The effects of thyroid hormones on hepatic insulin-like growth factor (IGF) II gene expression and their interaction with cortisol in the ontogenic control of this gene were investigated in fetal sheep during late gestation (term 145 ± 2 days) and after experimental manipulation of fetal plasma hormone concentrations. In intact fetuses, a significant decrease in hepatic IGF-II mRNA abundance was observed between 127-130 and 142-145 days of gestation, which coincided with the normal prepartum rise in plasma cortisol and triiodothyronine (T₃) concentrations. This ontogenic decline in hepatic IGF-II gene expression was abolished in fetuses in which the prepartum rise in plasma T₃, but not cortisol, was prevented by fetal thyroidectomy. At 127-130 days, downregulation of hepatic IGF-II mRNA abundance was induced prematurely in intact fetuses by an infusion of cortisol for 5 days (2-3 mg · kg¹ · day¹ iv). Plasma concentrations of cortisol and T₃ in the cortisol-infused intact fetuses were increased to values seen close to term. Similar findings were observed in thyroidectomized fetuses, in which, despite thyroidectomy, cortisol infusion significantly increased plasma T₃ concentrations and caused a premature decrease in hepatic IGF-II mRNA levels. However, in intact fetuses at 127-130 days, the increasing of T₃ concentrations alone by exogenous T₃ infusion (8-12 µg · kg¹ · day¹ iv for 5 days) had no effect on hepatic IGF-II mRNA levels. Overall, a decrease in hepatic IGF-II mRNA abundance was only observed in fetuses in which there were concurrent increases in plasma cortisol and T₃ concentrations. When observations from all fetuses were considered, irrespective of gestational age or treatment, hepatic IGF-II mRNA levels were negatively correlated with plasma cortisol and T₃ but not thyroxine concentrations. Partial correlation analysis of hepatic IGF-II, cortisol, and T₃ values showed that the plasma concentration of cortisol in the fetus had the predominant effect on hepatic IGF-II mRNA abundance. These findings show that T₃ may mediate, in part, the maturational effects of cortisol on hepatic IGF-II gene expression but that it is ineffective without a concomitant rise in fetal plasma cortisol. Hence, increased concentrations of both cortisol and T₃ appear necessary to induce downregulation of hepatic IGF-II mRNA abundance in fetal sheep close to term.