In many turtles the temperature during the middle of incubation determines the gonadal sex of the hatchling. In the red-eared slider turtle (Trachemys scripta), an incubation temperature of 26 °C results in all male offspring, whereas an incubation temperature of 31 °C results in all female offspring; at temperatures intermediate to these (e.g. 29, 29·2, 29·4 °C) a mixed sex ratio is obtained. Administration of exogenous oestrogens will overcome the effects of an all-male producing incubation temperature to cause female sex determination, whereas administration of exogenous dihydrotestosterone (DHT) or testosterone to eggs incubating at an all-female temperature will have no discernible effect. Administration of DHT will cause male sex determination only if administered at intermediate incubation temperatures whereas administration of testosterone to eggs incubating at all male-producing and male-biased intermediate temperatures results in a significant number of female offspring, an effect presumably due to aromatization of testosterone to oestradiol (OE2), Since testosterone serves as the precursor to both DHT and OE2, being metabolized by reductase and aromatase respectively, three experiments were conducted to determine whether various putative reductase and aromatase inhibitors would overcome the effect of incubation temperature. First, while administration of testosterone to eggs incubating at all male-producing and male-biased intermediate temperatures produced females in a dose- and temperature-dependent manner, significant numbers of intersex individuals resulted from high dosage testosterone treatment to eggs incubating at a female-biased intermediate temperature. The reductase inhibitors 4MA and MK906 were capable of producing female offspring if administered at intermediate temperatures, but not in a dose-dependent fashion. Administration of the aromatase inhibitors CGS16949A and CGS20267 resulted in male offspring at both female-biased intermediate and at all female-producing temperatures in a dose-dependent fashion. Second, similar findings were obtained with combined doses of testosterone and reductase or aromatase inhibitors. Combined treatment of eggs at male-biased intermediate incubation temperatures with testosterone and reductase inhibitor resulted in female hatchlings, whereas combined treatment of testosterone and aromatase inhibitor at both female-biased intermediate and at all female-producing temperatures resulted in male hatchlings. Finally, treatment with reductase inhibitor and aromatase inhibitor combined resulted in only male offspring at all incubation temperatures with the exception of the all-female incubation temperature; in the latter instance almost all offspring were female. These studies indicate that in the red-eared slider turtle (i) male and female sex determination are independent cascades residing equally in each individual and regulated by incubation temperature, (ii) steroid hormones are involved in temperature-dependent sex determination, and (iii) testosterone plays a pivotal role in this process. The data also suggest that aromatase and oestrogen receptors may be involved in the initiation of an ovary determining cascade and that reductase and androgen receptors may be involved in the initiation of a testis determining cascade.