Pflügers Archiv

, Volume 422, Issue 6, pp 539–545 | Cite as

Regulation of chloride channels in the human colon carcinoma cell line HT29.cl19A

  • Maarten Kansen
  • Rajesh B. Bajnath
  • Jack A. Groot
  • Hugo R. de Jonge
  • Bob Scholte
  • André T. Hoogeveen
  • Jan Bijman
Transport Processes, Metabolism and Endocrinology; Kidney, Gastrointestinal Tract, and Exocrine Glands


Chloride (Cl) channels are important in the regulation of salt and water transport in secretory epithelial cells. A disturbed Cl secretion is the most consistent characteristic in the genetic disease cystic fibrosis. An outwardly rectifying Cl channel (OR) with a conductance of 25–50 pS had been proposed to play a major role in Cl secretion. Activation by Ca2+ and the protein kinases (PK) A and C (at less than 10 nM Ca2+) as well as inhibition by PKC (at 1 μM Ca2+) has been reported. In the present study, we have identified and characterized the OR in HT29.cl19A human colon carcinoma cells. The OR displayed a conductance of 31±4 pS (n=25). Its open probability in 10 nM Ca2+ was voltage-dependent in 50% of the patches, starting from 0.2 at -70 mV to 0.8 at 70 mV. The spontaneous activation in excised inside-out patches at −60 mV was Ca2+-dependent and decreased from 29% in 1 mM Ca2+ to 2% in 10 nM Ca2+. Active OR were found in (a) 25% of patches exposed to 10 nM Ca2+, ATP and cAMP only, (b) 42% of the patches exposed to 10 nM Ca2+, ATP and the catalytic subunit of PKA (CAK) and (c) 67% of the patches exposed to 1 mM Ca2+, ATP plus CAK. Inhibition of voltage-activated channels by addition of PKC in 1 μM or 1 mM Ca2+ was not observed. Attempts to activate the OR in cell-attached patches by increasing cAMP levels under different experimental conditions were unsuccessful. Our data suggest that the OR may not be as important in Cl secretion as has been thought.

Key words

Chloride channel Patch clamp Intestine 


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  1. 1.
    Bajnath RB, Augeron C, Laboisse CL, de Jonge HR, Groot JA (1991) Electrophysiological studies of the effects of forskolin on the human colon carcinoma cell line HT-29 cl. 19A. J Membr Biol 122:239–250Google Scholar
  2. 2.
    Bear CE (1988) Phosphorylation-activated chloride channels in human skin fibroblasts. FEBS Lett 237:145–149Google Scholar
  3. 3.
    Bubien JK, Kirk KL, Rado ThA, Frizzell RA (1990) Cell cycle dependence of chloride permeability in normal and cystic fibrosis lymphocytes. Science 248:1416–1419Google Scholar
  4. 4.
    Buchanan JA, Yeger H, Tabcharani JA, Jensen TJ, Auerbach W, Hanrahan JW, Riordan JR, Buchwald M (1990) Transformed sweat gland and nasal epithelial cell lines from control and cystic fibrosis individuals. J Cell Sci 95:109–123Google Scholar
  5. 5.
    Champigny G, Verrier B, Gérard C, Mauchamp J, Lazdunski M (1990) Small conductance chloride channels in the apical membrane of thyroid cells. FEBS Lett 259:263–268Google Scholar
  6. 6.
    Chen JH, Schulman H, Gardner P (1989) A cAMP-regulated chloride channel in lymphocytes that is affected in cystic fibrosis. Science 243:657–660Google Scholar
  7. 7.
    Christensen O, Simon M, Randlev T (1989) Anion channels in a leaky epithelium. A patch-clamp-study of Necturus choroid plexus. Pflügers Arch 415:37–46Google Scholar
  8. 8.
    Clancy JP, McCann JD, Li M, Welsh MJ (1990) Calciumdependent regulation of airway epithelial chloride channels. Am J Physiol 258:L25-L32Google Scholar
  9. 9.
    Cook DI, Poronnik P, Young JA (1990) Characterization of a 25 pS nonselective cation channel in a cultured secretory epithelial cell line. J Membr Biol 114:37–52Google Scholar
  10. 10.
    de Jonge HR, van den Berghe N, Tilly BC, Kansen M, Bijman J (1989) (Dys)regulation of epithelial chloride channels. Biochem Soc Trans 17:816–818Google Scholar
  11. 11.
    Frizzell RA, Halm DR, Rechkemmer GR, Shoemaker RL (1986) Chloride channel regulation in secretory epithelia. Fed Proc 45:2727–2731Google Scholar
  12. 12.
    Giraldez F, Murray KJ, Sepúlveda FV, Seppard DN (1989) Characterization of a phosphorylation-activated Cl-selective channel in isolated Necturus enterocytes. J Physiol (Lond) 416:517–537Google Scholar
  13. 13.
    Gray MA, Pollard CE, Coleman HL, Greenwell JR, Argent BE (1990) Anion selectivity and block of the small-conductance chloride channel on pancreatic duct cells. Am J Physiol 259:C752-C761Google Scholar
  14. 14.
    Greger R, Schlatter E, Gögelein H (1987) Chloride channels in the luminal membrane of the rectal gland of the dogfish (Squalus acanthis). Pflügers Arch 409:114–121Google Scholar
  15. 15.
    Guggino WB (1989) Chloride channels in cystic fibrosis patients. Response. Science 247:222Google Scholar
  16. 16.
    Hagiwara G, Krouse M, Müller U, Wine J (1989) Is regulation of a chloride channel in lymphocytes affected in cystic fibrosis? Science 246:1049Google Scholar
  17. 17.
    Halm DR, Rechkemmer GR, Schomacher RA, Frizzell RA (1988) Apical membrane chloride channels in a colonic cell line activated by secretory agonist. Am J Physiol 254:C505-C511Google Scholar
  18. 18.
    Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 395:85–100Google Scholar
  19. 19.
    Hayslett JP, Gögelein H, Kunzelmann K, Greger R (1987) Characteristics of apical chloride channels in human colon cells (HT29). Pflügers Arch 410:487–494Google Scholar
  20. 20.
    Hwang T-C, Lu L, Zeitlin PL, Gruenert DC, Huganir R, Guggino WB (1989) Cl channels in CF: lack of activation by protein kinase C and cAMP-dependent protein kinase. Science 244:1351–1353Google Scholar
  21. 21.
    Jetten AM, Yankaskas JR, Stutts MJ, Willumsen NJ, Boucher RC (1989) Persistence of abnormal chloride conductance regulation in transformed cystic fibrosis epithelia. Science 244:1472–1475Google Scholar
  22. 22.
    Kansen M, Keulemans J, Hoogeveen AT, Scholte B, Vaandrager AB, van der Kamp AWM, Sinaasappel M, Bot AGM, de Jonge HR, Bijman J (1992) Regulation of chloride transport in cultured normal and cystic fibrosis keratinocytes. Biochim Biophys Acta 1139:49–65Google Scholar
  23. 23.
    Kartner N, Hanrahan JW, Jensen TJ, Naismith AL, Sun S, Ackerlay CA, Reyes EF, Tsui L-C, Rommens JM, Bear CE, Riordan JR (1991) Expression of the cystic fibrosis gene in non-epithelial invertebrate cells produces a regulated anion conductance. Cell 84:681–691Google Scholar
  24. 24.
    Krick W, Disser J, Hazama A, Burckhardt G, Frömter E (1991) Evidence for a cytosolic inhibitor of epithelial chloride channels. Pflügers Arch 418:491–499Google Scholar
  25. 25.
    Krouse ME, Hagiwara G, Chen J, Lewiston NJ, Wine JJ (1989) Ion channels in normal and cystic fibrosis sweat gland cells. Am J Physiol 257:C129-C140Google Scholar
  26. 26.
    Kunzelmann K, Pavenstädt H, Greger R (1989) Properties and regulation of chloride channels in cystic fibrosis and normal airway cells. Pflügers Arch 415:172–182Google Scholar
  27. 27.
    Li M, McCann JD, Anderson MP, Clancy JP, Liedtke CM, Nairn AC, Greengard R Welsh MJ (1989) Regulation of chloride channels by protein kinase C in normal and cystic fibrosis airway cells. Science 244:1353–1356Google Scholar
  28. 28.
    Li M, McCann JD, Liedtke CM, Nairn AC, Greengard P, Welsh MJ (1988) Cyclic AMP-dependent protein kinase opens chloride channels in normal but not cystic fibrosis airway epithelium. Nature 331:358–360Google Scholar
  29. 29.
    Marunaka Y, Eaton DC (1990) Chloride channels in the apical membrane of a distal nephron A6 cell line. Am J Physiol 258:C352-C368Google Scholar
  30. 30.
    Milton RL, Caldwell JH (1990) Na current in membrane blebs: implications for channel mobility and patch clamp recording. J Neurosci 10:885–893Google Scholar
  31. 31.
    Sandle GI, Fraser G, Long S, Warhurst G (1990) A cAMPactivated chloride channel in the plasma membrane of cultured human gastric cells (HGT-1). Pflügers Arch 417:259–263Google Scholar
  32. 32.
    Scholte BJ, Kansen M, Hoogeveen AT, Willemse R, Rhim JS, Van der Kamp AWM, Bijman J (1989) Immortalization of nasal polyp epithelial cell from cystic fibrosis patients. Exp Cell Res 182:559–571Google Scholar
  33. 33.
    Schoumacher RA, Shoemaker RL, Halm DR, Tallant EA, Wallace RW, Frizzell RA (1987) Phosphorylation fails to activate chloride channels from cystic fibrosis airway cells. Nature 330:752–754Google Scholar
  34. 34.
    Schoumacher RA, Ran J, Iannuzzi MC, Bradbury NA, Wallace RW, Tom Hon C, Kelly DR, Schmid SM, Gelder FB, Rado TA, Frizzell RA (1990) A cystic fibrosis pancreatic adenocarcinoma cell line. Proc Natl Acad Sci USA 87:4012–4016Google Scholar
  35. 35.
    Tabcharani JA, Hanrahan JW (1991) On the activation of outwardly rectifying anion channels in excised patches. Am J Physiol 261:G992-G999Google Scholar
  36. 36.
    Tabcharani JA, Low W, Elie D, Hanrahan JW (1990) Low conductance chloride channel activated by cAMP in the epithelial cell line T84. FEBS Lett 270:157–164Google Scholar
  37. 37.
    Tilly BC, Kansen M, van Gageldonk PGM, van den Berghe N, Galjaard J, Bijman J, de Jonge HR (1991) G-proteins mediate intestinal chloride channel activation. J Biol Chem 266: 2036–2040Google Scholar
  38. 38.
    Ward CL, Krouse ME, Gruenert DC, Kopito RR, Wine JJ (1991) Cystic fibrosis gene expression is not correlated with rectifying Cl channels. Proc Natl Acad Sci USA 88:5277–5281Google Scholar
  39. 39.
    Welsh MJ, Li M, McCann JD (1989) Activation of normal and cystic fibrosis chloride channels by voltage, temperature and trypsin. J Clin Invest 84:2002–2007Google Scholar
  40. 40.
    Wine JJ, Solc CK (1989) Chloride channels in cystic fibrosis patients. Science 247:2722Google Scholar
  41. 41.
    Worrell RT, Butt AG, Cliff WH, Frizzell RA (1989) A volume-sensitive chloride conductance in human colonic cell line T84. Am J Physiol 256:C1111-C1119Google Scholar

Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • Maarten Kansen
    • 1
  • Rajesh B. Bajnath
    • 3
  • Jack A. Groot
    • 3
  • Hugo R. de Jonge
    • 2
  • Bob Scholte
    • 1
  • André T. Hoogeveen
    • 1
  • Jan Bijman
    • 1
  1. 1.Department of Cell Biology and Genetics, Medical FacultyErasmus University RotterdamRotterdamThe Netherlands
  2. 2.Department of Biochemistry I, Medical FacultyErasmus University RotterdamRotterdamThe Netherlands
  3. 3.Department of Experimental ZoologyUniversity of AmsterdamAmsterdamThe Netherlands

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