In vitro steroidogenesis of the gonads of the protogynous Pacific wrasse, Haliochoeres trimaculatus

Abstract

Testicular and ovarian fragments of the protogynous Pacific wrasse Haliochoeres trimaculatus were incubated in vitro with [3H]pregnenolone ([3H]P5), [3H]17‐hydroxyprogesterone ([3H]17OHP4), non‐radioactive (nr) 17β‐oestradiol (nrE2) or nrP5 to identify the major gonadal steroidogenic pathways and steroid products in females and in the two male variants of this species, the terminal phase (TP) and initial phase (IP) males. Both testis and ovarian tissues exhibited 7 hydroxylase activity resulting in the formation of 7α‐hydroxypregnenolone (7OHP5) from [3H]P5, and many HPLC peaks were identified as products of testicular (c. 29) and ovarian (c. 23) steroidogenesis, and only c. 50% of these metabolites co‐eluted with authentic reference standards; only very small amounts of conjugated steroid were synthesized from any of the precursors. [3H]P5 was converted by testis mainly to 7αOHP5, and two unknown steroids, whereas [3H]17OHP4 metabolism gave rise to [3H]17,20β‐dihydroxy‐4‐pregnen‐3‐one (DHP), 11‐ketotestosterone (11KT), and two unknown steroids. For ovarian tissues, [3H]17OHP4 and [3H]P5 were metabolized to form E2, oestrone (E1), androstenedione (A4), 20α‐ and 20β‐dihydroprogesterone (20αDHP and 20βDHP), 7αOHP5 (from [3H]P5) and a major unknown. The HPLC steroid profiles for testis incubations for IP and TP males were similar, however, the steroidogenic response of the testis of TP males to human chorionic gonadotrophin, in vitro (determined by hormone assay), was significantly higher than that of IP males.

Sex-change and gonadal steroids in sequentially-hermaphroditic teleost fish

“…In particular, this method of study has confirmed the role of steroid hormones as mediators of sex-change processes in fish. Another method of study is the in vitro incubation of pre-, peri-and post-transitional gonad tissues with radio-labeled steroid precursors, followed by chromatographic identification of the steroidal products (Guiguen et al, 1995;Lee et al, 1995;Leatherland et al, 2003). This technique has been most useful for identifying novel steroids in hermaphroditic fishes, and for characterizing changes in steroidogenic pathways that occur during sexchange.…”

Cortisol Inhibition of 17b-Estradiol Secretion by Rainbow Trout Ovarian Follicles Involves Modulation of Star and P450scc Gene Expression

“…Freshly-dissected ovarian tissue from each fish was placed in icecold Cortland's medium (supplemented with 0.1% glucose, 0.1% bovine serum albumin, and 0.1% streptomycin sulfate at pH 7.55) and kept at a low temperature throughout the preparation of individual ovarian follicles for incubation, as described previously [31][32][33].…”

“…Radioimmunoassays (RIAs) were used to measure the concentrations of E 2 and T in the medium as described previously [32,33]. To determine the specificity of the assays, free steroids that had been extracted from the medium were separated by HPLC; the fractions were then dried under nitrogen, reconstituted in phosphate-buffered saline and the steroid levels measured in the reconstituted fractions.…”

“…The unconjugated (free) steroid fraction was eluted by passing 5 ml of diethyl ether through the cartridge, and the conjugated steroid fraction was then eluted by passing 5 ml of methanol through the cartridge. Both diethyl ether and methanol fractions were dried under nitrogen at < 45 o C and the sulfate and glucuronide conjugates were sequentially separated as free steroids following acid solvolysis (sulfate) and glucuronidase treatments (glucuronide), respectively [31][32][33]. To measure the recovery of tritium-labeled steroid at each separation step, a 100 μl aliquot was taken from each fraction, mixed with 4 ml of scintillation cocktail (Ecolite, MP Biochemicals, Solon, OH), and counted in a Searle Delta 300 counter.…”

Fecundity of banded wrasse(Notolabrus fucicola)from Otago, Southern New Zealand

“…Possible explanations are that hCG is not, or poorly, recognised by the wrasse ovary, akin to the situation in amago salmon (Oncorhynchus rhodurus; Young et al 1983). However, in vitro sensitivity to hCG in other wrasse species, such as the bambooleaf wrasse, Pseudolabrus japonicus (Matsuyama et al 1998) and Pacific wrasse, Haliochoeres trimaculatus (Leatherland et al 2003), suggests this is an unlikely explanation. Similarly, it is possible that 17a,20P-P is not the MIH, although this is unlikely in light of the findings on other wrasse (e.g., Matsuyama et al 2002).…”