Differential regulation of Ca 2+-dependent Cl − currents by FP prostanoid receptor isoforms in Xenopus oocytes

Abstract

The FP A and FP B prostanoid receptor isoforms are G-protein-coupled receptors that are activated by prostaglandin F 2α (PGF 2α). Differences in their carboxyl termini prompted us to examine the intracellular calcium (Ca 2+) signaling of these receptor isoforms using the Xenopus oocyte expression system. Protein expression was determined by immunofluorescence microscopy and whole cell binding with [ 3 H ]PGF 2α. Positive immunolabeling was observed on the outer membranes of oocytes expressing FLAG-tagged FP receptor isoforms, but not on control (water-injected) oocytes. Intracellular signaling was examined using a two-electrode voltage clamp. Specific whole-cell binding was also detected for both receptor isoforms. Bath application of 10 μM PGF 2α to FP A-expressing oocytes produced a chloride (Cl −) current response similar to that of an injection of inositol 1,4,5-trisphosphate (InsP 3) (5.76±0.6 μA, peak current; N=23) that returned to control levels within 25 min. In FP B-expressing oocytes the activation of the Cl − current was delayed or completely absent (1.38±0.2 μA, peak current; N=18). Control oocytes were not responsive to the application of PGF 2α (0.87±0.1 μA, peak current; N=10). Activation of Cl − currents for both FP receptor isoforms was dependent upon intracellular Ca 2+ stores as a 30-min pretreatment with thapsigargin (1 μM; N=5) blocked the PGF 2α induction of the Cl − current. These data indicate that the FP prostanoid receptor isoforms differ in their ability to activate Ca 2+-dependent Cl − channels when expressed in Xenopus oocytes. The difference appears to be in the ability of the two FP prostanoid receptor isoforms to mobilize intracellular calcium.