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Methods for the study of ionic currents and Ca2+-signals in isolated colonic crypts

Abstract

Isolated epithelial cells from intestinal mucosae are a suitable object for the study of the regulation of ion transport in the gut. This regulation possesses a great importance for human and veterinary medicine, as diarrheal diseases, which often are caused by an inadequate activation of intestinal anion secretion, are one of the major lethal diseases of children or young animals. The aim of this paper is to describe a method for the isolation of intact colonic crypts, e.g. for the subsequent investigation of the regulation of anion secretion by the intracellular second messenger, Ca2+ using electrophysiological and imaging techniques.

References

  1. 1.

    Binder HJ, Sandle GJ. Electrolyte transport in the mammalian colon. In: Johnson LR (ed) Physiology of the gastrointestinal tract. 3rd ed. New York: Raven Press; 1994. p. 2133–2171.

    Google Scholar 

  2. 2.

    Diener M, Eglème C, Rummel W. Phospholipase C-induced anion secretion and its interaction with carbachol in the rat colonic mucosa. Eur J Pharmacol 1991;200:267–276.

    PubMed  Article  CAS  Google Scholar 

  3. 3.

    Raimondi F, Kao JPY, Kaper JB, Guandalini S, Fasano A. Calcium-dependent intestinal chloride secretion by vibrio parahaemolyticus thermostable direct hemolysin in a rabbit model. Gastroenterology 1995;109:381–386.

    PubMed  Article  CAS  Google Scholar 

  4. 4.

    Böhme M, Diener M, Rummel W. Calcium- and cyclic-AMP-mediated secretory responses in isolated colonic crypts. Pflügers Arch 1991;419:144–151.

    PubMed  Article  Google Scholar 

  5. 5.

    Bleich M, Riedemann N, Warth R, Kerstan D, Leipziger J, Hör M, Van Driessche W, Greger R. Ca2+ regulated K+ and nonselective cation channels in the basolateral membrane of rat colonic crypt base cells. Pflügers Arch 1996;432:1011–1022.

    PubMed  Article  CAS  Google Scholar 

  6. 6.

    Del Castillo JR. The use of hyperosmolar, intracellular-like solutions for the isolation of epithelial cells from guinea-pig small intestine. Biochim Biophys Acta 1987;901:201–208.

    PubMed  Article  Google Scholar 

  7. 7.

    Strabel D., Diener M. Evidence against direct activation of chloride secretion by carbachol in the rat distal colon. Eur J Pharmacol 1995;274:181–191.

    PubMed  Article  CAS  Google Scholar 

  8. 8.

    Siemer C, Gögelein H. Activation of nonselective basolateral cation channels in the basolateral membrane of rat distal colon crypt cells. Pflügers Arch 1992;420:319–328.

    PubMed  Article  CAS  Google Scholar 

  9. 9.

    Warth R, Hamm K, Bleich M, Kunzelmann K, von Hahn T, Schreiber R, Ullrich E, Mengel M, Trautmann N, Kindle P, Schwab A, Greger R. Molecular and functional characterization of the small Ca2+-regulated K+ channel (rSK4) of colonic crypts. Pflügers Arch 1999;438:437–444.

    PubMed  Article  CAS  Google Scholar 

  10. 10.

    Lindström CG, Rosengren JE, Fork FT. Colon of the rat. An anatomic, histologic and radiographic investigation. Acta Radiol Diagn 1979;20:523–536.

    Google Scholar 

  11. 11.

    Parsons DS, Paterson CR. Fluid and solute transport across rat colonic mucosa. Q J Exp Physiol 1965;50:220–231.

    CAS  Google Scholar 

  12. 12.

    Frings M, Schultheiss G, Diener M. Electrogenic Ca2+ entry in the rat colonic epithelium. Pflügers Arch 1999;439:39–48.

    PubMed  Article  CAS  Google Scholar 

  13. 13.

    Grynkiewicz G, Poenie M, Tsien RY. A new generation of Ca2+ indicators with improved fluorescence properties. J Biol Chem 1985;260:3440–3450.

    PubMed  CAS  Google Scholar 

  14. 14.

    Hartmann F, Owen R, Bissell DM. Characterization of isolated epithelial cells from rat small intestine. Am J Physiol 1985;242:G147-G155.

    Google Scholar 

  15. 15.

    Diener M, Rummel W, Mestres P, Lindemann B. Single chloride channels in colon mucosa and isolated colonic enterocytes of the rat. J Membr Biol 1989;108:21–30.

    PubMed  Article  CAS  Google Scholar 

  16. 16.

    Ziomek CA, Schulman S, Edidin M. Redistribution of membrane proteins in isolated intestinal epithelial cells. J Cell Biol 1980;86:849–857.

    PubMed  Article  CAS  Google Scholar 

  17. 17.

    Bjorkman DJ, Allan CH, Hagen SJ, Trier J. Structural features of absorptive cell and microvillus preparations from rat small intestine. Gastroenterology 1986;91:1401–1414.

    PubMed  CAS  Google Scholar 

  18. 18.

    Diener M, Nobles M, Rummel W. Activation of basolateral Cl channels in the rat colonic epithelium during regulatory volume decrease. Pflügers Arch 1992;421:530–538.

    PubMed  Article  CAS  Google Scholar 

  19. 19.

    Lipkin M. Proliferation and differentiation of normal and diseased gastrointestinal cells. In: Johnson LR (ed) Physiology of the gastrointestinal tract. 2nd ed. New York: Raven press; 1994. p. 255–284.

    Google Scholar 

  20. 20.

    Schultheiss G, Ribeiro R, Diener M. Fatty acids inhibit anion secretion in rat colon: apical and basolateral action sites. Pflügers Arch 2001;442:603–613.

    PubMed  Article  CAS  Google Scholar 

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Correspondence to Martin Diener.

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Published: April 8, 2002

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Schultheiss, G., Kocks, S.L. & Diener, M. Methods for the study of ionic currents and Ca2+-signals in isolated colonic crypts. Biol Proced Online 3, 70–78 (2001). https://doi.org/10.1251/bpo25

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Indexing terms

  • Electrophysiology
  • Epithelial Cells
  • Ion channels
  • Patch clamp