The late gestation plasma cortisol surge in the sheep fetus is critical for stimulating organ development and parturition. Increased adrenal responsiveness is one of the key reasons for the surge, however the underlying mechanisms are not fully understood. Our recent studies suggest that ACTH mediated increased expression of ACTH receptor (ACTH-R) and steroid acute regulatory protein (StAR) may play a role in enhancing responsiveness. Hence we examined the effects of ACTH infusion in fetal sheep on mRNA expression of these two mediators of adrenal responsiveness, and also assessed the functional consequences of this treatment in vitro. Fetuses of around 118 and 138 days gestational age (dGA) were infused with ACTH_1-24 for 24 h. Controls received saline infusions. Arterial blood was sampled throughout the infusion. Adrenals were isolated and analyzed for ACTH-R and StAR mRNA, or the cells cultured for 48 h. Cells were stimulated with ACTH, and medium collected for cortisol measurement. Fetal plasma ACTH and cortisol concentrations increased over the infusion period in both groups. ACTH-R mRNA levels were significantly higher in ACTH infused fetuses in both groups (118 dGA: 0.51 ± 0.06 vs. 0.28 ± 0.05, 138 dGA: 0.91 ± 0.04 vs. 0.60 ± 0.06; values are ACTH-R mRNA:28SrRNA). StAR mRNA increased significantly in both the 138 (0.56 ± 0.06 to 0.74 ± 0.02 StAR mRNA:28SrRNA), and the 118 (0.23 ± 0.05 to 0.40 ± 0.08) dGA groups. Adrenal cells from ACTH infused fetuses were significantly more responsive to ACTH stimulation in terms of cortisol secretion than those from saline infused controls. These findings demonstrate that increases in circulating ACTH levels promote increased expression of ACTH-R and StAR mRNA, and are coupled to heightened adrenal responsiveness.