Immunocytochemical localization and biological activity of hydroxysteroid sulfotransferase in the frog brain

Biosynthesis of the neuroactive steroids pregnenolone sulfate (Delta(5)PS) and dehydroepiandrosterone sulfate (DHEAS) is catalyzed by the enzyme hydroxysteroid sulfotransferase (HST), which transfers the sulfonate moiety from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) on the 3-hydroxy site of steroids. Although high concentrations of Delta(5)PS and DHEAS have been detected in the rat brain, the anatomical localization of HST in the CNS has never been determined. Using an antiserum against rat liver HST, we have investigated the distribution of HST-like immunoreactivity in the CNS of the frog Rana ridibunda. Two populations of HST-immunoreactive neurons were observed in the hypothalamus, and several bundles of positive nerve fibers were visualized in the telencephalon and diencephalon. Incubation of frog brain homogenates with [S-35]PAPS and [H-3]pregnenolone yielded the formation of several H-3,S-35-labeled compounds, including Delta(5)PS and testosterone sulfate. When [H-3]dehydroepiandrosterone and [S-35]PAPS were used as precursors, one of the H-3,S-35-labeled metabolites coeluted with DHEAS. Neosynthesis of [H-3]Delta(5)PS and [H-3]DHEAS was reduced significantly by 2,4-dichloro-6-nitrophenol, a specific inhibitor of sulfotransferases. The present study provides the first immunocytochemical mapping of HST in the brain. Our data also demonstrate for the first time that biosynthesis of the highly potent neuroactive steroids Delta(5)PS and DHEAS occurs in the CNS of nonmammalian vertebrates.