Extracellular nucleotides stimulate Cl- currents in biliary epithelia through receptor-mediated IP3 and Ca2+ release.
Extracellular ATP regulates bile formation by binding to P2 receptors on cholangiocytes and stimulating transepithelial Cl(-) secretion. However, the specific signaling pathways linking receptor binding to Cl(-) channel activation are not known. Consequently, the aim of these studies in human Mz-Cha-1 biliary cells and normal rat cholangiocyte monolayers was to assess the intracellular pathways responsible for ATP-stimulated increases in intracellular Ca(2+) concentration ([Ca(2+)](i)) and membrane Cl(-) permeability. Exposure of cells to ATP resulted in a rapid increase in [Ca(2+)](i) and activation of membrane Cl(-) currents; both responses were abolished by prior depletion of intracellular Ca(2+). ATP-stimulated Cl(-) currents demonstrated mild outward rectification, reversal at E(Cl(-)), and a single-channel conductance of approximately 17 pS, where E is the equilibrium potential. The conductance response to ATP was inhibited by the Cl(-) channel inhibitors NPPB and DIDS but not the CFTR inhibitor CFTR(inh)-172. Both ATP-stimulated increases in [Ca(2+)](i) and Cl(-) channel activity were inhibited by the P2Y receptor antagonist suramin. The PLC inhibitor U73122 and the inositol 1,4,5-triphosphate (IP3) receptor inhibitor 2-APB both blocked the ATP-stimulated increase in [Ca(2+)](i) and membrane Cl(-) currents. Intracellular dialysis with purified IP3 activated Cl(-) currents with identical properties to those activated by ATP. Exposure of normal rat cholangiocyte monolayers to ATP increased short-circuit currents (I(sc)), reflecting transepithelial secretion. The I(sc) was unaffected by CFTR(inh)-172 but was significantly inhibited by U73122 or 2-APB. In summary, these findings indicate that the apical P2Y-IP3 receptor signaling complex is a dominant pathway mediating biliary epithelial Cl(-) transport and, therefore, may represent a potential target for increasing secretion in the treatment of cholestatic liver disease.
Dutta
Amal K
AK
Department of Pediatrics, University of Texas Southwestern Medical Center, Dallas, TX 75390-9063, USA.
Woo
Kangmee
K
Doctor
R Brian
RB
Fitz
J Gregory
JG
Feranchak
Andrew P
AP
D000230:Adenocarcinoma; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D018118:Chloride Channels; D002712:Chlorides / Q000378:metabolism*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005453:Fluorescence; D005704:Gallbladder / Q000166:cytology; D005706:Gallbladder Neoplasms; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism*; D009711:Nucleotides / Q000494:pharmacology*; D058910:Purinergic P2 Receptor Agonists; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000378:metabolism
D018118:Chloride Channels; D002712:Chlorides; D007295:Inositol Phosphates; D009711:Nucleotides; D058910:Purinergic P2 Receptor Agonists; D018048:Receptors, Purinergic P2; C059370:inositol 1,3,6-triphosphate; D000255:Adenosine Triphosphate; D002118:Calcium
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