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cAMP-dependent activation of small-conductance Cl− channels in HT29 colon carcinoma cells

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Abstract

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 umol/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 cellattached 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. 5-Nitro-2-(3-phenylpropylamino)-benzoate (1 μmol/l) was without significant effects in resting and in forskolin-stimulated HT29 cells. The agonist-induced conductance increase of the cell-attached nystatin patches was not paralleled by the appearance of detectable single-channel events in these membranes. These data suggest activation of small, non-resolvable Cl− channels by cAMP.

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Authors and Affiliations

  1. Physiologisches Institut der Albert-Ludwigs-Universität, Hermann-Herder-Strasse 7, W-7800, Freiburg, Federal Republic of Germany

    K. Kunzelmann, M. Grolik, R. Kubitz & R. Greger

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  1. K. Kunzelmann
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  2. M. Grolik
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  3. R. Kubitz
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  4. R. Greger
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Supported by DFG Gr 480/10 and BMFT 01 GA 88/6

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Kunzelmann, K., Grolik, M., Kubitz, R. et al. cAMP-dependent activation of small-conductance Cl− channels in HT29 colon carcinoma cells. Pflügers Arch 421, 230–237 (1992). https://doi.org/10.1007/BF00374832

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  • DOI: https://doi.org/10.1007/BF00374832

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