A fetoneonatal estradiol-binding plasma protein, often referred to as alpha fetoprotein (AFP), is present in high levels in serum of neonatal rats. The existence of elevated serum FSH levels, in spite of high estrogen titers and the presence of adult type estrogen receptors in the hypothalamus and pituitary, suggests that the lack of a fully expressed estrogen negative feedback in these animals is not due to a decreased function of the receptors, but rather to binding of estrogen to AFP. To evaluate this hypothesis, rats were ovariectomized at day 10 and injected sc daily for 3 days with different doses of estradiol benzoate (Eb), for which AFP has a very high binding affinity, diethylstilbestrol (DES), which binds very weakly to AFP, or ll-β-methoxyethynylestradiol (RU2858), which does not bind to AFP. Only the largest doses of EB (0.05 and 0.10 μg/100 g BW) were effective in preventing the postcastration rise in FSH. The smallest dose (0.0025 μg/100 g BW) of DES significantly reduced the postcastration rise in FSH while larger doses brought FSH levels below those of intact sham-operated controls. Even the smallest dose (0.0025 μg/100 g BW) of RU2858 was effective in reducing plasma FSH to less than half of that of control rats. To study estrogen positive feedback, intact female rats received a sc injection of Eb, DES, or RU2858 in different doses on the morning of day 10, followed by a 2nd injection 2 days later. The animals were killed at various times after the 1st and/or 2nd injection. As a control of the effectiveness of this procedure, 26 day old animals received an identical Eb treatment. No estrogen positive feedback on gonadotropin release was observed in infantile rats at any time studied regardless of dose or type of estrogen given. Moreover, plasma FSH was reduced following estrogen treatment. Plasma prolactin was not altered by Eb but was slightly increased by DES and further increased by RU2858. A marked increase in plasma LH, prolactin, and to a lesser extent FSH was elicited by Eb in 26 day old rats. The results are consistent with the concept that although the infantile hypothalamic-pituitary unit is capable of sensing changes in estrogen levels, the manifestation of estrogen negative feedback is prevented by the presence of AFP. By contrast, immaturity of a central mechanism appears to be responsible for the failure of estrogen positive feedback to operate in neonatal rats. © 1977 by The Endocrine Society.