Differential regulation of Ca(2+)-dependent Cl- currents by FP prostanoid receptor isoforms in Xenopus oocytes.
The FP(A) and FP(B) prostanoid receptor isoforms are G-protein-coupled receptors that are activated by prostaglandin F(2alpha) (PGF(2alpha)). 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 [3H]PGF(2alpha). 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 microM PGF(2alpha) 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 microA, 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 microA, peak current; N=18). Control oocytes were not responsive to the application of PGF(2alpha) (0.87+/-0.1 microA, 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 microM; N=5) blocked the PGF(2alpha) 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.
Anthony
Todd L
TL
Glaucoma Center, Department of Ophthalmology, University of California at San Diego, La Jolla, CA 92037, USA. tanthony@arenapharm.com
Fujino
Hiromichi
H
Pierce
Kristen L
KL
Yool
Andrea J
AJ
Regan
John W
JW
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D018118:Chloride Channels / Q000502:physiology*; D015237:Dinoprost / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D009865:Oocytes / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors; D020033:Protein Isoforms / Q000378:metabolism; D011982:Receptors, Prostaglandin / Q000378:metabolism*; D014982:Xenopus laevis
D018118:Chloride Channels; D004791:Enzyme Inhibitors; D020033:Protein Isoforms; D011982:Receptors, Prostaglandin; C034671:prostaglandin F2alpha receptor; D015237:Dinoprost; D010744:Phosphoric Monoester Hydrolases; D002118:Calcium