BACKGROUND/AIMS: The biliary epithelium contributes to bile formation through absorption and secretion of fluid and electrolytes. The effects of extracellular nucleotides on membrane ion transport were assessed in isolated bile duct cells from rats and Mz-ChA-1 cells from a human cholangiocarcinoma. METHODS: The rates of efflux of 125I and 86Rb were used to assess membrane Cl- and K+ permeabilities, respectively. Patch clamp recordings of whole cell currents were used to evaluate the properties of adenosine triphosphate (ATP)-activated currents. RESULTS: Purinergic receptor agonists ATP and uridine triphosphate stimulated 125I and 86Rb efflux about twofold above basal levels. The effects were reproduced by a nonhydrolyzable analogue of ATP (adenosine 5'-O-[3-thiophosphate]) and were unaffected by an adenosine receptor blocker xanthine amine congener. 125I efflux was also stimulated by adenosine and its receptor agonists 5'-N-ethylcarboxamidoadenosine, N6-(2-phenylisopropyl)adenosine; these effects were inhibited by xanthine amine congener, suggesting a separate adenosine receptor. ATP, adenosine 5'-O-(3-thiophosphate), and uridine triphosphate each stimulated release of Ca2+ from intracellular stores, whereas adenosine had no effect. In whole cell recordings of Mz-ChA-1 cells, ATP activated an early transient outward current consistent with a K+ conductance and a later, sustained inward current consistent with a Cl- conductance. CONCLUSIONS: Biliary cells possess at least two classes of nucleotide receptors that modulate membrane ion permeability through Ca(2+)-dependent and -independent pathways, and ATP may be involved in the regulation of biliary secretion.