Adult male rat hippocampus synthesizes estradiol from pregnenolone by cytochromes <a id="__tag_777755835" class="tag_hotlink" href="/protein/P45017" ref="/protein/P45017"><span class="tag-json" style="display:none">{"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}</span>P45017</a>α and P450 aromatase localized in neurons

Yasushi Hojo

Taka Hattori

Taihei Enami

Aizo Furukawa

William Janssen+4 authors

^{Department of Biophysics and Life Sciences and}^{Core Research for Evolutional Science and Technology Project of Japan Science and Technology Corporation, Graduate School of Arts and Sciences, University of Tokyo at Komaba, Meguro, Tokyo 153, Japan;}^{Department of Psychiatry, Tokyo Metropolitan Matsuzawa Hospital, 2-1-1 Kamikitazawa, Setagaya, Tokyo 156, Japan;}^{Kastor Neurobiology of Aging Laboratories, Fishberg Research Center for Neurobiology, Mount Sinai School of Medicine, New York, NY 10029;}^{Faculty of Integrated Arts and Sciences, Hiroshima University, Higashi-hiroshima 739, Japan; and}^{Department of Biochemistry, School of Medicine, Fujita Health University, Toyoake, Aichi 470, Japan}

^{To whom correspondence should be addressed. E-mail:}pj.ca.oykot-u.c.syhp@otawak.

Edited by Bruce S. McEwen, The Rockefeller University, New York, NY, and approved October 16, 2003

Received 2003 Mar 5

Abstract

In adult mammalian brain, occurrence of the synthesis of estradiol from endogenous cholesterol has been doubted because of the inability to detect dehydroepiandrosterone synthase, {"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}P45017α. In adult male rat hippocampal formation, significant localization was demonstrated for both cytochromes {"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}P45017α and P450 aromatase, in pyramidal neurons in the CA1–CA3 regions, as well as in the granule cells in the dentate gyrus, by means of immunohistochemical staining of slices. Only a weak immunoreaction of these P450s was observed in astrocytes and oligodendrocytes. ImmunoGold electron microscopy revealed that {"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}P45017α and P450 aromatase were localized in pre- and postsynaptic compartments as well as in the endoplasmic reticulum in principal neurons. The expression of these cytochromes was further verified by using Western blot analysis and RT-PCR. Stimulation of hippocampal neurons with N-methyl-d-aspartate induced a significant net production of estradiol. Analysis of radioactive metabolites demonstrated the conversion from [^{H]pregnenolone to [}^{H]estradiol through dehydroepiandrosterone and testosterone. This activity was abolished by the application of specific inhibitors of cytochrome P450s. Interestingly, estradiol was not significantly converted to other steroid metabolites. Taken together with our previous finding of a P450scc-containing neuronal system for pregnenolone synthesis, these results imply that 17β-estradiol is synthesized by}{"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}P45017α and P450 aromatase localized in hippocampal neurons from endogenous cholesterol. This synthesis may be regulated by a glutamate-mediated synaptic communication that evokes Ca^signals.

Abstract

The hippocampal formation, essentially involved in learning and memory processes, is known to be a target for the neuromodulatory actions of hormones produced in the gonads. As both estradiol and testosterone may reach the brain via blood circulation, and extensive studies have been performed to investigate their role in modulating hippocampal plasticity and function (1–4). Evidence is emerging that estrogen exerts not only the chronic/genomic effects but also a rapid/nongenomic influence on hippocampal synaptic plasticity (2, 5). In addition to endocrine-derived hormones, recent experiments have demonstrated that hippocampal neurons may also be exposed to locally synthesized brain neurosteroids, such as pregnenolone (PREG) and its sulfated ester (6–9). Dehydroepiandrosterone (DHEA) has been found in the mammalian brain at concentrations greater than that in plasma (6, 10). Because DHEA concentrations do not decrease after adrenalectomy and castration, many experiments have been performed with the aim of demonstrating the de novo synthesis of DHEA within the brain. DHEA biosynthesis has been demonstrated in cultured glial cells and neurons from the neonatal rat brain (11, 12). In the adult brain, however, a concrete demonstration of the synthesis of DHEA, androstenedione (AD), testosterone, or estradiol directly from endogenous cholesterol has yet to be reported. Sex steroids such as estradiol and testosterone, therefore, have not classically been considered to be “brain-derived neurosteroids.” Furthermore, the localization and activity of cytochrome {"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}P45017α (CYP17) in adult mammalian brain has also not been demonstrated, despite many sophisticated studies employing immunohistochemistry, molecular biology, and enzyme activity assays (13–16). {"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}P45017α has, therefore, been regarded to be only transiently expressed in the brain at embryonic and neonatal stages (11, 12, 14). Recently, a few studies have reported the presence of cytochrome P450 aromatase (P450arom, CYP19) in the adult rat and human hippocampal formation (17, 18).

The present study was designed to examine, in the adult male hippocampus, the cellular localization of {"type":"entrez-protein","attrs":{"text":"P45017","term_id":"1171764","term_text":"P45017"}}P45017α and P450arom and the endogenous metabolism of neurosteroids from PREG. Analyses revealed the neuronal localization of these P450s and the synthesis of DHEA, testosterone, and 17β-estradiol, which suggests their intracrine/paracrine actions on the plasticity of neurons in adult rat brain.

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Acknowledgments

We thank Dr. J. Rose for critical reading of the manuscript. We thank Novartis for the kind gift of SU-10603. This work was supported in part by National Institutes of Health Grant PO1AG16765 (to J.H.M.).

Acknowledgments

Notes

This paper was submitted directly (Track II) to the PNAS office.

Abbreviations: PREG, pregnenolone; DHEA, dehydroepiandrosterone; AD, androstenedione; P450arom, cytochrome P450 aromatase; 17β-HSD, 17β-hydroxysteroid dehydrogenase; DHT, dihydrotestosterone.

Notes

This paper was submitted directly (Track II) to the PNAS office.

Abbreviations: PREG, pregnenolone; DHEA, dehydroepiandrosterone; AD, androstenedione; P450arom, cytochrome P450 aromatase; 17β-HSD, 17β-hydroxysteroid dehydrogenase; DHT, dihydrotestosterone.

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