Tissue- and sex-specific regulation of CYP19A1 expression in the Atlantic croaker (Micropogonias undulatus)

To better define the tissue- and sex-specific roles of aromatase in fishes, we have isolated a CYP19A1 cDNA sequence from a well-developed model of teleost reproduction, the Atlantic croaker (Micropogonias undulatus). This cDNA encodes a protein which has high identity (57-90%) to known CYP19A1 proteins and segregates with teleost CYP19A1 proteins in molecular phylogenetic analysis. In both sexes, the gene encoding Atlantic croaker CYP19A1 is expressed primarily in gonadal tissue, but also in the brain and other tissues at much lower levels, as determined relative to ribosomal 18S RNA expression by real-time quantitative RT-PCR. In females, the highest levels of CYP19A1 mRNA are found in the developing ovary compared to spawning, regressing and resting ovaries. In contrast, testicular CYP19A1 expression is lowest in developing testes and increases in spawning and regressing testes, although there were no statistically significant differences between stages. Brain CYP19A1 mRNA levels are lower in animals with developing gonads compared to spawning fish. In vitro treatment with human chorionic gonadotropin (10 IU/ml) for 6 or 24h increases CYP19A1 mRNA approximately 16- and 43-fold, respectively, in isolated Atlantic croaker ovarian follicles, but has no effect on CYP19A1 mRNA in testicular or brain minces. Six hour in vitro treatment with sex steroids (estradiol, testosterone or 17,20 beta,21-trihydroxy-4-pregnen-3-one; 290 nM) does not alter CYP19A1 mRNA in ovary, testis or brain. The regulation of CYP19A1 in the Atlantic croaker therefore differs in a tissue- and sex-specific manner.     

Effects of follicle stimulating hormone on estradiol-17 beta production and P-450 aromatase (CYP19) activity and mRNA expression in brown trout vitellogenic ovarian follicles in vitro

In order to determine whether follicle stimulating hormone (FSH) regulates P-450 aromatase (P-450 arom) in salmonid fish, we investigated the in vitro effects of FSH on estradiol (E(2)) production and P-450 arom activity and expression in brown trout (Salmo trutta) vitellogenic ovarian follicles. Brown trout ovarian follicles were incubated in the presence of coho salmon FSH and the production of E(2) into the medium was measured by RIA, the activity of P-450 arom by the tritiated water release assay and the expression of P-450 arom by Northern blotting using a homologous cDNA probe obtained by RT-PCR. Results from this study indicate that the dose- and time-dependent stimulatory effects of FSH on E(2) production are dependent on new RNA and protein synthesis. The basal and FSH-stimulated E(2) production was completely blocked by fadrozole, a specific aromatase inhibitor. FSH was capable of stimulating P-450 arom activity but this stimulation was only detectable with short incubation times (30 min) since longer incubation times with FSH resulted in the inhibition of P-450 arom activity. In addition, FSH increased the steady-state P-450 arom mRNA levels. In conclusion, our results indicate, for the first time in teleost fish, that FSH stimulates the expression of P-450 arom, as well as its activity, albeit after a short-term treatment with FSH, and that FSH plays a fundamental role in the regulation of the production of E(2) in the salmonid ovary.     

Masculinization of the European sea bass (Dicentrarchus labrax) by treatment with an androgen or aromatase inhibitor involves different gene expression and has distinct lasting effects on maturation

The objective of this study was to contribute to our understanding of the role of sex steroids in fish sex differentiation and male maturation. Sexually undifferentiated sea bass were administered 17α-methyldihydrotestosterone (MDHT), estradiol-17β (E₂), fadrozole (Fz), cyproterone acetate (CPA) or tamoxifen (Tx). MDHT produced 100% males whereas E₂ and Tx resulted in 100% females. Fz produced 95% males while CPA did not alter sex ratios. E₂ treatment did not affect gonadal aromatase (cyp19a) expression levels, supporting the possibility that the feminizing effect of exogenous E₂ are not directly related to cyp19a regulation. Both MDHT and Fz decreased cyp19a expression. Moreover, androgen receptor (ar) expression levels increased during development in all but the MDHT group, suggesting that early exposure to an androgen down-regulates subsequent ar expression in males and that Fz does not interact with the androgen receptor. Together, these observations indicate that although MDHT and Fz result in a similar phenotype, the molecular pathways involved are likely different, and show that Fz masculinization is the consequence of inhibited ovarian differentiation rather than of a direct androgenic effect. Further, since CPA did not alter sex ratios when administered during the period of highest androgen sensitivity, we suggest that androgens are not required for initial testicular differentiation in the sea bass. MDHT and Fz did not alter the number of precocious males but reduced and increased, respectively, their gonadosomatic index (GSI). In addition, Fz had lasting effects on the GSI of precocious and non-precocious males, probably due to alterations of estrogen function in the testis.