Steroid hormone action during brain development exerts profound effects on reproductive physiology and behavior that last into adulthood. A variety of in vitro studies indicate that steroid receptors require nuclear receptor coactivators for efficient transcriptional activity. To determine the functional significance of the nuclear receptor coactivator SRC-1 in developing brain, we investigated the consequence of reducing SRC-1 protein during sexual differentiation of the brain. We report that reducing SRC-1 protein interferes with the defeminizing actions of estrogen in neonatal rat brain. Our data indicate that SRC-1 protein expression is critically involved in the hormone-dependent development of normal male reproductive behavior and brain morphology.