To investigate the role of the retino-hypothalamic tract on fetal prolactin regulation, we examined the effect of ocular enucleation on fetal plasma prolactin. Eleven fetuses of Suffolk ewes were chronically catheterized during fall, and six of them were subjected to bilateral ocular enucleation. All ewes were kept at 12h:12h light:dark cycle (lights on at 0800 and off at 2000). The experiments were performed 5-9 days after surgery (GA control fetuses 125 +/- 1.5, optical enucleation 121.3 +/- 1.5 days). Blood samples were taken from fetuses hourly around the clock, and plasma prolactin and cortisol were measured by radioimmunoassay (RIA). Luteinizing hormone (LH) and Growth hormone (GH) were measured in pooled plasma samples from control and enucleated fetuses by RIA. Average plasma prolactin was 5-fold lower in enucleated than in control fetuses (9.6 +/- 0.5 and 54.2 +/- 3.3 ng/ml, SEM; P < 0.005). Both control and enucleated fetuses presented circadian rhythm of prolactin with acrophase between 1400 and 1830 h. An enucleated fetus was tested for response of prolactin to TRH. Prolactin increased as described in the literature. There was no change in plasma concentration of cortisol, LH or GH after ocular enucleation. Our data indicate that the optical pathway participates in prolactin regulation in the fetal sheep.