Activity and localization of 3β-hydroxysteroid dehydrogenase/Δ5-Δ4-isomerase in the zebrafish central nervous system

Little information is available for neurosteroidogenesis in the central nervous system (CNS) of lower vertebrates. Therefore, in the present study, we examined the enzymatic activity and localization of 3 beta -hydroxysteroid dehydrogenase/Delta (5)-Delta (4)-isomerase (3 beta HSD), a key steroidogenic enzyme, in the CNS of adult male zebrafish to clarify central progesterone biosynthesis. Biochemical studies together with HPLC analysis revealed that the zebrafish brain converted pregnenolone to progesterone, suggesting the enzymatic activity of 3 beta HSD. This conversion was significantly reduced by trilostane, a specific inhibitor of 3 beta HSD. By using Western immunoblotting with the polyclonal. antiserum. directed against purified bovine adrenal 3 beta HSD, a 3 beta HSD-like substance was found in homogenates of the zebrafish brain. Immunocytochemical analysis was then undertaken to investigate the localization of the 3 beta HSD-like substance in the zebrafish brain and spinal cord. Clusters of immunoreactive cell bodies were localized in the dorsal telencephalic. areas (D), central posterior thalamic nucleus (CP), preoptic nuclei (NPO), posterior tuberal nucleus (PTN), paraventricular organ (PVO), and nucleus of medial longitudinal fascicle (NMLF). 3 beta HSD-like immunoreactivity was also observed in somata of cerebellar Purkinje neurons. A widespread distribution of immunoreactive fibers was found throughout the brain and spinal cord. In addition, positively stained cells were restricted to other organs, such as the pituitary and retina. Preabsorbing the antiserum with purified bovine adrenal microsome resulted in a complete absence of 3 beta HSD-like immunoreactivity. These results suggest that the fish CNS possesses steroidogenic enzyme 3 beta HSD and produces progesterone. The present study further provides the first immunocytochemical mapping of the site of 3 beta HSD expression in the fish CNS. (C) 2001 Wiley-Liss, Inc.