Chloride channels mediate the response to gonadotropin-releasing hormone (GnRH) in Xenopus oocytes injected with rat anterior pituitary mRNA.
Functional expression of receptors for GnRH was studied using Xenopus laevis oocytes injected with poly(A)+ mRNA extracted from rat anterior pituitary glands. Whole-cell currents were monitored using two-electrode voltage-clamp techniques. In oocytes which responded to both GnRH and TRH, the GnRH response showed a longer latency and time-to-peak than the TRH response. The response to GnRH or an agonist of GnRH receptors, buserelin (1 nM-1 microM) consisted of current fluctuations and occurred in a dose-dependent manner. This GnRH response was blocked by the Cl- channel blockers 9-AC (9-anthracene carboxylic acid; 1 mM), 4,4'-diisothiocyanastilbene-2,2'-disulfonic acid (0.1 mM), and diphenylamine-2-carboxylic acid (0.1 mM). The reversal potential for the GnRH-induced current fluctuations was -25 mV, comparable with the reported Cl- equilibrium potential in Xenopus oocytes, and its shift, when the external concentration of Cl- was changed, was reasonably described by the Nernst equation. These results indicate that the GnRH-induced response was dependent on the activity of Cl- channels. Ca2+ also plays a role, as the GnRH-induced response was reversibly suppressed by a calmodulin inhibitor, chlordiazepoxide (0.2 microM), and by a blocker of intracellular Ca2+ release, TMB-8 (8-(N.N-diethylamino) octyl-3,4,5-trimethoxybenzoate; 0.1-0.2 mM). It is concluded that GnRH (and TRH) receptors, expressed in Xenopus oocytes by injecting exogenous mRNA from rat anterior pituitary glands, operate via activation of Ca2+-dependent Cl- channels.
Yoshida
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MRC Reproductive Biology Unit, Centre for Reproductive Biology, Edinburgh, United Kingdom.
Plant
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Taylor
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Eidne
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D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D018118:Chloride Channels; D002712:Chlorides / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008845:Microinjections; D009865:Oocytes; D010903:Pituitary Gland, Anterior; D010906:Pituitary Hormone-Releasing Hormones / Q000494:pharmacology*; D012333:RNA, Messenger; D011967:Receptors, Gonadotropin / Q000187:drug effects / Q000235:genetics; D014982:Xenopus laevis; D062367:ortho-Aminobenzoates / Q000494:pharmacology
D000873:Anthracenes; D018118:Chloride Channels; D002712:Chlorides; D008565:Membrane Proteins; D010906:Pituitary Hormone-Releasing Hormones; D012333:RNA, Messenger; D011967:Receptors, Gonadotropin; D062367:ortho-Aminobenzoates; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; C010030:9-anthroic acid; C001209:fenamic acid; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D002118:Calcium