Presence of Gonadotropin-Releasing Hormone (GnRH) Binding Sites and Compounds with GnRH-Like Activity in the Ovary of African Catfish, Clarias Gariepinus1

Abstract

GnRH binding was characterized in the African catfish ovary by use of an analog of salmon GnRH (sGnRH- ; [D-Arg6, Trp7, Leu8, Pro9-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 (approximately 22 degrees C) at pH 7.6. Addition of unlabeled sGnRH-A displaced the bound 125I-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 (Kd) of 0.27 +/- 0.036 nM. Bound 125I-sGnRH-A was also found to be displaceable by catfish GnRH (cfGnRH; [His5, Leu7, Asn8]-GnRH), chicken GnRH-II (cGnRH-II; [His5, Trp7, Tyr8]-GnRH), and salmon GnRH (sGnRH; [Trp7, Leu8]-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 from goldfish pituitary, as well as displacing 125I-sGnRH-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.

Post-ovulatory secretion of pituitary gonadotropins GtH I and GtH II in the rainbow trout (Oncorhynchus mykiss): regulation by steroids and possible role of non-steroidal gonadal factors

“…Indeed, the presence of a GnRH-like factor has been demonstrated in the African catfish (Habibi et al 1994) and goldfish ovary (Pati & Habibi 1998). However, it is improbable that GnRH would stimulate GtH II secretion but inhibit that of GtH I.…”

Hormonal regulation of follicular atresia in teleost fish

“…In that study, however, the early eluting compound with GnRH-like activity could not be identified, suggesting the presence of either a novel form of GnRH or a GnRH variant resulting from differential splicing of GnRH transcripts (Pati and Habibi, 1998;Nabissi et al, 2000). The presence of compounds with GnRH-like activity was also demonstrated in the ovary of African catfish (Habibi et al, 1994) and seabream (Nabissi et al, 1997). In spite of extensive attempts to purify GnRH peptides from the ovary, cGnRH-II peptide, which is present in the goldfish brain, was not detected in goldfish ovarian extracts suggesting that cGnRH-II expression in the ovary of this species is either very low or transient.…”

“…The latter is a relatively new hypothesis, which is based on recent studies carried out in goldfish and seabream. In this regard, there is unambiguous evidence demonstrating the presence of GnRH and GnRH-R in the gonadal tissue of fish and other vertebrate species (Pati and Habibi, 1992Habibi et al, 1994Habibi et al, , 2001Gazourian et al, 1997;Nabissi et al, 1997). Further studies have provided strong support for the hypothesis that locally produced GnRH plays an autocrine/paracrine role in the regulation of gonadal function.…”

“…Further studies have provided strong support for the hypothesis that locally produced GnRH plays an autocrine/paracrine role in the regulation of gonadal function. In fish, ovarian extract was shown to contain compounds with GnRH-like activity Habibi et al, 1994;Nabissi, et al, 1997;Habibi, 1999). In these studies, treatment of goldfish cultured pituitary fragments with ovarian extracts was found to stimulate LH release and subunit gene expression, which could be inhibited by co-treatment with a GnRH antagonist.…”

Localization of gonadotrophin-releasing hormone I, bradykinin and their receptors in the ovaries of non-mammalian vertebrates

“…Among vertebrates, the presence of GnRH I-receptor has previously been demonstrated mainly in the ovaries of several species of fishes: African catfish, Clarias gariepinus (Habibi et al 1994), sea bream, Sparus aurata (Nabissi et al 1997), goldfish, Carassius auratus (Pati & Habibi 1998) and a teleost, Fugu rubripes (Moncaut et al 2005). The presence of GnRH I-receptor in the vertebrate ovary suggests that GnRH I acts through a receptormediated process (Schirman-Hildesheim et al 2005).…”