In vivoeffect of recombinant FSH and LH administered to meagre (Argyrosomus regius) at the initial stages of sex differentiation

ABSTRACT Recombinant gonadotropins, follicle stimulating (rFSH) and luteinizing hormone (rLH), offer the potential to induce gametogenesis in prepubertal fish. This study aimed to determine the in vivo effect of Argyrosomus regius rFSH (arrFSH) and Argyrosomus regius rLH (arrLH) administered to prepubertal meagre juveniles at the initial stages of sex differentiation. Meagre single-chain recombinant gonadotropins, arrFSH and arrLH were produced with the CHO expression system by Rara Avis Biotec, S. L. Juvenile meagre, 9-months old with mean weight of 222 ± 36 g (mean ± SD) were randomly distributed into seven experimental groups (n = 8 per group) that were treated weekly for three weeks with an acute injection of 6, 12 or 18 μg kg -1 of arrFSH (groups, 6-arrFSH, 12-arrFSH and 18-arrFSH) or 6, 12 or 18 μg kg -1 of arrLH (groups, 6-arrLH, 12-arrLH and 18-arrLH) or saline solution (Control group). Two more groups (n=8) were set up and treated for 6 weeks, with 12 μg kg -1 of arrFSH or saline Control. The fish were held in a 10 m 3 tank with natural photoperiod (Feb. – March) and temperature 16.1 ± 0.4°C. At the start of the experiment (n = 8) and end of the three week experiment all fish were sacrificed and gonads dissected. Gonads were weighed, fixed in Bouin’s solution and processed for histological analysis. Blood was sampled from all fish at the start and end of the experiment (week 3 and 6) for 17β-estradiol (E2) and 11-ketotestosterone (11-KT) analysis. Juvenile meagre at the start of the experiment were in the initial stages of sexual differentiation, indicated by the presence of the ovarian or testes lumen that was surrounded by undifferentiated embryonic germ stem cells and somatic cells. At the end of the experiment, there was no significant difference in gonadosomatic index (GSI) amongst control (initial and saline treated) and the experimental groups. After three weeks of application of arrFSH, arrLH or saline all fish presented a similar gonadal structure as at the start of the experiment. However, the incidence of isolated developing germ cells (principally spermatogonia, spermatocytes, spermatids, but also perinucleolar stage oocytes) generally increased in arrGTH treated meagre. A mean of 44 % of meagre treated with arrFSH or arrLH presented isolate developing germ cells, mainly male cells. Plasma steroid levels of E2 decreased significantly from the start of the experiments to the end. At the end of the experiment there were no differences in plasma E2 amongst Control fish and rGTH treated fish. Plasma 11-KT showed no change from the start of the experiment to week 3. However, a significant increase was observed in the arrFSH group after six weeks of treatment compared to the start of the experiment and the control group on week 6. The application of arrFSH or arrLH to meagre at the initial stages of sex differentiation did not stimulate steroid production until week six and had a limited, but evident effect on the development of isolated germ cells. The rGTHs, arrFSH or arrLH did not stimulate large developmental changes in undifferentiated gonads. HIGHLIGHTS Exogenous recombinant gonadotropins administered to meagre did not advance sexual differentiation. Exogenous recombinant gonadotropins administered to meagre did not affect plasma estradiol. Exogenous recombinant FSH increased the proportion of meagre with isolated male germ cells. Exogenous recombinant FSH increased plasma 11-ketotestosterone in meagre treated for 6 weeks.