PRESENCE OF GONADOTROPIN-RELEASING-HORMONE (GNRH) BINDING-SITES AND COMPOUNDS WITH GNRH-LIKE ACTIVITY IN THE OVARY OF AFRICAN CATFISH, CLARIAS-GARIEPINUS

GnRH binding was characterized in the African catfish ovary by use of an analog of salmon GnRH (sGnRH-A; [D-Arg(6), Trp(7), Leu(8), Pro(9)-NEt]GnRH) as a labeled ligand. Binding of sGnRH-A to catfish ovarian membrane preparation was found to be saturable, displaceable, reversible, and dependent on time, temperature, and tissue concentration. Optimal binding was achieved after 70 min of incubation at room temperature (similar to 22 degrees C) at pH 7.6. Addition of unlabeled sGnRH-A displaced the bound I-125-sGnRH-A in a dose-related manner. Hill plot as well as Scatchard analysis indicated the presence of one class of high-affinity binding sites with a equilibrium dissociation constant (K-d) of 0.27 +/- 0.036 nM. Bound I-125-sGnRH-A was also found to be displaceable by catfish GnRH (cfGnRH; [His(5), Leu(7), Asn(8)]-GnRH), chicken GnRH-II (cGnRH-II; [His(5), Trp(7), Tyr(8)]-GnRH), and salmon GnRH (sGnRH; [Trp(7), Leu(8)]-GnRH); all the peptides were found to bind with lower affinities than sGnRH-A to the catfish ovarian GnRH binding sites. Further experiments using ovarian extracts indicated the presence of compounds with GnRH-like activity in the ovary of African catfish. The crude ovarian extract was found to stimulate pituitary gonadotropin release h om goldfish pituitary, as well as displacing I-125-sGnRN-A binding in the catfish ovary. HPLC analysis of the catfish ovarian extract revealed the presence of two fractions that bind specifically to the catfish ovary and release gonadotropin from cultured goldfish pituitary. These fractions include an early eluting peak that does not correspond with the retention time of known GnRH forms in addition to a fraction that co-elutes with the mammalian GnRH. Overall, the study provided characterization of GnRH binding sites in the catfish ovary, and evidence for the presence of compounds with GnRH-like activity in the catfish ovary.