Abstract
Sex steroids are involved in sex determination in almost all vertebrates, including species with temperature-dependent sex determination (TSD). It is well established that aromatase and estrogens are involved in ovary determination in TSD species. In contrast, the role of non-aromatizable androgens in TSD is less clear. In this study, we used dihydrotestosterone (DHT) and an antagonist of the mammalian androgen receptor (flutamide) to examine the impact of androgens on sex determination in the snapping turtle. We incubated eggs at a male-producing temperature and treated embryos with drug delivery vehicle (5 L ethanol), DHT in vehicle, or flutamide in vehicle during the sex-determining period. We then measured expression of markers for ovarian and testicular development and genes involved in steroidogenesis. A subset of embryos and hatchlings were collected for histological analysis of gonad differentiation and sex determination. DHT and flutamide both induced ovarian development: 100% of vehicle-treated hatchlings had testes, while 60% of DHT-treated and 32% flutamide-treated hatchlings had ovaries. DHT and flutamide treatments also had feminizing effects on gene expression patterns and the structure of embryonic gonads. DHT treatment increased expression of FoxL2, androgen receptor, aromatase and several steroidogenic genes. Flutamide produced a similar, but weaker, pattern of gene expression. Genes involved in testis development (Sox9 and Amh) were influenced by flutamide treatment. Our findings support the hypothesis that androgens and the androgen receptor are involved in ovary determination in the common snapping turtle.