cAMP-activated Cl channels in CFTR-transfected cystic fibrosis pancreatic epithelial cells

W. H. Cliff, Robert Schoumacher, R. A. Frizzell

Research output: Contribution to journalArticle

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Abstract

Retrovirus-mediated transfection of cDNA for the cystic fibrosis (CF) gene into the CF pancreatic cell line, CFPAC-1, confers adenosine 3',5'-cyclic monophosphate (cAMP)-dependent regulation of Cl conductance. We used patch- clamp techniques to identify the single-channel basis of this conductance pathway and to study its properties. Forskolin or cAMP activated Cl channels with a conductance of 9 ± 1 pS in 26 of 62 cell-attached patches of cystic fibrosis transmembrane conductance regulator (CFTR)-transfected CFPAC-1 cells. The current-voltage (I-V) relation showed slight outward rectification (chord conductance of 10 ± 2 pS at +80 mV vs. 7 ± 1 pS at -80mV) with high Cl concentrations (170 mM) in the pipette solution. Channel kinetics were voltage sensitive, with longer openings at positive clamp voltages. Channel properties were unaffected by the substitution of N-methyl-D-glucamine for pipette Na or by the addition of disulfonic stilbenes (100 μM DNDS or DIDS) to the pipette. The channels usually inactivated within seconds of patch excision, but in three of nine patches, activity could be maintained by addition of the catalytic subunit of protein kinase A and ATP. With equal Cl concentrations on both membrane surfaces, the single-channel I-V relation was linear, suggesting that the outward rectification of the cell-attached channel is due to a pipette-to-cell Cl gradient. Anion substitution on the extracellular side of the membrane indicates a halide permselectivity of Br ~ Cl > I. The cAMP responsiveness, linear I-V relation, stilbene insensitivity, and halide permselectivity of this small-conductance Cl channel suggest that it is responsible for the cAMP-activated whole cell Cl current.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume262
Issue number5 31-5
StatePublished - Jun 30 1992
Externally publishedYes

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Cystic Fibrosis Transmembrane Conductance Regulator
Cystic Fibrosis
Cyclic AMP
Epithelial Cells
Stilbenes
4,4'-dinitro-2,2'-stilbenedisulfonic acid
Clamping devices
Electric potential
Substitution reactions
Cyclic AMP-Dependent Protein Kinase Catalytic Subunits
4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid
Membranes
Colforsin
Anions
Patch-Clamp Techniques
Retroviridae
Complementary DNA
Genes
Adenosine Triphosphate
Cells

All Science Journal Classification (ASJC) codes

  • Physiology
  • Cell Biology

Cite this

cAMP-activated Cl channels in CFTR-transfected cystic fibrosis pancreatic epithelial cells. / Cliff, W. H.; Schoumacher, Robert; Frizzell, R. A.

In: American Journal of Physiology - Cell Physiology, Vol. 262, No. 5 31-5, 30.06.1992.

Research output: Contribution to journalArticle

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