Sperm quality of artificially matured shortfinned eel is not affected by human chorionic gonadotropin dose and route of administration.

Background: Acquisition of high quality sperm is key to the artificial propagation of eels in captivity, but fertility drugs are expensive and repeated handling is stressful to the fish. An interrupted treatment regime (an initial hormone injection to stimulate spermatogenesis, followed several weeks later by weekly booster injections to induce sperm maturation) for acquisition of sperm in captive male eels has promise for high sperm quality on the one hand, and animal welfare benefits on the other. To further develop this approach for shortfinned eel, Anguilla australis, we evaluated the efficacy of (i) different initial doses of human chorionic gonadotropin (hCG) and (ii) route of administration. Methods: Male eels were artificially induced to mature with a single injection of 0, 250, 500 or 1,000 IU/fish of hCG, administered either intramuscularly (IM) or intraperitoneally (IP). Sperm maturation was induced with 150 IU hCG/fish from week 5 onwards and sperm collected for evaluation of quality by computer-assisted sperm analysis. Results: Control males did not mature and hence, sperm could not be retrieved and analysed, but all other treatments were effective in inducing testicular maturation. Milt volume tended to be higher for fish injected IM compared to those injected IP, whereas hCG dose had no effect. Conversely, the concentration of spermatozoa tended to be higher for several sperm collection time points in IP-injected than in IM-injected fish. Sperm quality, represented by percent motility, percent progressive motility and curvilinear velocity, was equal in fish given an initial dose of 250 IU hCG to those given higher initial doses of hCG. Conclusions: We recommend that an initial dose of 250 IU hCG/fish be administered to induce spermatogenesis in male A. australis, and, after a period of 4-5 weeks, weekly booster injections of ∼150 IU hCG/fish be administered in the day prior to sperm collection; both routes of administration (IM or IP) are equally effective. We contend that an interrupted treatment regime has notable benefits for induced maturation in male anguillids, as it reduces fish handling and manipulation and reduces the resources required to produce high quality sperm.

Steroidogenic acute regulatory protein transcript abundance in the eel, Anguilla australis: changes during the induced reproductive cycle and effects of follicle-stimulating hormone during previtellogenesis.

Steroidogenic acute regulatory protein (StAR) mRNA levels in the eel ovary were assayed by quantitative PCR and related to plasma steroid levels throughout oogenesis in order to shed light on the previously considered 'aberrant' prematurational increase in plasma levels of estradiol-17ß (E2). Total ovarian StAR transcript abundance mirrored circulating levels of E2, but not of 11-ketotestosterone (11KT). The study was complemented by evaluation of in vitro effects of follicle-stimulating hormone (FSH) on ovarian StAR transcript abundance and on short-term ('acute') radiolabelled pregnenolone-supported steroid metabolism by ovarian fragments to understand how the production of steroids during previtellogenic oocyte growth is regulated. We observed a significant effect of FSH on StAR mRNA levels within 24h of incubation, but these were no longer evident by 4 days of culture. Unexpectedly, FSH had no effect on substrate-supported steroidogenesis, as comparable yields of steroid products were detected using semi-quantitative HPLC and scintillation counting. We conclude that the eel ovarian follicle can respond to FSH from a very early stage of development (early oil droplet stage) by increasing StAR mRNA levels, but that there is no evidence for acute effects of FSH on bioactive steroid production downstream of cytochrome P450 side-chain cleavage. Furthermore, the prematurational increase in StAR mRNA in vivo is in keeping with general teleost models and is likely to be a 'normal' response to reaching advanced stages of development.

Effects of reproductive stage and 11-ketotestosterone on LPL mRNA levels in the ovary of the shortfinned eel.

To understand the dynamics of lipid uptake into the ovary and the potential role that lipoprotein lipase plays in this event, changes in LPL transcript abundance during oogenesis were measured in both wild-caught and pituitary homogenate-induced artificially maturing eels. Also, the effects of 11-ketotestosterone (11-KT) on LPL mRNA levels were investigated in vivo and in vitro. Normalized ovarian LPL transcript abundance increased as oogenesis advanced, and it rose particularly rapidly during midvitellogenesis, corresponding to pronounced increases in ovarian lipid deposits and LPL activity. Furthermore, LPL mRNA levels were dramatically increased following 11-KT treatment in vivo, findings that were reinforced as trends in ovarian tissue incubated in vitro. Ovarian LPL appears to be directly involved in the uptake of lipids into the eel ovary, an involvement that appears to be controlled, at least in part, by the androgen 11-KT.

11-Ketotestosterone and IGF-I increase the size of previtellogenic oocytes from shortfinned eel, Anguilla australis, in vitro.

Previtellogenic ovarian fragments from eel, Anguilla australis, were cultured in vitro in a chemically defined medium containing steroids and/or peptide hormones for 18 days in order to investigate their involvement in control of early oocyte growth. 11-Ketotestosterone (11-KT), but not estradiol-17beta, induced a significant 10-20% increase in diameters of previtellogenic oocytes and oocyte nuclei in a dose-dependent manner. Effects were greatest for 100 nM 11-KT, a dose that is within the physiological range seen in very early vitellogenic eels in the wild. The effect was not accompanied by obvious ultrastructural changes in the oocytes other than an apparent increase in nuclear size. Similarly, treatment with recombinant human IGF-I resulted in increased oocyte diameters, whereas no such effect was seen after treatment with heterologous insulin, GH, leptin, or human chorionic gonadotropin. Interestingly, lipid supplementation also resulted in an increase in oocyte diameter, and greater radioactivity in ovarian explants following incubation with (14)C-triglycerides and 11-KT, but not FSH, suggesting that the androgen may play a role in lipid accumulation into the oocyte. Our results implicate hormones from both the reproductive and the metabolic axes in control of previtellogenic oocyte growth in a teleost fish.