Blocking kinetics of Cl- channels in colonic carcinoma cells (HT29) as revealed by 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB)

• Published in: Biochimica et biophysica acta Vol. 946; no. 1; pp. 135 - 142
• Main Authors: DREINHÖFER, J, GÖGELEIN, H, GREGER, R
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier 08.12.1988; North-Holland
• Subjects: Biological and medical sciences; Cell Line; Cell physiology; Chloride Channels; Chlorides - antagonists & inhibitors; Colonic Neoplasms - metabolism; Fundamental and applied biological sciences. Psychology; Humans; Kinetics; Mathematics; Membrane and intracellular transports; Membrane Proteins - antagonists & inhibitors; Molecular and cellular biology; Nitrobenzoates - pharmacology; Benzoic acid; Human; Cell culture; Molecular structure; Blocking; Hill coefficient; Tumor; Ionic channel; Kinetics; Colon; Chlorides Ions
• ISSN: 0006-3002; 1878-2434
• DOI: 10.1016/0005-2736(88)90466-X

The blocking effect of 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) was investigated on single Cl- channels of the cultured human colon carcinoma cells, HT29. In the absence of NPPB, the open-time histogram yielded two time constants, with 0.9 ms and 33 ms, whereas the closed-time distribution could be fitted by a single exponential with a time constant of 0.7 ms. Addition of NPPB in the range 1-50 microM induced brief closing events of the single-channel current. This resulted in a decrease of the long open-time constant to 2.1 ms and in an increase of the closed-time constant to 1.8 ms at 50 microM NPPB concentration. The short open-time constant did not change at low blocker concentration (1 microM), but could no longer be resolved at higher concentrations. The open-state probability decreased from 0.9 (control conditions) to 0.5 at 50 microM NPPB. The Hill plot yielded a Hill coefficient of about 0.7, compatible with one NPPB molecule inhibiting one channel molecule. The kinetics of channel gating are described by a sequential model with one closed and two open states. Since in the presence of NPPB no additional time constant appeared in the time histograms, we assumed the same kinetic scheme as under control conditions, and hypothesize that NPPB has an influence on rate constants.