Thermal and endocrine regulation of gonadal apoptosis during sex differentiation in pejerrey Odontesthes bonariensis.

The fate of the differentiating gonads in pejerrey Odontesthes bonariensis is determined by the environmental water temperature experienced by the larvae during the critical period of sex determination. We previously reported a link between apoptosis, temperature and sex differentiation in this species. To clarify this link, we subjected larvae to thermal and endocrine treatments between hatching and the onset of histological sex differentiation of the gonads and assessed the patterns of gonadal development and apoptosis by light microscopic histology and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) analysis, respectively. Apoptotic labeling was widespread among somatic cells of the anterior region of the right gonads of fish reared at the male-producing temperature (MPT) and part of the fish at the mixed sex-producing temperature prior to sex differentiation. In contrast, TUNEL-positive cells were rarely observed in gonads at the female-producing temperature (FPT). Administration of exogenous estrogen completely prevented MPT-induced masculinization, induced feminization and reduced gonadal apoptosis, whereas an aromatase inhibitor (fadrozole) induced TUNEL signals in the gonads of FPT-reared larvae. These results provide strong evidence that apoptosis in somatic cells in the right lobe of the gonads might play a key role in testicular differentiation in pejerrey and that estrogens are involved in the regulation of this process.

Effects of light and temperature conditions on the expression of GnRH and GtH genes and levels of plasma steroids in Odontesthes bonariensis females.

In this study we examined the endocrine mediation between environmental factors (temperature and photoperiod) and the brain-pituitary-gonadal axis in females of pejerrey Odontesthes bonariensis. Changes in the expression of brain gonadotropin-releasing hormones (GnRHs) and gonadotropin (GtH) subunit [follicle stimulating-beta (FSH-beta), luteinizing hormone-beta (LH-beta), glycoprotein hormone-alpha (GPH-alpha)] genes, plasma gonadal steroids [estradiol (E(2)) and testosterone (T)], gonadal histology, and gonadosomatic index (GSI) in adult females exposed to combinations of short-day (8 h) or long-day (16 h) photoperiods and low (12 degrees C) or high (20 degrees C) temperatures after winter conditions (8 h light, 12 degrees C) were analyzed. Pejerrey females kept under the short photoperiod had low GSIs, and their ovaries contained only previtellogenic oocytes regardless of the experimental temperature. In contrast, females exposed to the long photoperiod had high GSIs and ovaries with vitellogenic oocytes at both temperatures. These fish also showed a significantly higher expression of sGnRH, pjGnRH, cGnRH-II (the three different GnRH variants found to date in the pejerrey brain), FSH-beta, LH-beta and GPH-alpha genes and plasma E(2 )levels than those at the shorter photoperiod. No significant changes were observed in plasma T levels. Based on these results, we concluded that the increase in day length but not that of temperature triggers the maturation of pejerrey females after the winter period of gonadal rest and that this occurs by an integrated stimulation of the various components of the brain-pituitary-gonad axis.

mRNA expression of GnRH variants and receptors in the brain, pituitary and ovaries of pejerrey (Odontesthes bonariensis) in relation to the reproductive status.

The present study examined the differential mRNA expression levels of three forms of GnRH (sGnRH, pjGnRH and cGnRH-II) and two forms of GnRH receptor (pjGnRH-R I and pjGnRH-R II) in the brain, pituitary, and ovaries of pejerrey in relation to the reproductive status. The analysis revealed the presence of significant amounts of mRNA of the three GnRH forms while the ovaries showed only two (sGnRH and pjGnRH). The GnRH receptor II was found ubiquitously in the brain, pituitary, and ovaries while the form I was detected only in the brain. The levels of pjGnRH mRNA in the brain and pjGnRH-R II in the pituitary gland varied in correlation with the ovarian condition. However, brain sGnRH and pjGnRH-R I mRNA levels reached a maximum during early stages of ovarian development. In contrast, the brain levels of cGnRH-II mRNA showed no variation. The present study also shows a good correlation of ovarian sGnRH and pjGnRH-R II mRNA levels with the reproductive condition, suggesting that these molecules are may be involved in the regulation of pejerrey ovarian function.

Dimorphic expression of dmrt1 and cyp19a1 (ovarian aromatase) during early gonadal development in pejerrey, Odontesthes bonariensis.

The pejerrey (Odontesthes bonariensis) is a teleost fish with strong temperature-dependent sex determination (TSD). Several studies have shown that dmrt1 and gonadal aromatase (cyp19a1) are implicated in the sex differentiation process in teleosts but little is known on the expression balance and endocrine regulation of these two genes during TSD. This study was designed to clarify the expression patterns of both genes during gonadal sex differentiation of pejerrey reared at female-, male- and mixed-sex-producing temperatures (FPT, MPT, and MixPT, respectively). The expression of dmrt1 was found to be significantly higher during gonadal sex differentiation at MPT compared to FPT. Conversely, cyp19a1 expression clearly increased during differentiation at FPT but not at MPT. The expression of both genes at MixPT showed a dimorphic profile with individual values resembling either those at the MPT or FPT. Administration of exogenous 17beta-estradiol down- and up-regulated the expression of dmrt1 and cyp19a1, respectively, regardless of temperature, and rescued the female phenotype at the MPT. However, treatment with the aromatase inhibitor Fadrozole caused masculinization without changing the pattern of gene expression. These results are strong evidence of the involvement of both genes in the gonadal differentiation process of pejerrey. The involvement of estradiol is discussed.

The significance of cortisol on acclimation to salinity in pejerrey Odontesthes bonariensis

The role of cortisol on the osmoregulation of pejerrey Odontesthes bonariensis at different salinities was investigated in adult fish injected with 0.7mg hydrocortisone per 100g body weight of fish, and transferred to 0, 5 and 20ppt of NaCl. Blood cortisol was 566ng/ml at the beginning of the experiment (0h) but surged to 1250ng/ml within 3h in cortisol-injected fish. Cortisol levels were influenced not only by treatment but also by time, being higher at 3h compared to 24h. Salinity level, time of exposure and their interaction, but not cortisol treatment, significantly affected plasma osmolality and the concentration of ions Cl- and Na+. This study showed that exogenous cortisol does not seem to play a significant role on the regulation of plasma osmolality and concentration of individual ions in pejerrey

Investigou-se a participação do cortisol na osmoregulação de peixe-rei Odontesthes bonariensis, em diferentes salinidades, em peixes adultos injetados com 0,7mg hidrocortisona por 100g de peso corporal, e transferidos para 0, 5 e 20ppt de NaCl. No inicio do experimento (0h), o cortisol encontrado no plasma foi de 566ng/ml, aumentando para 1250ng/ml em 3h em peixes injetados com cortisol. A concentração de cortisol foi influenciada não somente pelo tratamento, mas também pelo tempo, sendo maior 3h após a inoculação, comparada à 24h. A salinidade, o tempo de exposição e a interação desses dois fatores, mas não o tratamento com o cortisol, afetaram significativamente a osmolaridade e a concentração dos íons Cl- e Na+ do plasma. Este estudo mostrou que, o cortisol exógeno não influi significativamente na regulação da osmolaridade e da concentração de íons Na+ e Cl- no plasma em peixe-rei