In vivo effect 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 the administration of Argyrosomus regius rFsh and rLh on the reproductive development of prepubertal meagre juveniles at the initial stages of sexual differentiation. Juvenile meagre, 9-months old with mean weight of 219 ± 3.9 g (mean ± SEM) were randomly distributed into nine groups (n = 8 per group). Experimental groups were treated weekly with an acute injection of either rFsh or rLh. Control groups were injected with saline solution. In a 3-week experiment, different groups were administered with different doses 6, 12 or 18 µg kg⁻¹ of rFsh or rLh or saline solution. In a 6-week experiment a group was administered with 12 µg kg⁻¹ of rFsh and a second group with saline solution. The fish were held in a single 10 m³ tank with natural photoperiod (Feb. – March) and temperature 16.1 ± 0.4 °C. At the start of the experiment (n = 8) and at the end of the 3-week experiment, fish were blood sampled and sacrificed. Blood was analysed for 17β-estradiol (E2) and 11-ketotestosterone (11-KT). Gonads and liver were dissected and weighed. Gonads were fixed in Bouińs solution and processed for histological analysis. Juvenile meagre at the start of the experiment were in the initial stages of sexual differentiation, indicated by the presence of the ovarian cavity or testes duct that was surrounded by undifferentiated embryonic germ stem cells and somatic cells. At the end of the 3-week 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 rFsh, rLh or saline all fish presented a similar gonadal structure as at the start of the experiment. However, the incidence of sporadic developing germ cells (principally spermatogonia, spermatocytes, spermatids, but also perinucleolar stage oocytes) generally increased in rGth treated meagre. A mean of 44 % of meagre treated with rFsh or rLh presented sporadic isolated 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 experiments 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 a proportion of the rFsh group after six weeks of treatment compared to the start of the experiment and the saline control group on week 6. The application of rFsh or rLh to meagre at the initial stages of sex differentiation did not stimulate steroid production until week six (11-KT) and had a limited, but evident effect on the development of sporadic isolated germ cells. However, we conclude that rGth, rFsh or rLh did not stimulate large developmental changes in sexually undifferentiated meagre gonads.