The present study was designed to investigate the relationship between fetal sheep pituitary oestrogen receptor (ER) α expression and changes in fetal and maternal plasma 17β-oestradiol (E2) concentrations during gestation. The results revealed that immunoreactivity for ERα was located in the nuclei and distributed throughout the fetal pituitary gland during gestation. The percentage of ERα-positive cells was approximately 2% of the total cell population in female fetuses at Day 60 of gestation, increased to approximately 7% and 13% of the total cell population at Days 90 and 120 of gestation, respectively, and then declined to approximately 10% at birth. The fetal plasma E2 concentrations were approximately 19 and 71 pg mL^-1 at Days 90 and 120 of gestation, respectively, and decreased to 22 pg mL^-1 after birth. In male fetuses, plasma E2 concentrations and the percentage of ERα-positive cells were similar to values in female fetuses throughout gestation, except on Day 120 when the plasma E2 level in female fetuses was significantly higher than in male fetuses. These data demonstrate that changes in the percentage of fetal pituitary ERα-positive cells parallel fetal plasma E2 concentrations throughout gestation.