cAMP-dependent activation of small-conductance Cl- channels in HT29 colon carcinoma cells.
The present study was performed to examine the conductance properties in the colon carcinoma cell line HT29 and the activation of Cl- channels by cAMP. A modified cell-attached nystatin patch-clamp technique was used, allowing for the simultaneous recording of the cell membrane potential (PD) and the conductance properties of the cell-attached membrane. In resting cells, PD was -56 +/- 0.4 mV (n = 294). Changing the respective ion concentrations in the bath indicate that these cells possess a dominating K+ conductance and a smaller Cl- conductance. A significant non-selective cation conductance, which could not be inhibited by amiloride, was only observed in cells examined early after plating. The K+ conductance was reversibly inhibited by 1 - 5 mmol/l Ba2+. Stimulation of the cells by the secretagogues isoproterenol and vasointestinal polypeptide (VIP) depolarized PD and induced a Cl- conductance. Similar results were obtained with compounds increasing cytosolic cAMP: forskolin, 3-isobutyl-1-methylxanthine, cholera toxin and 8-bromoadenosine cyclic 3',5'-monophosphate (8-Br-cAMP). VIP (1 nmol/l, n = 10) and isoproterenol (1 mumol/l, n = 12) depolarized the cells dose-dependently and reversibly by 12 +/- 2 mV and 13 +/- 2 mV. The maximal depolarization was reached after some 20 s. The depolarization was due to increases in the fractional Cl- conductance. Simultaneously the conductance of the cell-attached membrane increased from 155 +/- 31 pS to 253 +/- 40 pS (VIP, n = 4) and from 170 +/- 43 pS to 268 +/- 56 pS (isoproterenol, n = 11), reflecting the gating of Cl- channels in the cell-attached membrane.(ABSTRACT TRUNCATED AT 250 WORDS)
Kunzelmann
K
K
Physiologisches Institut der Albert-Ludwigs-Universität, Freiburg, Federal Republic of Germany.
Grolik
M
M
Kubitz
R
R
Greger
R
R
D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D002277:Carcinoma / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism*; D003110:Colonic Neoplasms / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism*; D007473:Ion Channels / Q000378:metabolism*; D009579:Nitrobenzoates / Q000494:pharmacology; D009761:Nystatin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000318:Adrenergic beta-Agonists; D002712:Chlorides; D007473:Ion Channels; D009579:Nitrobenzoates; D009761:Nystatin; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D000242:Cyclic AMP