Expression of CFTR controls cAMP-dependent activation of epithelial K+ currents.
The perforated-patch configuration of the patch-clamp technique was used to record whole cell currents from human epithelial CFPAC-1 cells defective for functional cystic fibrosis transmembrane conductance regulator (CFTR). In CFPAC-1 cells, adenosine 3',5'-cyclic monophosphate (cAMP) stimulation with forskolin (10 microM) plus 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate (400 microM) activated neither Cl- nor K+ currents. In the same cells transfected with wild-type CFTR gene, cAMP stimulation produced activation of both Cl- and K+ currents. In Cl(-)-depleted medium (gluconate as a substitute), cAMP stimulation evoked a K+ current in CFTR-transfected but not in untransfected CFPAC-1 cells. This cAMP-evoked K+ current was the sum of two components: 1) a time-independent inwardly rectifying component, and 2) a slowly relaxing component activated at positive voltages. Increasing intracellular Ca2+ with ionomycin (1 microM) activated K+ currents in either transfected or untransfected cells. In transfected cells, blocking the CFTR conductance with high-concentration glibenclamide (100 microM) reduced the K+ current when activated by cAMP but not when activated by Ca2+. Pretreating CFTR-transfected cells for 48 h with interferon-gamma downregulated CFTR gene expression and reduced cAMP but not Ca2+ activation of the whole cell K+ current. From these results, we conclude that functional membrane CFTR protein influences activation by cAMP of epithelial K+ currents.
Loussouarn
G
G
Laboratoire de Physiopathologie et de Pharmacologie Cellulaires et Moléculaires, Hôpital G & R Laënnec, Nantes, France.
Demolombe
S
S
Mohammad-Panah
R
R
Escande
D
D
Baró
I
I
D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D018999:Charybdotoxin / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000096:biosynthesis / Q000502:physiology*; D004848:Epithelium; D005905:Glyburide / Q000494:pharmacology; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D010179:Pancreas; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D013873:Thionucleotides / Q000494:pharmacology; D014162:Transfection
C505032:CFTR protein, human; D015221:Potassium Channels; D011994:Recombinant Proteins; D013873:Thionucleotides; D018999:Charybdotoxin; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D015759:Ionomycin; D000242:Cyclic AMP; D005905:Glyburide; D002118:Calcium