During the neonatal period in male macaques, the testis produces adult-like levels of plasma testosterone (T), but the function of this in development is not understood. To investigate the interaction of T with the neonatal brain, 4 male and 5 female cynomolgus monkeys were gonadectomized 2-5 days after birth, and were injected subcutaneously 3 days later with 500 microCi [3H]-testosterone ([3H]-T). 60 min later, brains and other tissue samples were removed. Purified nuclear pellets were prepared by centrifugation through 2 M sucrose, extracted into ether and analyzed by high-performance liquid chromatography. The aromatized metabolite, [3H]-estradiol [( 3H]-E2), was found only in the hypothalamus (HYP) and amygdala (AMG). In HYP, [3H]-E2 represented 55 +/- 3% of the radioactivity in males and 53 +/- 3% in females. In AMG, [3H]-E2 represented 40 +/- 9% of the radioactivity in males and 47 +/- 3% in females. Concentrations of unchanged [3H]-T were higher than those of [3H]-dihydrotestosterone [( 3H]-DHT). Both androgens were present in nuclear pellets from all 8 brain regions studied, and concentrations were significantly higher in females than in males (p less than 0.005). [3H]-T was also the main form of radioactivity in nuclear pellets from pituitary gland, adrenal gland, uterus and liver, but very high levels of [3H]-DHT were found in seminal vesicles, prostate and penis. Comparisons were made with previous results from orchidectomized fetuses at 122 days gestation and from fully adult male castrates, and the largest developmental changes occurred in the AMG where concentrations of [3H]-E2 were 20-fold higher in adults than in fetuses, and most of this increase took place after the neonatal stage. Nuclear concentrations of [3H]-T also increased markedly during development in most brain regions except the cerebellar cortex where they declined.