Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Basolateral K channel activated by carbachol in the epithelial cell line T84


Cholinergic stimulation of chloride secretion involves the activation of a basolateral membrane potassium conductance, which maintains the electrical gradient favoring apical Cl efflux and allows K to recycle at the basolateral membrane. We have used transepithelial short-circuit current (I SC), fluorescence imaging, and patch clamp studies to identify and characterize the K channel that mediates this response in T84 cells. Carbachol had little effect on I SC when added alone but produced large, transient currents if added to monolayers prestimulated with cAMP. cAMP also enhanced the subsequent I SC response to calcium ionophores. Carbachol (100 μm) transiently elevated intracellular free calcium ([Ca2+] i ) by ∼3-fold in confluent cells cultured on glass coverslips with a time course resembling the I sc response of confluent monolayers that had been grown on porous supports. In parallel patch clamp experiments, carbachol activated an inwardly rectifying potassium channel on the basolateral aspect of polarized monolayers which had been dissected from porous culture supports. The same channel was transiently activated on the surface of subconfluent monolayers during stimulation by carbachol. Activation was more prolonged when cells were exposed to calcium ionophores. The conductance of the inward rectifier in cell-attached patches was 55 pS near the resting membrane potential (−54 mV) with pipette solution containing 150 mm KCl (37°C). This rectification persisted when patches were bathed in symmetrical 150 mm KCl solutions. The selectivity sequence was 1 K > 0.88 Rb > 0.18 Na ≫ Cs based on permeability ratios under bi-ionic conditions. The channel exhibited fast block by external sodium ions, was weakly inhibited by external TEA, was relatively insensitive to charybdotoxin, kaliotoxin, 4-aminopyridine and quinidine, and was unaffected by external 10 mm barium. It is referred to as the KBIC channel based on its most distinctive properties (Ba-insensitive, inwardly rectifying, Ca-activated). Like single KBIC channels, the carbachol-stimulated I SC was relatively insensitive to several blockers on the basolateral side and was unaffected by barium. These comparisons between the properties of the macroscopic current and single channels suggest that the KBIC channel mediates basolateral membrane K conductance in T84 cell monolayers during stimulation by cholinergic secretagogues.

This is a preview of subscription content, log in to check access.


  1. 1.

    Al-Bazzaz, F., Jayaram, T. 1981. Ion transport by canine tracheal mucosa: Effect of elevation of cellular calcium. Exp. Lung Res. 2:121–130

  2. 2.

    Anderson, M.P., Welsh, M.J. 1991. Calcium and cAMP activate different chloride channels in the apical membrane of normal and cystic fibrosis epithelia. Proc. Natl. Acad. Sci. USA 88:6003–6007

  3. 3.

    Bolton, J.E., Field, M. 1977. Ca ionophore-stimulated ion secretion in rabbit ileal mucosa: Relation to actions of cyclic 3′,5′-AMP and carbamylcholine. J. Membrane Biol. 35:159–173

  4. 4.

    Burgess, G.M., Bird, G.St.J., Obie, J.F., Putney, J.W., Jr. 1991. The mechanism for synergism between phospholipase C and adenylyl cyclase-linked hormones in liver. cAMP-dependent protein kinase augments IP3 mobilization without increasing the cellular levels of inositol polyphosphates. J. Biol. Chem. 266:4772–4781

  5. 5.

    Butt, A.G., Clapp, W.L., Frizzell, R.A. 1990. Potassium conductances in tracheal epithelium activated by secretion and cell swelling. Am. J. Physiol. 258:C630-C638

  6. 6.

    Cartwright, C.A., McRoberts, J.A., Mandel, K.G., Dharmsathaphorn, K. Synergistic action of cyclic adenosine monophosphate- and calcium-mediated chloride secretion in a colonic epithelial cell line. J. Clin. Invest. 76:1837–1842

  7. 7.

    Cereijido, M., Robbins, E.S., Dolan, W.J., Rotunno, C.A., Sabatini, D.D. 1978. Polarized monolayers formed by epithelial cells on a permeable and translucent support. J. Cell Biol. 77:853–880

  8. 8.

    Cohn, J.A. 1990. Protein kinase C mediates cholinergically regulated protein phosphorylation in a Cl-secreting epithelium. Am. J. Physiol. 258:C227-C233

  9. 9.

    Dawson, D.C., Richards, N.W. 1990. Basolateral K conductance: role in regulation of NaCl absorption and secretion. Am. J. Physiol. 259:C181-C195

  10. 10.

    Devor, D.C., Frizzell, R.A. 1993. Calcium-mediated agonists activate an inwardly rectified K+ channel in colonic secretory cells. FASEB J. 7:A350 (Abstr.)

  11. 11.

    Devor, D.C., Frizzell, R.A. 1993. Calcium-mediated agonists activate an inwardly rectified K+ channel in colonic secretory cells. Am. J. Physiol. 265:C1271-C1280

  12. 12.

    Dharmsathaphorn, K., Cohn, J., Beuerlein, G. 1989. Multiple calcium-mediated effector mechanisms regulate chloride secretory responses in T84-cells. Am. J. Physiol. 256:C1224-C1230

  13. 13.

    Dharmsathaphorn, K., McRoberts, J.A., Mandel, K.G., Tisdale, L.D., Masui, H. 1984. A human colonic tumor cell line that maintains vectorial electrolyte transport. Am. J. Physiol. 246:G204-G208

  14. 14.

    Dharmsathaphorn, K., Pandol, S.J. 1986. Mechanism of chloride secretion induced by carbachol in a colonic epithelial cell line. J. Clin. Invest. 77:348–354

  15. 15.

    Frizzell, R.A. 1977. Active chloride secretion by rabbit colon: Calcium-dependent stimulation by ionophore A23187. J. Membrane Biol. 35:175–187

  16. 16.

    Germann, W.J., Ernst, S.A., Dawson, D.C. 1986. Resting and osmotically induced basolateral potassium conductances in turtle colon. J. Gen. Physiol. 88:253–274

  17. 17.

    Germann, W.J., Lowy, M.E., Ernst, S.A., Dawson, D.C. 1986. Differentiation of two distinct potassium conductances in the basolateral membrane of turtle colon. J. Gen. Physiol. 88:237–251

  18. 18.

    Grygorczyk, C., Grygorczyk, R. 1991. Dual action of carbachol on chloride secretion in colonic epithelial cells (T84). Biophys. J. 59:646a (Abstr.)

  19. 19.

    Hanrahan, J.W., Tabcharani, J.A. 1990. Inhibition of an outwardly rectifying anion channel by HEPES and related buffers. J. Membrane Biol. 116:65–77

  20. 20.

    Hanrahan, J.W., Tabcharani, J.A. 1993. Basolateral K channel in T84 cells activated by carbachol. FASEB J. 7:A351 (Abstr.)

  21. 21.

    Hille, B. 1992. Ionic channels of excitable membranes. Second edition. Sinauer Associates, Sunderland, MA

  22. 22.

    Ho, K., Nichols, C.G., Lederer, W.J., Lytton, J., Vassilev, P.M., Kanazirska, M.V., Hebert, S.C. 1993. Cloning and expression of an inwardly rectifying ATP-regulated potassium channel. Nature 362:31–38

  23. 23.

    Isaacs, P.E.T., Corbet, C.L., Riley, A.K., Hawker, P.C., Turnberg, L.A. 1976. In vitro behaviour of human intestinal mucosa. The influence of acetylcholine on ion transport. J. Clin. Invest. 58:535–542

  24. 24.

    Kachintorn, U., Vajanaphanich, M., Barrett, K.E., Traynor-Kaplan, A.E. 1993. Elevation of inositol tetrakisphosphate parallels inhibition of Ca2+-dependent Cl secretion in T84 cells. Am. J. Physiol. 264:C671-C676

  25. 25.

    Kachintorn, U., Vajanaphanich, M., Traynor-Kaplan, A.E., Dharmsathaphorn, K., Barrett, K.E. 1993. Activation by calcium alone of chloride secretion in T84 epithelial cells. Br. J. Pharmacol. 109:510–517

  26. 26.

    Kachintorn, U., Vongkovit, P., Vajanaphanich, M., Dinh, S., Barrett, K.E., Dharmsathaphorn, K. 1992. Dual effects of a phorbol ester on calcium-dependent chloride secretion by T84 epithelial cells. Am. J. Physiol. 262:C15-C22

  27. 27.

    Grynkiewicz, G., Peonie, M., Tsien, R.Y. 1985. A new generation of Ca2+ indicators with greatly improved fluorescence properties. J. Biol. Chem. 260:3440–3450

  28. 28.

    Mandel, K.G., McRoberts, J.A., Beuerlein, G., Foster, E.S., Dharmsathaphorn, K. 1986. Ba2+ inhibition of VIP- and A23187-stimulated Cl secretion by T84 cell monolayers. Am. J. Physiol. 250:C486-C494

  29. 29.

    Marty, A., Tan, Y.P., Trautmann, A. 1984. Three types of calciumdependent channel in rat lacrimal glands. J. Physiol. 357:293–325

  30. 30.

    Maruyama, Y., Gallacher, D.V., Petersen, O.H. 1984. Voltage and Ca2+-activated K+ channel in baso-lateral acinar cell membranes of mammalian salivary glands. Nature 302:827–829

  31. 31.

    McCann, J.D., Ghalla, R.C., Welsh, M.J. 1989. Release of intracellular calcium by two different second messengers in airway epithelium. Am. J. Physiol. 257:L116-L124

  32. 32.

    McCann, J.D., Matsuda, J., Garcia, M., Kaczorowski, G., Welsh, M.J. 1990. Basolateral K+ channels in airway epithelia I. Regulation by Ca2+ and block by charybdotoxin. Am. J. Physiol. 258:L334-L342

  33. 33.

    McCann, J.D., Welsh, M.J. 1990. Basolateral K+ channels in airway epithelia. II. Role in Cl secretion and evidence for two types of K+ channel. Am. J. Physiol. 258:L343-L348

  34. 34.

    McRoberts, J.A., Beuerlein, G., Dharmsathaphorn, K. 1985. Cyclic AMP and Ca2+-activated K+ transport in a human colonic epithelial cell line. J. Biol. Chem. 260:14163–14172

  35. 35.

    Miller, C., Moczydlowski, E., Latorre, R., Phillips, M. 1985. Charybdotoxin, a protein inhibitor of single Ca2+-activated K+ channels from mammalian skeletal muscle. Nature 313:316–318

  36. 36.

    Reenstra, W.W. 1993. Inhibition of cAMP- and Ca-dependent Cl secretion by phorbol esters: inhibition of basolateral K+ channels. Am. J. Physiol. 264:C161-C168

  37. 37.

    Reinlib, L., Mikkelsen, R., Zahniser, D., Dharmsathaphorn, K., Donowitz, M. 1989. Carbachol-induced cytosolic free Ca2+ increases in T84 colonic cells seen by microfluorimetry. Am. J. Physiol. 257:G950-G960

  38. 38.

    Rich, A., Dixon, T.E., Clausen, C. 1990. Changes in membrane conductances and areas associated with bicarbonate secretion in turtle bladder. J. Membrane Biol. 113:211–219

  39. 39.

    Richards, N.W., Lowy, R.J., Ernst, S.A., Dawson, D.C. 1989. Two K+ channel types, muscarinic agonist-activated and inwardly rectifying, in a Cl secretory epithelium: the avian salt gland. J. Gen. Physiol. 93:1171–1194

  40. 40.

    Roch, B., Baró, I., Hongre, A.S., Escande, D. 1993. Charybdotoxin-sensitive Ca2+-activated K+ channels in human epithelial cells. Ped. Pulmonol. Suppl. 9:227

  41. 41.

    Shorofsky, S.R., Field, M., Fozzard, H.A. 1983. Electrophysiology of Cl secretion in canine trachea. J. Membrane Biol. 72:105–115

  42. 42.

    Smith, P.L., Frizzell, R.A. 1984. Chloride secretion by canine tracheal epithelium: IV. Basolateral membrane K permeability parallels secretion rate. J. Membrane Biol. 77:187–199

  43. 43.

    Supplisson, S., Loo, D.D.F., Sachs, G. 1991. Diversity of K+ channels in the basolateral membrane of resting Necturus oxyntic cells. J. Membrane Biol. 123:209–221

  44. 44.

    Tabcharani, J.A., Boucher, A., Eng, J.W.L., Hanrahan, J.W. (1994). Regulation of an inwardly rectifying K channel in the T84 epithelial cell line by calcium, nucleotides and kinases. J. Membrane Biol. 142:(in press)

  45. 45.

    Tabcharani, J.A., Hanrahan, J.W. 1991. Single chloride and potassium channels in the epithelial cell line T84. Biophys. J. 59:6a (Abstr.)

  46. 46.

    Tabcharani, J.A., Low, W., Elie, D., Hanrahan, J.W. 1990. Lowconductance chloride channel activated by cAMP in the epithelial cell line T84. FEBS Lett. 270:157–164

  47. 47.

    Trautmann, A., Marty, A. 1984. Activation of Ca-dependent K channels by carbamoylcholine in rat lacrimal glands. Proc. Natl. Acad. Sci. USA 81:611–615

  48. 48.

    Tsien, R.Y., Harootunian, A.T. 1990. Practical design criteria for a dynamic ratio imaging system. Cell Calcium 11:93–109

  49. 49.

    Vergara, C., Moczydlowski, E., Latorre, R. 1984. Conduction, blockade and gating in a Ca2+-activated K+ channel incorporated into planar bilayers. Biophys. J. 45:73–76

  50. 50.

    Walters, R.J., Sepúlveda, F.V. 1991. A basolateral K+ conductance modulated by carbachol dominates the membrane potential of small intestinal crypts. Pfuegers Arch. 419:537–539

  51. 51.

    Welsh, M.J., McCann, J.D. 1985. Intracellular calcium regulates basolateral potassium channels in a chloride-secreting epithelium. Proc. Natl. Acad. Sci. USA 82:8823–8826

  52. 52.

    Welsh, M.J., Smith, P.L., Frizzell, R.A. 1983. Chloride secretion by canine tracheal epithelium: III. Membrane resistances and electromotive forces. J. Membrane Biol. 71:209–218

  53. 53.

    Wolosin, J.M., Candia, O.A. 1987. Cl secretagogues increase basolateral K+ conductance of frog corneal epithelium. Am. J. Physiol. 253:C555-C560

  54. 54.

    Wong, S.M.E., Lindeman, R.P., Parangi, S., Chase, H.S., Jr. 1989. Role of calcium in mediating action of carbachol in T84 cells. Am. J. Physiol. 257:C976-C985

  55. 55.

    Wong, S.M.E., Tesfaye, A., DeBell, M.C., Chase, H.S., Jr. 1990. Carbachol increases basolateral K+ conductance in T84 cells. J. Gen. Physiol. 96:1271–1285

Download references

Author information

Additional information

We thank Dr. Marcel Crest (Laboratoire de Neurobiologie, CNRS, Marseille) for providing a sample of kaliotoxin. This work was supported by the Canadian Cystic Fibrosis Foundation and the Respiratory Health Network of Centres of Excellence. J.W.H. is a Chercheur-Boursier of the Fonds de la recherche en santé du Québec.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Tabcharani, J.A., Harris, R.A., Boucher, A. et al. Basolateral K channel activated by carbachol in the epithelial cell line T84 . J. Membarin Biol. 142, 241–254 (1994).

Download citation

Key words

  • T84
  • Inward rectifier
  • KBIC
  • Cystic fibrosis
  • Chloride transport
  • Intestinal secretion