The distribution of neurons which concentrate testosterone o r i t s metabolites in the brain of 2-day old female r a t s was determined using autoradiography. A specific topographic pattern of neurons which concent r a t e testosterone o r i t s metabolites was obtained similar, b u t not ident i c a l , t o the one found in adult male r a t s . The demonstrated s i t e s of nuclear uptake and retention of testosterone o r i t s metabolites by specific neurons in the neonatal brain probably provide the anatomical substrate f o r sexual d i f f e r e n t i a t i o n of the brain.The presence of androgen has repeatedly been shown t o have irreversible e f f e c t s on the developing central nervous system (CNS) of the r a t (Gorski, ' 7 1 ) .These e f f e c t s include permanent a1 terations in gonadotropin secretion (Gorski , ' 7 1 ) , the number of dendritic spines in the preoptic region of the brain (Raisman and Field, ' 7 3 ) , the number of dense core vesicles in the neurons of the arcuate nucleus (Ratner and Adamo, ' 7 1 ) , and sexual behavior (Beach, '68). However, attempts t o demonstrate a preferential uptake of radioactivity a f t e r 3H-testosterone by the neonatal r a t brain before day 10 have f a i l e d (Sheratt, Exley and Rogers, '69; Alvarez and Ramirez, '70; Kincl, '70; Weisz and Philpott, '71; Lob1 and Schapiro, '72; Barnea, Weinstein and Linder, '72; Chamberlain and Rogers, '72; Heffner and Van Tienhoven, '73; Vertes e t a l . , '73). Since autoradiography had proven t o be successful in demonstrating selective uptake and retention of testosterone o r i t s metabolites in specific areas of the CNS of the adult male r a t before biochemical techniques were successful (Sar and Stumpf , '72), and 589