Regulation of membrane chloride currents in rat bile duct epithelial cells

J. Gregory Fitz, Srisaila Basavappa, James McGill, Ola Melhus, Jonathan A. Cohn

Research output: Contribution to journalArticle

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Abstract

This study examines the conductive properties of the plasma membrane of cells isolated from the intrahepatic portion of bile ducts. Membrane Cl- conductance was measured in single cells using whole-cell patch clamp recording techniques and in cells in short-term culture using 36Cl and 125I efflux. Separate Ca2+-and cAMP-dependent Cl- currents were identified. Ca2+-dependent Cl- currents showed outward rectification of the current-voltage relation, time-dependent activation at depolarizing potentials, and reversal near the equilibrium potential for Cl-. Ionomycin (2 μM) increased this current from 357±72 pA to 1,192±414 pA (at +80 mV) in 5:7 cells, and stimulated efflux of 125I > 36Cl in 15:15 studies. Ionomycin-stimulated efflux was inhibited by the Cl- channel blocker 4,4′-diisothiocyano-2,2′-stilbene disulfonic acid (DIDS) (150 μM). A separate cAMP-activated Cl- current showed linear current-voltage relations and no time dependence. Forskolin (10 μM) or cpt-cAMP (500 μM) increased this current from 189±50 pA to 784±196 pA (at +80 mV) in 11:16 cells, and stimulated efflux of 36Cl > 125I in 16:16 studies. cAMP-stimulated efflux was unaffected by DIDS. Because the cAMP-stimulated Cl- conductance resembles that associated with cystic fibrosis transmembrane conductance regulator (CFTR), a putative Cl- channel protein, the presence of CFTR in rat liver was examined by immunoblot analyses. CFTR was detected as a 150-165-kD protein in specimens with increased numbers of duct cells. Immunoperoxidase staining confirmed localization of CFTR to bile duct cells but not hepatocytes. These findings suggest that Ca2+- and cAMP-regulated Cl- channels may participate in control of fluid and electrolyte secretion by intrahepatic bile duct epithelial cells, and that the cAMP-regulated conductance is associated with endogenous expression of CFTR. Abnormal ductular secretion may contribute to the pathogenesis of cholestatic liver disease in cystic fibrosis.

Original languageEnglish (US)
Pages (from-to)319-328
Number of pages10
JournalJournal of Clinical Investigation
Volume91
Issue number1
StatePublished - Jan 1993

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Cystic Fibrosis Transmembrane Conductance Regulator
Bile Ducts
Chlorides
Epithelial Cells
Membranes
4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid
Intrahepatic Bile Ducts
Ionomycin
Fluids and Secretions
Stilbenes
Patch-Clamp Techniques
Colforsin
Plasma Cells
Cystic Fibrosis
Electrolytes
Liver Diseases
Hepatocytes
Proteins
Cell Count
Cell Membrane

Keywords

  • Cystic fibrosis
  • Immunohistochemistry
  • Liver
  • Patch clamp
  • Western blot

ASJC Scopus subject areas

  • Medicine(all)

Cite this

Fitz, J. G., Basavappa, S., McGill, J., Melhus, O., & Cohn, J. A. (1993). Regulation of membrane chloride currents in rat bile duct epithelial cells. Journal of Clinical Investigation, 91(1), 319-328.

Regulation of membrane chloride currents in rat bile duct epithelial cells. / Fitz, J. Gregory; Basavappa, Srisaila; McGill, James; Melhus, Ola; Cohn, Jonathan A.

In: Journal of Clinical Investigation, Vol. 91, No. 1, 01.1993, p. 319-328.

Research output: Contribution to journalArticle

Fitz, JG, Basavappa, S, McGill, J, Melhus, O & Cohn, JA 1993, 'Regulation of membrane chloride currents in rat bile duct epithelial cells', Journal of Clinical Investigation, vol. 91, no. 1, pp. 319-328.
Fitz, J. Gregory ; Basavappa, Srisaila ; McGill, James ; Melhus, Ola ; Cohn, Jonathan A. / Regulation of membrane chloride currents in rat bile duct epithelial cells. In: Journal of Clinical Investigation. 1993 ; Vol. 91, No. 1. pp. 319-328.
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AB - This study examines the conductive properties of the plasma membrane of cells isolated from the intrahepatic portion of bile ducts. Membrane Cl- conductance was measured in single cells using whole-cell patch clamp recording techniques and in cells in short-term culture using 36Cl and 125I efflux. Separate Ca2+-and cAMP-dependent Cl- currents were identified. Ca2+-dependent Cl- currents showed outward rectification of the current-voltage relation, time-dependent activation at depolarizing potentials, and reversal near the equilibrium potential for Cl-. Ionomycin (2 μM) increased this current from 357±72 pA to 1,192±414 pA (at +80 mV) in 5:7 cells, and stimulated efflux of 125I > 36Cl in 15:15 studies. Ionomycin-stimulated efflux was inhibited by the Cl- channel blocker 4,4′-diisothiocyano-2,2′-stilbene disulfonic acid (DIDS) (150 μM). A separate cAMP-activated Cl- current showed linear current-voltage relations and no time dependence. Forskolin (10 μM) or cpt-cAMP (500 μM) increased this current from 189±50 pA to 784±196 pA (at +80 mV) in 11:16 cells, and stimulated efflux of 36Cl > 125I in 16:16 studies. cAMP-stimulated efflux was unaffected by DIDS. Because the cAMP-stimulated Cl- conductance resembles that associated with cystic fibrosis transmembrane conductance regulator (CFTR), a putative Cl- channel protein, the presence of CFTR in rat liver was examined by immunoblot analyses. CFTR was detected as a 150-165-kD protein in specimens with increased numbers of duct cells. Immunoperoxidase staining confirmed localization of CFTR to bile duct cells but not hepatocytes. These findings suggest that Ca2+- and cAMP-regulated Cl- channels may participate in control of fluid and electrolyte secretion by intrahepatic bile duct epithelial cells, and that the cAMP-regulated conductance is associated with endogenous expression of CFTR. Abnormal ductular secretion may contribute to the pathogenesis of cholestatic liver disease in cystic fibrosis.

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