RECEPTOR-BINDING OF GONADOTROPIN-RELEASING-HORMONE ANTAGONISTS THAT INHIBIT RELEASE OF GONADOTROPIN-II AND GROWTH-HORMONE IN GOLDFISH, CARASSIUS-AURATUS

In goldfish, GnRH stimulates gonadotropin-II (GTH-II) and growth hormone (GH) release. The two native forms of GnRH, salmon GnRH (sGnRH) and chicken GnRH-II (cGnRH-II), bind to two classes of GnRH binding sites: high-affinity/low-capacity sites and low-affinity/high-capacity sites. Our previous in vitro perifusion studies of goldfish pituitary fragments showed that [Ac-Delta(3)-Pro(1), 4FD-Phe(2), D-Trp(3,6)]-mGnRH (analog E), [Ac-Delta(3)-Pro(1), 4FD-Phe(2), D-Trp(3,6)]-sGnRH (analog C), and [Ac-D(2)Nal(1), 4Cl-D-Phe(2), D-(3)Pal(3,6)]-cGnRH-II (analog N) inhibited both sGnRH- and cGnRH-II-stimulated GTH-II and GH release. Interestingly, analog C stimulated GH release but not GTH-II release. The objectives of the present study were 1) to test the site of action of GnRH antagonists in goldfish, 2) to test the relationship between receptor binding affinity of antagonists and their in vitro inhibitory potencies and apparent duration of action, and 3) to compare the binding characteristics of analog C with its differential action on GTH-LI and GH release. As in previous studies, analog E suppressed sGnRH-stimulated GTH-II and GH release from perifused pituitary fragments. Similarly, analog E suppressed both sGnRH- and cGnRH-II-stimulated GTH-II and GH release from perifused dispersed goldfish pituitary cells, indicating the direct action of GnRH antagonists at the pituitary cell level. In the receptor binding studies, analog E displaced I-125-[D-Arg(6), Pro(9)NHEt]-sGnRH (sGnRH-A) from crude goldfish pituitary membrane preparations in a dose-dependent manner. The binding affinities to high-affinity GnRH binding sites of analog E, analog C, and analog N were significantly higher than those of native sGnRH and sGnRH-A. The rank order of high affinity was analog N greater than or equal to C > E, and binding affinity had no positive relation with the inhibitory effects of these analogs on GTH-II or GH release in vitro. In conclusion, GnRH antagonists inhibit native GnRH-stimulated GTH-II and GH release by competitively binding to binding sites at the pituitary cells, although there was no positive relation between receptor binding affinity and in vitro GTH-II or GH release-inhibiting potency of the analogs tested.