This study demonstrates that in the rat there are specific patterns of in vitro phosphorylation of cytosolic proteins for the preoptic-hypothalamus, cortex, and amygdala. Furthermore, there are sex-specific patterns for the male and female preoptic-hypothalamus. These sex-specific patterns are controlled by the sex steroid environment of the neonatal rat. If testosterone was removed by neonatal castration of a male, the female pattern of in vitro phosphorylated proteins was found in the adult preoptic-hypothalamus. Conversely, if a neonatal female was androgenized at 2 days by a single injection of testosterone, a male-like pattern was found in the adult preoptic-hypothalamus. Treatment of neonatal females or males did not alter the adult patterns of in vitro protein phosphorylation in the cortex, but such treatments did give rise to anomalous patterns in the amygdala. Thus, as with behavioral and structural characteristics of the preoptic-hypothalamus, the spectrum of phosphorylated proteins in this region also seems to be organized by neonatal testosterone.