Cortisol Extraction from Sturgeon Fin and Jawbone Matrices.

The aims of this study were to develop a technique for the extraction of cortisol from sturgeon fins using two washing solvents (water and isopropanol) and quantify any differences in fin cortisol levels among three main sturgeon species. Fins were harvested from 19 sacrificed sturgeons including seven beluga (Huso huso), seven Siberian (Acipenser baerii), and five sevruga (A. stellatus). The sturgeons were raised in Iranian farms for 2 years (2017-2018), and cortisol extraction analysis was conducted in South Korea (January-February 2019). Jawbones from five H. huso were also used for cortisol extraction. Data were analyzed using the general linear model (GLM) procedure in the SAS environment. The intra- and inter-assay coefficients of variation were 14.15 and 7.70, respectively. Briefly, the cortisol extraction technique involved washing the samples (300 ± 10 mg) with 3 mL of solvent (ultrapure water and isopropanol) twice, rotation at 80 rpm for 2.5 min, air-drying the washed samples at room temperature (22-28 °C) for 7 days, further drying the samples using a bead beater at 50 Hz for 32 min and grinding them into powder, applying 1.5 mL methanol to the dried powder (75 ± 5 mg), and slow rotation (40 rpm) for 18 h at room temperature with continuous mixing. Following extraction, samples were centrifuged (9,500 x g for 10 min), and 1 mL supernatant was transferred into a new microcentrifuge tube (1.5 mL), incubated at 38 °C to evaporate the methanol, and analyzed via enzyme-linked immunosorbent assay (ELISA). No differences were observed in fin cortisol levels among species or in fin and jawbone cortisol levels between washing solvents. The results of this study demonstrate that the sturgeon jawbone matrix is a promising alternative stress indicator to solid matrices.

Effects of 11-ketotestosterone and temperature on inhibin subunit mRNA levels in the ovary of the shortfinned eel, Anguilla australis.

Members of the transforming growth factor-b (TGFb) superfamily are important during early oogenesis in mammals. In this study, we tested whether documented effects of 11-ketotestosterone (11KT) on previtellogenic eel ovaries are mediated through affecting the expression of key ovarian TGFb genes. Furthermore, we investigated whether 11KT effects interacted with temperature. Accordingly, three thermal regimes were compared and their interaction with 11KT-mediated actions on expression of TGFb superfamily genes (chiefly inhibin subunits) evaluated in the eel (Anguilla australis). Inhibin subunit mRNA levels were also measured in ovarian explants cultured in vitro with 11KT and in ovaries from eels collected from the wild. In wild eels, inhibin-bA mRNA levels were higher in early than in previtellogenic eels; inhibin-a expression did not differ between stages, whereas that of inhibin-bB first decreased, then recovered with advanced developmental stage. Temperature was ineffective in modulating any of the end points, at least as long as a Q10 adjustment was made to correct for 'metabolic dose'. However, 11KT affected the expression of inhibin-a compared to control fish, while those of inhibin-b subunit genes remained unaffected. In contrast, 11KT dramatically reduced mRNA levels of inhibin-b subunits in vitro, but had inconsistent effects on inhibin-a transcript abundance. We conclude that 11KT affects ovarian inhibin subunit gene expression, but effects are not in keeping with the changes seen during early oogenesis in eels from the wild. We further contend that in vivo temperature experiments are easily biased and that Q10 corrections may be required to identify 'true' temperature effects.

Androgen-specific regulation of FSH signalling in the previtellogenic ovary and pituitary of the New Zealand shortfinned eel, Anguilla australis.

The evidence for androgens having a pivotal role in the functioning of the female reproductive axis--such as initiating puberty or vitellogenesis--is mounting. However, the use of aromatizable androgens and the tissue-specific focus of most studies often make it unclear if androgenic effects throughout the axis proceed via androgen or estrogen signalling mechanisms. In this study, we assessed the effects of 11-ketotestosterone (11KT, a non-aromatizable androgen) on the pituitary and ovary of previtellogenic (PV) freshwater eels Anguilla australis, comparing them with eels naturally undergoing early vitellogenesis (EV). We found that 11KT treatment produces molecular and morpho-physiological phenotypes that were generally intermediate between PV and EV. Most notably, we demonstrated that 11KT induces effects on follicle-stimulating hormone (FSH) signalling in the pituitary and ovaries that are in opposition to each other. Thus, 11KT significantly reduced fshß subunit expression in the pituitary. At the same time, 11KT dramatically increased mRNA levels of ovarian FSH receptor and plasma levels of estradiol-17ß, very likely sensitizing the previtellogenic follicle to the FSH signal. Androgens therefore may be important in facilitating puberty in the eel.