Ca2+ and cAMP activate different K+ conductances in the human intestinal goblet cell line HT29-Cl.16E.
The mechanism of regulated Cl- secretion was evaluated in the mucin-secreting cell line HT29-Cl.16E by transepithelial electrophysiology and fura 2 measurements of cytosolic Ca2+. Carbachol by itself was a weak secretagogue, but augmented adenosine 3',5'-cyclic monophosphate (cAMP)-mediated secretion more than twofold, consistent with activation of a rate-limiting K+ conductance. To characterize this conductance, monolayers were apically permeabilized with amphotericin B. At least two types of K+ conductances were identified. One type was activated by elevated cytosolic cAMP levels and inhibited by Ba2+ (inhibitor constant 0.3 mM) in the basolateral solution but was not affected by quinidine or elevated cytosolic Ca2+. The other type was activated by carbachol via cytosolic Ca2+ and was partially inhibited by quinidine (60% inhibition by 2.5 mM quinidine) but was not affected by Ba2+ up to 1 mM. Both conductances appear to be involved in active, transepithelial Cl- secretion in intact monolayers but under different conditions because 1) the cAMP-stimulated short-circuit current (Isc) can be partially inhibited by 1 mM Ba2+ (50%) but not quinidine, 2) the Ba2+ inhibition does not affect the carbachol-induced increase in Isc in cells with elevated cAMP levels, and 3) the carbachol-dependent Isc can be inhibited by quinidine. Therefore, the contribution of the cAMP-dependent K+ conductance appears important for maintaining the membrane potential and therewith Cl- secretion when cAMP is the only messenger for secretion signals, whereas the Ca(2+)-dependent K+ conductance is responsible for the carbachol-stimulated increase in Isc.
Merlin
D
D
Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106-4970, USA.
Guo
X
X
Laboisse
C L
CL
Hopfer
U
U
D000666:Amphotericin B; D001464:Barium / Q000494:pharmacology; D002034:Bumetanide / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D002463:Cell Membrane Permeability; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D004072:Digitonin; D004159:Diphenylamine / Q000031:analogs & derivatives / Q000494:pharmacology; D004553:Electric Conductivity; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects / Q000502:physiology; D016257:Fura-2; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D008564:Membrane Potentials / Q000187:drug effects; D008954:Models, Biological; D010042:Ouabain / Q000494:pharmacology; D011188:Potassium / Q000378:metabolism*; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology*; D011802:Quinidine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology
D015221:Potassium Channels; D013873:Thionucleotides; D002034:Bumetanide; C059537:3',5-dichlorodiphenylamine-2-carboxylic acid; D005576:Colforsin; D001464:Barium; D014660:Vasoactive Intestinal Peptide; D000001:Calcimycin; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D010042:Ouabain; D000666:Amphotericin B; D002217:Carbachol; D004159:Diphenylamine; D000242:Cyclic AMP; D011802:Quinidine; D004072:Digitonin; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium; D002118:Calcium; D016257:Fura-2