REGULATION OF BILIARY-SECRETION THROUGH APICAL PURINERGIC RECEPTORS IN CULTURED RAT CHOLANGIOCYTES

To evaluate whether ATP in bile serves as a signaling factor regulatin g ductular secretion, voltage-clamp studies were performed using a nov el normal rat cholangiocyte (NRC) model. In the presence of amiloride (100 mu M) to block Na+ channels, exposure of the apical membrane to A TP significantly increased the short-circuit current (I-sc) from 18.2 +/- 5.9 to 52.8 +/- 12.7 mu A (n = 18). The response to ATP is mediate d by basolateral-to-apical Cl- transport because it is inhibited by 1) the Cl- channel blockers 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (1 mM), diphenylanthranilic acid (1.5 mM), or 5-nitro-2-(3-phenyl propylamino)benzoic acid (50 or 100 mu M) in the apical chamber, 2) th e K+ channel blocker Ba2+ (5 mM), or 3) the Na+-K+-2Cl(-) cotransport inhibitor bumetanide (200 mu M) in the basolateral chamber. Other nucl eotides stimulated an increase in Isc with a rank order potency of UTP = ATP = adenosine 5'-O-(3)-thiotriphosphate, consistent with P-2u pur inergic receptors. ADP, AMP, 2-methylthioadenosine 5'-triphosphate, an d adenosine had no effect. A cDNA encoding a rat P-2u receptor (rP(2u) R) was isolated from a liver cDNA library, and functional expression o f the corresponding mRNA in Xenopus laevis oocytes resulted in the app earance of ATP-stimulated currents with a similar pharmacological prof ile. Northern analysis identified hybridizing mRNA transcripts in NRC as well as other cell types in rat liver. These findings indicate that exposure of polarized cholangiocytes to ATP results in luminal Cl- se cretion through activation of P-2u receptors in the apical membrane. R elease of ATP into bile may serve as an autocrine or paracrine signal regulating cholangiocyte secretory function.