Activity and localization of 3 beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4)-isomerase in the zebrafish central nervous system

Little information is available for neurosteroidogenesis in the central ner vous 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 steroidoge nic enzyme, in the CNS of adult male zebrafish to clarify central progester one 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 re duced by trilostane, a specific inhibitor of 3 beta HSD. By using Western i mmunoblotting with the polyclonal. antiserum. directed against purified bov ine adrenal 3 beta HSD, a 3 beta HSD-like substance was found in homogenate s of the zebrafish brain. Immunocytochemical analysis was then undertaken t o investigate the localization of the 3 beta HSD-like substance in the zebr afish brain and spinal cord. Clusters of immunoreactive cell bodies were lo calized in the dorsal telencephalic. areas (D), central posterior thalamic nucleus (CP), preoptic nuclei (NPO), posterior tuberal nucleus (PTN), parav entricular 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 f ound throughout the brain and spinal cord. In addition, positively stained cells were restricted to other organs, such as the pituitary and retina. Pr eabsorbing the antiserum with purified bovine adrenal microsome resulted in a complete absence of 3 beta HSD-like immunoreactivity. These results sugg est that the fish CNS possesses steroidogenic enzyme 3 beta HSD and produce s progesterone. The present study further provides the first immunocytochem ical mapping of the site of 3 beta HSD expression in the fish CNS. (C) 2001 Wiley-Liss, Inc.