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Am J Physiol Gastrointest Liver Physiol 281: G438-G446, 2001;
0193-1857/01 $5.00
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Vol. 281, Issue 2, G438-G446, August 2001

Clminus -dependent secretory mechanisms in isolated rat bile duct epithelial units

Satish K. Singh1,2, Albert Mennone1, Alessandro Gigliozzi1, Flavia Fraioli1, and James L. Boyer1

1 Liver Center, Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06520-8019; and 2 Evans Biomedical Research Center, Department of Medicine, Boston University School of Medicine, Boston, Massachusetts 02118-2518

Cholangiocytes absorb and secrete fluid, modifying primary canalicular bile. In several Cl--secreting epithelia, Na+-K+-2Cl- cotransport is a basolateral Cl- uptake pathway facilitating apical Cl- secretion. To determine if cholangiocytes possess similar mechanisms independent of CO2/HCO<UP><SUB>3</SUB><SUP>−</SUP></UP>, we assessed Cl--dependent secretion in rat liver isolated polarized bile duct units (IBDUs) by using videomicroscopy. Without CO2/HCO<UP><SUB>3</SUB><SUP>−</SUP></UP>, forskolin (FSK) stimulated secretion entirely dependent on Na+ and Cl- and inhibited by Na+-K+-2Cl- inhibitor bumetanide. Carbonic anhydrase inhibitor ethoxyzolamide had no effect on FSK-stimulated secretion, indicating negligible endogenous CO2/HCO<UP><SUB>3</SUB><SUP>−</SUP></UP> transport. In contrast, FSK-stimulated secretion was inhibited ~85% by K+ channel inhibitor Ba2+ and blocked completely by bumetanide plus Ba2+. IBDU Na+-K+-2Cl- cotransport activity was assessed by recording intracellular pH during NH4Cl exposure. Bumetanide inhibited initial acidification rates due to NH<UP><SUB>4</SUB><SUP>+</SUP></UP> entry in the presence and absence of CO2/HCO<UP><SUB>3</SUB><SUP>−</SUP></UP>. In contrast, when stimulated by FSK, a 35% increase in Na+-K+-2Cl- cotransport activity occurred without CO2/HCO<UP><SUB>3</SUB><SUP>−</SUP></UP>. These data suggest a cellular model of HCO<UP><SUB>3</SUB><SUP>−</SUP></UP>-independent secretion in which Na+-K+-2Cl- cotransport maintains high intracellular Cl- concentration. Intracellular cAMP concentration increases activate basolateral K+ conductance, raises apical Cl- permeability, and causes transcellular Cl- movement into the lumen. Polarized IBDU cholangiocytes are capable of vectorial Cl--dependent fluid secretion independent of HCO<UP><SUB>3</SUB><SUP>−</SUP></UP>. Bumetanide-sensitive Na+-K+-2Cl- cotransport, Cl-/HCO<UP><SUB>3</SUB><SUP>−</SUP></UP> exchange, and Ba2+-sensitive K+ channels are important components of stimulated fluid secretion in intrahepatic bile duct epithelium.

ammonia; cholangiocyte; bumetanide; barium; pH


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