Characterization of a cyclic AMP-activated Cl-transport pathway in the apical membrane of a human colonic epithelial cell line.
This report describes a Cl- transport pathway in confluent monolayer cultures of the T84 human colonic carcinoma cell line which is: 1) activated by vasoactive intestinal polypeptide, or other agents which induce or mimic cAMP; 2) independent of extracellular Na+ or K+; 3) refractory to inhibition by 0.1 mM bumetanide and 1 mM 4-acetamido-4'-isothiocyanostilbene-2,-2'-disulfonic acid; 4) competitively inhibited by NO3-, I-, SCN-, and Br-; 5) inhibited in a noncompetitive-complex manner by the putative Cl- channel-blocking agent, N-phenylanthranilic acid; and 6) localized to the apical membrane of confluent monolayers. This Cl- transport system is, therefore, distinct from the bumetanide-sensitive, basolateral membrane-localized, Na+, K+, Cl- cotransport system previously described in these cells (Dharmsathaphorn, K., Mandel, K., Masui, H., and McRoberts, J.A. (1985) J. Clin. Invest. 75, 462-471). Kinetic studies revealed that Cl- transport by this pathway fit simple Michaelis-Menten kinetics with an apparent Km for Cl- of about 6 mM. Activation by vasoactive intestinal polypeptide increased the Vmax but did not alter the apparent Km. We discuss the possibility that this transport system is a Cl- channel which is intimately involved in hormonally mediated, electrogenic Cl- secretion across T84 cell monolayers.
D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000494:pharmacology; D000873:Anthracenes / Q000494:pharmacology; D001693:Biological Transport, Active; D001965:Bromides / Q000494:pharmacology; D003994:Bucladesine / Q000494:pharmacology; D002034:Bumetanide / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism*; D003106:Colon / Q000166:cytology*; D000242:Cyclic AMP / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004848:Epithelium / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012964:Sodium / Q000378:metabolism; D014634:Valinomycin / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology; D062367:ortho-Aminobenzoates / Q000494:pharmacology
D000873:Anthracenes; D001965:Bromides; D002712:Chlorides; D062367:ortho-Aminobenzoates; D002034:Bumetanide; D014634:Valinomycin; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D014660:Vasoactive Intestinal Peptide; D000001:Calcimycin; D003994:Bucladesine; C010030:9-anthroic acid; C001209:fenamic acid; D012964:Sodium; D000242:Cyclic AMP