Heterotrimeric G-proteins activate Cl^- channels through stimulation of a cyclooxygenase-dependent pathway in a model liver cell line

Circulating hormones produce rapid changes in the Cl^- permeability of liver cells through activation of plasma membrane receptors coupled to heterotrimeric G-proteins. The resulting effects on intracellular pH, membrane potential, and Cl^- content are important contributors to the overall metabolic response. Consequently, the purpose of these studies was to evaluate the mechanisms responsible for G-protein-mediated changes in membrane Cl^- permeability using HTC hepatoma cells as a model. Using patch clamp techniques, intracellular dialysis with 0.3 mM guanosine 5′-O-(3-thiotriphosphate) (GTPγS) increased membrane conductance from 10 to 260 picosiemens/picofarads due to activation of Ca²⁺-dependent Cl^- currents that were outwardly rectifying and exhibited slow activation at depolarizing potentials. These effects were mimicked by intracellular AlF₄^- (0.03 mM) and inhibited by pertussis toxin (PTX), consistent with current activation through Gα_i. Studies using defined agonists and inhibitors indicate that Cl^- channel activation by GTPγS occurs through an indomethacin-sensitive pathway involving sequential activation of phospholipase C, mobilization of Ca²⁺ from inositol 1,4,5-trisphosphate-sensitive stores, and stimulation of phospholipase A₂ and cyclooxygenase (COX). Accordingly, the conductance responses to GTPγS or to intracellular Ca²⁺ were inhibited by COX inhibitors. These results indicate that PTX-sensitive G-proteins regulate the Cl^- permeability of HTC cells through Ca²⁺-dependent stimulation of COX activity. Thus, receptor-mediated activation of Gα_i may be essential for hormonal regulation of liver transport and metabolism through COX-dependent opening of a distinct population of plasma membrane Cl^- channels.