Ca2+ and cAMP activate different chloride efflux pathways in HT-29.cl19A colonic epithelial cell line.
The mechanism of adenosine 3',5'-cyclic monophosphate (cAMP)- and Ca(2+)-induced Cl- secretion was studied in monolayers of the colon carcinoma cell line HT-29.cl19A by combined short-circuit current (Isc) and 125I- or 36Cl- efflux measurements. Forskolin, a specific adenylate cyclase activator, was found to induce a large increase in Isc and a two- to threefold increase in 36Cl- efflux solely across the apical border. The fractional efflux of 36Cl-compared with 125I- (basal ratio 1.71 +/- 0.28) did not change significantly in the presence of forskolin (1.91 +/- 0.45). In contrast, the Ca2+ ionophore A23187 did not appreciably affect the Isc but enhanced 36Cl- and 125I- efflux at the apical and basolateral side of the monolayer. Furthermore, the fractional efflux ratio of 36Cl- to 125I- changed dramatically to a value of 0.36 +/- 0.14. Both forskolin- and A23187-induced 36Cl- or 125I- efflux were only weakly inhibited by the putative Cl- channel blocker 5-nitro-2-(3-phenylpropylamino)-benzoicacid. Carbachol, a Ca(2+)-linked agonist, mimicked the effects of A23187 on the 36Cl- and 125I- efflux but additionally provoked a significant increase in Isc. These data show that Ca2+ and cAMP activate different Cl-efflux pathways in HT-29.cl19A cells. Most likely these pathways represent a cAMP-activated conductance in the apical membrane and a separate Ca(2+)-activated Cl- conductance expressed in both apical and basolateral membranes. Apparently cholinergic agonists induce net electrogenic Cl- secretion through an intracellular signaling pathway (e.g., protein kinase C activation) different from the one activated by Ca2+/Ca2+ ionophore alone.
Vaandrager
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AB
Department of Biochemistry I, Medical Faculty, Erasmus University Rotterdam, The Netherlands.
Bajnath
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R
Groot
J A
JA
Bot
A G
AG
De Jonge
H R
HR
D001692:Biological Transport; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D002712:Chlorides / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D003106:Colon / Q000378:metabolism*; D003110:Colonic Neoplasms; D000242:Cyclic AMP / Q000378:metabolism*; D004848:Epithelium / Q000378:metabolism; D006801:Humans; D007454:Iodides / Q000378:metabolism; D007700:Kinetics; D011188:Potassium / Q000502:physiology; D015398:Signal Transduction; D014407:Tumor Cells, Cultured
D002121:Calcium Channel Blockers; D002712:Chlorides; D007454:Iodides; D005576:Colforsin; D000001:Calcimycin; D002217:Carbachol; D000242:Cyclic AMP; D011188:Potassium; D002118:Calcium