Ion Channels and Cell Signaling in Cell Cultures

  • Valerie Urbach
  • Deirdre Walsh
  • Maria Higgins
  • Isabelle Leguen
  • Christina Doolan
  • John Cuffe
  • Elizabeth Horwitz
  • Catherine Halligan
  • Ruth Gleeson
  • Anthony Cullinane
  • Brian Harvey
Part of the Springer Lab Manual book series (SLM)


Two major diseases, namely cancer and cystic fibrosis (CF), have driven the development of immortalized human airway epithelial cell lines. Primary cultures of airway epithelial cells have also played a central role in the study of other diseases such as airway inflammation and viral infection. However, primary lung culture is limited by the number of viable cells that can be generated from the tissue and by the availability of fresh tissue. Immortalized cell lines of airway epithelium were developed from lung carcinomas or from primary cultures transformed in vitro using viruses (SV40). The development of cell lines has greatly enhanced our understanding of the biochemical and proliferation properties of lung epithelium because these characteristics are not affected by viral transformation. However, many transformed cells lose certain differentiated functions, such as tight junction formation or physiological ionic transport properties. For this reason, most of the knowledge of ionic transport mechanisms and regulation come from studies on primary cell culture.


Cystic Fibrosis Cystic Fibrosis Transmembrane Conductance Regulator Airway Epithelial Cell Airway Epithelium Cortical Collect Duct 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


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  1. Ahmad I, Korbmacher C, Segal AS, Cheung P, Boulpaep EL, Barnstable CJ (1992) Mouse cortical collecting duct cells show nonselective cation channel activity and express a gene related to the cGMP-gated rod photoreceptor channel. Proc Natl Acad Sci USA 89:10262–10266PubMedCrossRefGoogle Scholar
  2. Anderson MP, Sheppard DN, Berger HA, Welsh MJ (1992) Chloride channels in the apical membrane of normal and cystic fibrosis airway and intestinal epithelia. Am J Physiol 263:L1–14PubMedGoogle Scholar
  3. Barrett KE (1993) Positive and negative regulation of Cl-secretion in colonic cells. Am J Physiol 265:C859–C868PubMedGoogle Scholar
  4. Benos DJ, Cunningham S, Baker RR, Beason KB Oh Y, Smith PR (1992) Molecular caracteristics of amiloride-sensitive sodium channels. Rev Physiol Biochem Pharmacol 120:32–113Google Scholar
  5. Brayden DJ, Krouse ME, Law T, Wine JJ (1993) Stilbenes stimulate colonic Cl-secretion by elevating Ca2+. Am J Physiol 264:G325–G333PubMedGoogle Scholar
  6. Brochiero E, Raschi C, Ehrenfeld J (1995) Na/Ca exchange in the basolateral membrane of A6 cell monolayer: role in ca homeostasis. Pflügers Arch 430:105–114PubMedCrossRefGoogle Scholar
  7. Boucher RC (1994) Human airway ion transport. Am J Resp Crit Care Med 150:271–281PubMedGoogle Scholar
  8. Buchanan JA, Yeger H,Tabcharani JA, Jensen T J, Auerbach W, Hanrahan JW, Riodan JR, Bucwald M (1990) Transformed sweat gland and nasal epithelial cell lines from control and CF individuals. J Cell Sci 109–123Google Scholar
  9. Candia O, Mia AJ, Yorio T (1993) Influence of filter supports on transport characteristics of cultured A6 kidney cells. Am J Physiol 265:0479–88Google Scholar
  10. Cantiello HF, Stow JL, Prat AG and Ausiello DA (1991) Actin filaments regulate epithelial Na channel activity. Am J Physiol 261:C882–C888PubMedGoogle Scholar
  11. Casavola-V, Guerra-L, Helmle-Kolb-C, Reshkin-SJ, Murer-H (1992) Na+/H(+)-exchange in A6 cells: polarity and vasopressin regulation. J Membr Biol 130:105–114PubMedGoogle Scholar
  12. Coca-Prados M, Chatt G (1986) Growth of nonpigmented ciliary epithelial cells in serum free hormone supplemented media. Exp Eye Res 43:617–620PubMedCrossRefGoogle Scholar
  13. Doolan CM, Harvey BJ (1996a) Modulation of cytosolic protein kinase C and calcium ion activity by steroid hormones in rat distal colon. J Biol Chem 271:8763–8767PubMedCrossRefGoogle Scholar
  14. Doi Y, Marunaka Y (1995) Antidiuretic hormone decreases the intracellular pH in distal nephron epithelium (A6) by inhibiting Na+/H+ exchange. Gen Pharmacol 26:723–726PubMedCrossRefGoogle Scholar
  15. Doolan CM, Harvey BJ (1996b) Rapid effects of steroid hormones on free intracellular calcium in T84 colonic epithelial cells. Am J Physiol 271:(Cell Physiol 40) C1935–C1941PubMedGoogle Scholar
  16. Coca-Prados M, Wax MB (1986) Transformation of Human ciliary epithelial cells by simion virus 40: induction of cell proliferation and retention of b2 -adrenergic receptors. Proc Natl Acad Sci USA 83: 8754–8760PubMedCrossRefGoogle Scholar
  17. Cott GR and Rao Hydrocortisone promotes the maturation of na dependent ion transport across the fetal pulmonry epithelium. Am J Respir Cell Mol Biol 9:166–171 1993PubMedGoogle Scholar
  18. Cotton CU, Stutts MJ, Knowles MR, Gatzy JT, Boucher RC (1987) Abnormal apical cell mebrane in cystic fibrosis respiratory epithelium. J Clin Invest 79:80–85PubMedCrossRefGoogle Scholar
  19. Coleman D L, Tuet IK, Widdicombe JH (1984) Electrical properties of dog tracheal epithelial cells grown in monolayer culture. Am J Physiol 245:C355–C359Google Scholar
  20. Cozens AL, Yezzi MJ, Kunzelman K, Ohrui T, Chin L, Eng K, Finbeiner WE, Widdicombe JH, Gruenert DC (1994) CFTR expression and chloride secretion in polarized immortal human bronchial epithelial cells. Am J Respir Cell Mol Biol Vol 10:38–47Google Scholar
  21. De Jong PM, Van Sterkenburg MA, Kempenaar JA, Dijkman JH, Ponec M (1993) Serial culturing of human bronchial epithelial cells derived from biopsies. In Vitro Cell Dev Biol 219A:379–387CrossRefGoogle Scholar
  22. Denning G M, Anderson M P, Amara J f, Marshall J, Smith AE, Welsh MJ (1992) Processing of mutant cystic fibrosis transmembrane conductance regulator is temperature-sensitive. Nature 358:761–764PubMedCrossRefGoogle Scholar
  23. Field M , Semrad CE (1993) Toxigenic diarrheas, congenital diarrheas, and cystic fibrosis: Disorders of intestinal ion transport. Annu Rev Physiol 55:631–655PubMedCrossRefGoogle Scholar
  24. Graham A, Steel DM, Alton EW, Alton FW, Geddes DM (1992) Second messenger regulation of sodium transport in mammalian airway epithelia. J Physiol 453:475–491PubMedGoogle Scholar
  25. Gruenert DC (1987) Differentiated properties of human epithelial cells transformed in vitro. Biotechniques 5(8):740–749Google Scholar
  26. Gruenert DC, Finkbeiner WE, Widicombe JH (1995) Cultured and transformation of human airway epithelial cells. Am J Physiol 268:L347–L360PubMedGoogle Scholar
  27. Gstraunthaler GJA (1988) Epithelial cells in tissue culture. Renal Physiol Biochem 11:1–42PubMedGoogle Scholar
  28. Hall AC, Horwitz ER, Wilkins RJ (1996) The cellular physiology of articular cartilage. J Exp Physiol 81:535–545Google Scholar
  29. Halm DH, Rechkemmer GR, Schoumacher RA, Frizzell RA (1988) Apical membrane chloride channels in a colonic cell line activated by secretory agonists. Am J Physiol 254:C505–C511PubMedGoogle Scholar
  30. Harvey BJ, Urbach V (1995) Regulation of ion and water transport by hydrogen ions in high resistance epithelia. In: Heisler N (ed) Advances in comparative and environmental physiology, vol 22. Springer, Berlin Heidelberg New York, pp 153–183Google Scholar
  31. Harvey BJ, Urbach V, Van Kerkhove E (1991) Inward rectifier K channels in principal cells of isolated frog skin epithelium and in cultured A6 cells. J Physiol 438:264PGoogle Scholar
  32. Haws C, Krouse ME, Xia Y, Gruenert DC, Wine JJ (1992) CFTR channels in immortalised human airway cells. Am J Physiol 263:L692–L707PubMedGoogle Scholar
  33. Hoffmann EK, Duham PB (1995) Membrane mechanisms and intracellular signalling in cell volume regulation. Int Rev Cytol 161:173–262PubMedCrossRefGoogle Scholar
  34. Horwitz ER, Higgins T, Harvey BJ (1996) Histamine induced cytosolic calcium increase in porcine articular chondrocytes. Biochim Biophys Acta 1313:95–100PubMedCrossRefGoogle Scholar
  35. Keely SJ, Stack WA, O’Donoghue DP, Baird AW (1995) Regulation of ion transport by histamine in human colon. Eur J Pharm 279:203–209CrossRefGoogle Scholar
  36. Kilburn KH (1968) A hypothesis for pulmonary clearance and its implications. Am Rev Respir Dis 98:449–463PubMedGoogle Scholar
  37. Knowles MR, Gatzy JT, Boucher RC (1985) Aldosterone metabolism and transepithe-lial potential difference in normal and cystic fibrosis subjects. Pediatr Res 19:676–679PubMedCrossRefGoogle Scholar
  38. Koefed-Johnsen V, Ussing HH (1958) The nature of frog skin potential. Acta Physiol Scand 42:293–308CrossRefGoogle Scholar
  39. Korbmacher C, Segal AS, Fejes-Toth G, Giebisch G, Boulpaep EL (1993) Whole-cell currents in single and confluent M-l mouse cortical collecting duct cells. J Gen Physiol 102:761–793PubMedCrossRefGoogle Scholar
  40. Kuchel OG, Kuchel GA (1991) Peripheral dopamine in pathophysiology of hypertension: Interaction with aging and lifestyle. Hypertension 18:709–721PubMedGoogle Scholar
  41. Kunzelmann K, Kathofer S, Greger R (1995) Na and Cl conductances in airway epithelial cells: increased Na conductance in cystic fibrosis. Pflugers Arch 431:1–9PubMedCrossRefGoogle Scholar
  42. Kunzelmann K, Kathofer S, Hipper A, Gruenert DC, Greger R (1996) Culture -dependent expression of Na+ conductances in airway epithelial cells. Pflugers Arch 431:578–586PubMedCrossRefGoogle Scholar
  43. Kunzelmann K, Schwiebert EM, Zeitlin PL, Kuo WL, Stanton BA, Gruenert DC (1993) An immortalized CF tracheal cell line homozygous for the AF 508 CFTR mutation. Am J Respir Cell Mol Biol 8:522–529PubMedGoogle Scholar
  44. Lechner JF, LaVeck MA (1985) A serum-free method for culturing normal human bronchial epithelial cells at clonal density. J Tissue Culture Methods 9:43–48CrossRefGoogle Scholar
  45. Leguen I, Urbach V, Harvey BJ (1995) Cytochalasin effect on mechanosensitive calcium chgannel activity and cell swelling-induced calcium mobilisation in renal A6 cells. 489:92PGoogle Scholar
  46. Letz B, Ackermann A, Canessa CM, Rossier BC, Korbmacher C (1995) Amiloride-sensitive sodium channels in confluent M-1 mouse cortical collecting duct cells. J Membr Biol 148:127–141PubMedGoogle Scholar
  47. Lewis SA, Eaton, DC, Clausen C, Diamond JM (1977) Nystatin as a probe for investigating the electrical properties of tight epithelium. J Gen Physiol 70:427–440PubMedCrossRefGoogle Scholar
  48. Lin M , Nairn AC, Guggino SE (1992) cGMP-dependent protein kinase regulation of a chloride channel in colonic cells. Am J Physiol 262:C1304–C1312PubMedGoogle Scholar
  49. Ling BN and Eaton DC (1989) Effect of luminal Na+ on single Na channels, a regulatory role for protein kinase C. Am J Physiol 256:F1094–F1103PubMedGoogle Scholar
  50. Maguire D, O’ Sullivan G, Harvey BJ (1994) Potassium ion channels and sodium absorption in human colon. Surgical Forum 80:195–197Google Scholar
  51. Maguire D , O’ Sullivan G, Harvey BJ (1995) Membrane and genomic mechanisms for aldosterone effect in human colon. Surgical Forum 81:203–205Google Scholar
  52. Mall M, Hipper A, Greger R, Kunzelman K (1996) Wild type but not AF 508 inhibits Na conductance when coexpressed in Xenopus oocytes. FEBS Letters 381:47–52PubMedCrossRefGoogle Scholar
  53. Malone DG, Irani AM, Schwartz B, Barrett KE, Metcalfe DD (1986) Mast cell numbers and histamine levels in synovial fluids from patients with diverse arthritides. Arthritis Rheum 29:944–955CrossRefGoogle Scholar
  54. Mason SJ, Paradiso AM, Boucher RC (1991) Regulation of transepitehlial ion transport and intracellular calcium by extracellular adenosine triphosphate in human normal and cystic fibrosis airway epithelium Br J Pharmacol 103:1649–1656PubMedGoogle Scholar
  55. Middleton JP, Mangel AW, Basavappa S, Fitz JG (1993) Nucleotide receptors regulate ion transport in renal epithelial cells. Am J Physiol 264:F867–F873PubMedGoogle Scholar
  56. Murphy E, Cheng E, Yankaskas J, Stutts MJ, Boucher RC (1988) Cell calcium levels of normal and cystic fibrosis nasal epithelium. Pediatr Res 24:79–84PubMedCrossRefGoogle Scholar
  57. Nilius B, Sehrer J, Heinke S, Droogmans G (1995) Ca2+ release and activation of K+ and Cl-currents by extracellular ATP in distal nephron epithelial cells. Am J Physiol 269:C376–C384PubMedGoogle Scholar
  58. Paradiso AM (1992) Identification of Na/H exchange in human normal and cystic fibrosis ciliated airway epithelium. Am J Physiol 262:L757–L764PubMedGoogle Scholar
  59. Paradiso AM, Cheng EHC, Boucher RC (1991) Effects of bradykinin on intracellular calcium regulation in human ciliated airway epithelium. Am J Physiol 261:L63–L69PubMedGoogle Scholar
  60. Rafferty KA (1975) Epithelial cells grown in culture of normal and neoplastic forms. Adv Cancer Res 21:249–272PubMedCrossRefGoogle Scholar
  61. Reenestra WW (1993) Inhibition of cAMP-and Ca2+-dependent Cl-secretion by phorbol esters: Inhibition of basolateral K+ channels. Am J Physiol 264:C161–C168Google Scholar
  62. Rich DP, Anderson MP, Gregory RJ, Cheng SH, Paul S, Jefferson DM, McCann JD, Klinger KW, Smith AE and Welsh MJ (1990) Expression of cystic fibrosis transmembranaire conductance regulator corrects defective chloride channel regulation in cystic fibrosis airway epithelial cells. Nature 347:358–363PubMedCrossRefGoogle Scholar
  63. Rodriguez-Commes J, Isales C, Kalghati L, Gasalla-Herraiz J, Hayslett JP (1994) Mechanism of insulin-stimulated electrogenic sodium transport. Kidney Int 46:666–674PubMedCrossRefGoogle Scholar
  64. Rouse D, Leite M, Suki WN (1994) ATP inhibits the hydrosmotic effect of AVP in rabbit CCT: evidence for a nucleotide P2u receptor. Am J Physiol 267:F289–F295PubMedGoogle Scholar
  65. Schafer JA, Troutman SL (1990) cAMP mediates the increase in apical membrane Na+ conductance produced in rat CCD by vasopressin. Am J Physiol 259:F823–F831PubMedGoogle Scholar
  66. Schwiebert EM, Egan ME, Hwang TH, Fulmer SB, Allen SS, Cutting GR, Guggino WB (1995) CFTR regulates ORCC thriugh an autorine mechanism inolving ATP. Cell: 1063–1073Google Scholar
  67. Scolte BJ, Kansen M, Hoogeveen AT, Willemse R, Rhim J S, Vander Kamp AWM, Bijman J (1991) Immortalization of nasal polyp epithelial cells from cystic fibrosis patients. Exp Cell Res 182:559–571CrossRefGoogle Scholar
  68. Stoos BA, Naray-Fejes-Toth A, Carretero OA, Ito S, Fejes-Toth G (1991) Characterisation of a mouse cortical collecting duct cell line. Kidney Int 39:1168–1175PubMedCrossRefGoogle Scholar
  69. Stutts MJ, Knowles MR, Gatzy JT, Boucher RC (1986) Oxygen consumption and ouabain binding sites in cystic fibrosis nasal epithelium. Pediatr Res 20:1316–1320PubMedCrossRefGoogle Scholar
  70. Stutts MJ, Chinet TC, Mason SJ, Fulton JM, Clarke LL, Boucher RC (1992) Regulation of chloride channels in normal and cystic fibrosis airway epithelial cells by extracellular ATP. Proc Natl Acad Sci USA 89:1621–1625PubMedCrossRefGoogle Scholar
  71. Stutts MJ, Canessa Cecilia, Olsen JC, Hamrick M, Cohn JA, Rossier BC, Boucher RC (1995) CFTR as a cAMP-dependent regulator of Na channels. Sciences 269:847–851CrossRefGoogle Scholar
  72. Taylor DJ, Yoffe JR, Brown DM, Woolley DE (1985) Histamine H2 receptors on chondrocytes derived from human, canine and bovine articular cartilage. Biochem J:315–319Google Scholar
  73. Taylor DJ, Yoffe JR, Brown DM, Woolley DE (1986) Evidence for both histamine Hl and H2 receptors on human articular chondrocytes. Arthritis Rheum 29:160–165PubMedCrossRefGoogle Scholar
  74. Taylor DJ, Woolley DE (1987) Histamine stimulates prostaglandin E production by rheumatoid synovial cells and human articular chondrocytes in culture. Ann Rheum Dis 46:431–435PubMedCrossRefGoogle Scholar
  75. Urbach V Prosser E, Raffin JP, Thomas S, Harvey B (1994) Activation of Cl channel by extracellular UTP in human lung epithelium. J Gen Physiol 104:84Google Scholar
  76. Van Scott MR, Yankaskas, Boucher RC (1986) Culture of airway epithelial cells: research techniques. Exp Lung Res 11:75–94PubMedCrossRefGoogle Scholar
  77. Walsh D, Urbach V, Harvey B (1996) Nucleotide regulation of intracellular calcium and ion channel activity in human normal and cystic fibrosis lung. The respiratory system in health and disease. Welcome Trust Meeting. March 1, 1996Google Scholar
  78. Ward CL, Krouse ME, Gruenert DC, Kopito RR, Wine JJ (1991) Cystic fibrosis gene expression is not correlated with rectifying Cl-channels. PNAS 88:5277–5281PubMedCrossRefGoogle Scholar
  79. Widdicombe JH, Welsh MJ, Finkbeiner WE (1985) Cystic fibrosis decreases the apical membrane chloride permeability of monolayers cultured from cells tracheal epithelium. Proc Natl Acad Sci USA 82:6167–6171PubMedCrossRefGoogle Scholar
  80. Willumsen NJ, Boucher RC (1992) Intracellular pH and its relationschip to regulation of ion transport in normal and cystic fibrosis human nasal epithelia. J Physiol 455:247–269PubMedGoogle Scholar
  81. Willumsen NJ, Davis CW, Boucher RC (1989) Cellular Cl transport in cultured cystic fibrosis airway epithelium Am J Physiol 256:C1045–C1053PubMedGoogle Scholar
  82. Wu R, Nolan E, Turner C (1985) Expression of trchelal differenciation in serum-free, hormone-supplemented medium. J Cell Physiol 127:167–181CrossRefGoogle Scholar
  83. Yanase M, Handler JS (1986) Activators of protein kinase C inhibit sodium transport in A6 epithelia. Am J Physiol 250:C517–C523PubMedGoogle Scholar
  84. Yankaskas JR, Cotton CU Knowles MR, Gatzy JT, Boucher RC (1985) Culture of human nasal epithelial cells on collagen matrix supports. Am Rev Respir Dis 132:1281–1287PubMedGoogle Scholar
  85. Yue G, Hu P, Oh Y, Jilling T, Shoemaker R L, Benos D J, Cragoe E J, Matalon S (1993) Culture-induced alterations in alveolar type II cell Na conductance. Am J Physiol 265:C630–C 640PubMedGoogle Scholar
  86. Urbach V, Andersen H, Hall J, Harvey BJ (1993) A calcium permeable cation channel activated by membrane stretch and cell swelling in renal A6 and frog skin epithelia. J Physiol 467:273PGoogle Scholar
  87. Urbach V, Leguen I, Harvey BJ (1995) Volume and mechano-sensitive calcium entry and release in renal A6 cells. J Physiol 489:87PGoogle Scholar
  88. Urban JPG, Hall AC (1992) In: Kuettner K et al. (ed) Articular cartilage and osteoarthritis. Raven, New York, pp 393–406Google Scholar
  89. Verrey F (1994) Antidiuretic hormone action in A6 cells: effect on apical Cl and Na conductances and synergism with aldosterone for NaCl reabsorption. J Membr Biol 138:65–76PubMedGoogle Scholar
  90. Weynier A, Huott PA, Liu W, McRoberts JA Dharmasthaphorn K (1985) Chloride secretory mechanism induced by prostaglandin E1 in a colonic epithelial cell line. J Clin Invest 76:1828–1836CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1998

Authors and Affiliations

  • Valerie Urbach
    • 1
  • Deirdre Walsh
  • Maria Higgins
  • Isabelle Leguen
  • Christina Doolan
  • John Cuffe
  • Elizabeth Horwitz
  • Catherine Halligan
  • Ruth Gleeson
  • Anthony Cullinane
  • Brian Harvey
  1. 1.Wellcome Trust Cellular Physiology Research Unit, Department of PhysiologyUniversity CollegeCorkIreland

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