Long-term exposure to environmentally relevant concentrations of progesterone and norgestrel affects sex differentiation in zebrafish (Danio rerio).

The aim of this study was to investigate the effects of progestins on the sex differentiation of zebrafish by measuring the sex ratio and transcriptions of genes related to sex differentiation (Amh, Dmrt1, Figa, Sox9a and Sox9b genes) as well as sex hormone levels and transcriptional expression profiles along the hypothalamic-pituitary-gonadal (HPG) and hypothalamic-pituitary-adrenal (HPA) axes in juvenile zebrafish. Exposure of zebrafish to 4, 33, 63ngL(-1) progesterone (P4) or 4, 34, 77ngL(-1) norgestrel (NGT) started at 20 days post fertilization (dpf) and ended at 60 dpf. The results showed that exposure to P4 caused a significant increase in proportion of females as well as significant down-regulation of Amh gene and up-regulation of Figa at a concentration of 63ngL(-1). However, the shift in the sex ratio toward males was observed following exposure to 34 and 77ngL(-1) NGT, which came along with the significant induction of Dmrt1 gene and inhibition of Figa gene. The sex hormones in exposed fish were measured with estrone being detected only in the fish exposed to the highest P4 concentration; whereas estradiol and androstenedione were detected only in the fish of the control and lowest NGT concentration. Furthermore, the increase in females was associated with the significant up-regulation of several key genes controlling the synthesis of sex hormones (i.e., Cyp17, Cyp19a1a and Hsd3b) following exposure to 63ngL(-1) P4 whereas the significant down-regulation of Cyp11a1, Cyp17, Cyp19a1a and Hsd3b genes was observed in the male-biased populations caused by 34 and 77ngL(-1) NGT. The overall results imply that both P4 and NGT could significantly affect sex differentiation in zebrafish, and that changes may be reflected by altered sex hormone levels and transcriptional expression profiles of genes related to synthesis of sex hormones.