Activity and localization of 3β-hydroxysteroid dehydrogenase/δ54-isomerase in the zebrafish central nervous system

Hirotaka Sakamoto, Kazuyoshi Ukena, Kazuyoshi Tsutsui

Research output: Contribution to journalArticle

75 Citations (Scopus)

Abstract

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 3β-hydroxysteroid dehydrogenase/Δ54-isomerase (3βHSD), 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 3βHSD. This conversion was significantly reduced by trilostane, a specific inhibitor of 3βHSD. By using Western immunoblotting with the polyclonal antiserum directed against purified bovine adrenal 3βHSD, a 3βHSD-like substance was found in homogenates of the zebrafish brain. Immunocytochemical analysis was then undertaken to investigate the localization of the 3βHSD-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). 3βHSD-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 3βHSD-like immunoreactivity. These results suggest that the fish CNS possesses steroidogenic enzyme 3βHSD and produces progesterone. The present study further provides the first immunocytochemical mapping of the site of 3βHSD expression in the fish CNS.

Original languageEnglish
Pages (from-to)291-305
Number of pages15
JournalJournal of Comparative Neurology
Volume439
Issue number3
DOIs
Publication statusPublished - 2001 Oct 22
Externally publishedYes

Fingerprint

3-Hydroxysteroid Dehydrogenases
Isomerases
Zebrafish
Central Nervous System
Progesterone
Brain
Immune Sera
Spinal Cord
Fishes
Posterior Thalamic Nuclei
Pregnenolone
Telencephalon
Preoptic Area
Paraventricular Hypothalamic Nucleus
Purkinje Cells
Carisoprodol
Enzymes
Microsomes
Vertebrates
Retina

Keywords

  • Biochemical analysis
  • Brain
  • Immunocytochemistry
  • Neurosteroids
  • Teleost
  • Western immunoblotting

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Activity and localization of 3β-hydroxysteroid dehydrogenase/δ54-isomerase in the zebrafish central nervous system. / Sakamoto, Hirotaka; Ukena, Kazuyoshi; Tsutsui, Kazuyoshi.

In: Journal of Comparative Neurology, Vol. 439, No. 3, 22.10.2001, p. 291-305.

Research output: Contribution to journalArticle

@article{3033d73c8ef14aad902182bb3b9be172,
title = "Activity and localization of 3β-hydroxysteroid dehydrogenase/δ5-δ4-isomerase in the zebrafish central nervous system",
abstract = "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 3β-hydroxysteroid dehydrogenase/Δ5-Δ4-isomerase (3βHSD), 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 3βHSD. This conversion was significantly reduced by trilostane, a specific inhibitor of 3βHSD. By using Western immunoblotting with the polyclonal antiserum directed against purified bovine adrenal 3βHSD, a 3βHSD-like substance was found in homogenates of the zebrafish brain. Immunocytochemical analysis was then undertaken to investigate the localization of the 3βHSD-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). 3βHSD-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 3βHSD-like immunoreactivity. These results suggest that the fish CNS possesses steroidogenic enzyme 3βHSD and produces progesterone. The present study further provides the first immunocytochemical mapping of the site of 3βHSD expression in the fish CNS.",
keywords = "Biochemical analysis, Brain, Immunocytochemistry, Neurosteroids, Teleost, Western immunoblotting",
author = "Hirotaka Sakamoto and Kazuyoshi Ukena and Kazuyoshi Tsutsui",
year = "2001",
month = "10",
day = "22",
doi = "10.1002/cne.1351",
language = "English",
volume = "439",
pages = "291--305",
journal = "Journal of Comparative Neurology",
issn = "0021-9967",
publisher = "Wiley-Liss Inc.",
number = "3",

}

TY - JOUR

T1 - Activity and localization of 3β-hydroxysteroid dehydrogenase/δ5-δ4-isomerase in the zebrafish central nervous system

AU - Sakamoto, Hirotaka

AU - Ukena, Kazuyoshi

AU - Tsutsui, Kazuyoshi

PY - 2001/10/22

Y1 - 2001/10/22

N2 - 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 3β-hydroxysteroid dehydrogenase/Δ5-Δ4-isomerase (3βHSD), 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 3βHSD. This conversion was significantly reduced by trilostane, a specific inhibitor of 3βHSD. By using Western immunoblotting with the polyclonal antiserum directed against purified bovine adrenal 3βHSD, a 3βHSD-like substance was found in homogenates of the zebrafish brain. Immunocytochemical analysis was then undertaken to investigate the localization of the 3βHSD-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). 3βHSD-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 3βHSD-like immunoreactivity. These results suggest that the fish CNS possesses steroidogenic enzyme 3βHSD and produces progesterone. The present study further provides the first immunocytochemical mapping of the site of 3βHSD expression in the fish CNS.

AB - 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 3β-hydroxysteroid dehydrogenase/Δ5-Δ4-isomerase (3βHSD), 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 3βHSD. This conversion was significantly reduced by trilostane, a specific inhibitor of 3βHSD. By using Western immunoblotting with the polyclonal antiserum directed against purified bovine adrenal 3βHSD, a 3βHSD-like substance was found in homogenates of the zebrafish brain. Immunocytochemical analysis was then undertaken to investigate the localization of the 3βHSD-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). 3βHSD-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 3βHSD-like immunoreactivity. These results suggest that the fish CNS possesses steroidogenic enzyme 3βHSD and produces progesterone. The present study further provides the first immunocytochemical mapping of the site of 3βHSD expression in the fish CNS.

KW - Biochemical analysis

KW - Brain

KW - Immunocytochemistry

KW - Neurosteroids

KW - Teleost

KW - Western immunoblotting

UR - http://www.scopus.com/inward/record.url?scp=0035934738&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0035934738&partnerID=8YFLogxK

U2 - 10.1002/cne.1351

DO - 10.1002/cne.1351

M3 - Article

VL - 439

SP - 291

EP - 305

JO - Journal of Comparative Neurology

JF - Journal of Comparative Neurology

SN - 0021-9967

IS - 3

ER -