Variation in anogenital distance (AGD) in female newborn rats depends upon androgenization secondary to transplacental/transmembraneous testosterone from adjacent intrauterine male siblings. Since the size of the sexually dimorphic nucleus of the preoptic area of the hypothalamus (SDN-POA) and the degree of pituitary sensitivity to GnRH are neuroendocrine markers of neonatal androgenization, we compared these to AGD in castrated adult male and female rats. Compared to 1-day-old female rats with short AGD (less than or equal to 1.4 mm), 1-day-old female rats with long AGD (greater than 1.4 mm) had significantly larger SDN-POA volumes as adults. In contrast, LH secretion following GnRH injection did not differ in the two subgroups. Our results emphasize that some endpoints of central nervous system sexual differentiation in the adult rat are predicted by the appearance of a masculinized genital tract at birth. It follows that the complete evaluation of potential androgenizing agents will require systematic assessment of multiple morphologic and functional endpoints.