Neurosteroids are steroids that are synthesized de novo in the brain from cholesterol and, in general, mediate their effects through ion-gated channel receptors such as γ-aminobutyric acid(A) (GABA(A)) and N-methyl-D- aspartate receptors rather than through classical nuclear steroid hormone receptors. Steroid hormones are known to exist not only as free compounds, but also as sulfated derivatives. Pharmacological studies indicate that unconjugated and sulfated steroids, such as pregnenolone and pregnenolone sulfate, may have opposite effects on GABA(A) receptors. Thus, pregnenolone acts as a potent positive allosteric modulator of γ-aminobutyric acid action at GABA(A) receptors, whereas pregnenolone sulfate acts as a potent negative modulator. Recent experiments also suggest that dehydroepiandrosterone and dehydroepiandrosterone sulfate may have distinct effects on growth of neurites from embryonic neocortical neurons in vitro. Thus, regulation of steroid sulfation may have profound behavioral and morphological effects on the nervous system. We, therefore, studied the developmental expression of the enzyme steroid sulfatase (STS), which converts sulfated steroids to free steroids. By in situ hybridization, STS messenger RNA was expressed in the embryonic mouse cortex, hindbrain, and thalamus during the last third of gestation. The sites of expression of STS were similar to those of P450c17, suggesting that these two enzymes may have concerted actions in similar functional processes.