potential duration. This result suggests the involvement of cystic fibrosis transmembrane conductance regulator (CFTR) chloride current in the electrophysiological effects of beta3-adrenoceptor stimulation in non-cystic fibrosis tissues. We therefore investigated the control of CFTR activity by human beta3-adrenoceptors in a recombinant system: A549 human cells were intranuclearly injected with plasmids encoding CFTR and beta3-adrenoceptors. CFTR activity was functionally assayed using the 6-methoxy-N-(3-sulfopropyl)quinolinium fluorescent probe and the patch-clamp technique. Injection of CFTR-cDNA alone led to the expression of a functional CFTR protein activated by cAMP or cGMP. Co-expression of CFTR (but not of mutated DeltaF508-CFTR) with high levels of beta3-adrenoceptor produced an increased halide permeability under base-line conditions that was not further sensitive to cAMP or beta3-adrenoceptor stimulation. Patch-clamp experiments confirmed that CFTR channels were permanently activated in cells co-expressing CFTR and a high level of beta3-adrenoceptor. Permanent CFTR activation was not associated with elevated intracellular cAMP or cGMP levels. When the expression level of beta3-adrenoceptor was lowered, CFTR was not activated under base-line conditions but became sensitive to beta3-adrenoceptor stimulation (isoproterenol plus nadolol, SR 58611, or CGP 12177). This later effect was not prevented by protein kinase A inhibitors. Our results provide molecular evidence that CFTR but not mutated DeltaF508-CFTR is regulated by beta3-adrenoceptors expression through a protein kinase A-independent pathway.
Leblais
V
V
Laboratoire de Physiopathologie et de Pharmacologie Cellulaires et Moléculaires, INSERM CJF 96-01, France.
Demolombe
S
S
Vallette
G
G
Langin
D
D
Baró
I
I
Escande
D
D
Gauthier
C
C
D000818:Animals; D019556:COS Cells; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D003550:Cystic Fibrosis / Q000378:metabolism*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000378:metabolism*; D006801:Humans; D009154:Mutation; D009206:Myocardium / Q000378:metabolism; D011943:Receptors, Adrenergic, beta / Q000235:genetics / Q000378:metabolism*; D022702:Receptors, Adrenergic, beta-3; D015398:Signal Transduction
C505032:CFTR protein, human; D011943:Receptors, Adrenergic, beta; D022702:Receptors, Adrenergic, beta-3; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D017868:Cyclic AMP-Dependent Protein Kinases