ORIGINAL ARTICLE Developmental Expression of Key Steroidogenic Enzymes in the Brain of Protandrous Black Porgy Fish, Acanthopagrus schlegeli S. Tomy,* G.-C. Wu,* H.-R. Huang,* S. Dufour and C.-F. Chang* *Department of Aquaculture, National Taiwan Ocean University, Keelung, Taiwan.  USM 0401, UMR 5178 CNRS ⁄ MNHN ⁄ UPMC Biologie des Organismes Marins et Ecosyste `mes, De ´partement des Milieux et Peuplements Aquatiques, Muse ´um National d’Histoire Naturelle, Paris, France. De novo steroidogenesis from cholesterol is a conserved property of vertebrate brains, and such steroids synthesised de novo in the brain are called neurosteroids (1, 2). The first indication that ster- oidogenesis occurs in the brain came from the observation that certain steroids, in particular, pregnenolone and dehydroepiandros- terone and their sulphate esters, accumulate in the brains of cas- trated and adrenalectomised mammals (3, 4). Neurosteroidogenesis begins with the conversion of cholesterol to pregnenolone, the major precursor of other neurosteroids. Cytochrome P450 side- chain cleavage enzyme (P450scc:CYP11A1) is the first and rate-lim- iting enzyme that catalyses the initial step in steroid biosynthesis, conversion of cholesterol to pregnenolone (5). During development, the CYP11A1 gene is expressed in the central nervous system (CNS) in rats and the expression persists during neonatal development Journal of Neuroendocrinology Correspondence to: Dr Ching-Fong Chang, Department of Aquaculture, National Taiwan Ocean University, Keelung 202, Taiwan (e-mail: b0044@mail.ntou.edu.tw). In the present study, we tested the hypothesis that the brain of the black porgy fish, Acantho- pagrus schlegeli, has the capacity for de novo steroidogenesis and that these neurosteroids may impact sex differentiation. Gonadal histology and Dmrt1 gene expression revealed that the fish were not sex differentiated until 155 dah (days after hatching). We further demonstrated the developmental expressions of the mRNAs encoding for four key neurosteroidogenic enzymes, namely, the cytochrome P450 side chain cleavage (CYP11A1), 3b-hydroxysteroid dehydroge- nase ⁄D 5 -D 4 isomerase (3bHSD), cytochrome P450c17 (CYP17) and aromatase (CYP19b) in the brain at different post-hatching developmental ages. The results indicated that steroidogenic genes are expressed in brain from the earliest sampling time, 60 dah. Quantitative real-time polymerase chain reaction analysis demonstrated significantly higher expression levels of these enzymes at 120 dah compared to 60 dah in all the brain regions. However, the increase for 3bHSD was significant only in hypothalamus and midbrain, whereas it was significant only in forebrain and hypothalamus for CYP19b. A decline in mRNA levels were observed for all the genes at 155 dah except in midbrain for CYP11A1 and in hindbrain for CYP19b. Analysis of aro- matase enzyme activity showed a significant increase in aromatase activity in the forebrain at 120 dah. Thus, the present study demonstrated for the first time an age- and ⁄ or region depend- ent expression of the mRNAs encoding the steroidogenic enzyme genes in the brain of black porgy. The presence of key steroidogenic enzymes as early as 60 dah, before gonadal sex differ- entiation, demonstrates that steroid biosynthetic capacity in brain precedes histological gonad differentiation. The mRNA transcripts of these genes showed a synchronous peak at 120 dah, suggesting that oestradiol may be locally formed in most parts of the brain. The study suggests an important role for brain aromatase in male black porgy brain sex differentiation, and consid- ers the possibility of a role for this enzyme in neurogenesis. Key words: 3bHSD, brain, CYP11A1, CYP17, CYP19b, development, fish, sex differentiation, stero- idogenic enzymes. doi: 10.1111/j.1365-2826.2007.01572.x Journal of Neuroendocrinology 19, 643–655 ª 2007 The Authors. Journal Compilation ª 2007 Blackwell Publishing Ltd