Simultaneous quantification of GABAergic 3α,5α/3α,5β neuroactive steroids in human and rat serum

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• 作者：Patrizia Porcu ; Todd K. O’ ; Buckley ; Sarah E. Alward ; Christine E. Marx ; Lawrence J. Shampine ; Susan S. Girdler ; A. Leslie Morrow
• 关键词：GABAergic neuroactive neurosteroid ; Pregnenolone ; Gas chromatography– ; mass spectrometry
• 刊名：Steroids
• 出版年：2009
• 出版时间：April-May 2009
• 年：2009
• 卷：74
• 期：4-5
• 页码：463-473
• 全文大小：457 K

文摘

The 3α,5α- and 3α,5β-reduced derivatives of progesterone, deoxycorticosterone, dehydroepiandrosterone and testosterone enhance GABAergic neurotransmission and produce inhibitory neurobehavioral and anti-inflammatory effects. Despite substantial information on the progesterone derivative (3α,5α)-3-hydroxypregnan-20-one (3α,5α-THP, allopregnanolone), the physiological significance of the other endogenous GABAergic neuroactive steroids has remained elusive. Here, we describe the validation of a method using gas chromatography–mass spectrometry to simultaneously identify serum levels of the eight 3α,5α- and 3α,5β-reduced derivatives of progesterone, deoxycorticosterone, dehydroepiandrosterone and testosterone. The method shows specificity, sensitivity and enhanced throughput compared to other methods already available for neuroactive steroid quantification. Administration of pregnenolone to rats and progesterone to women produced selective effects on the 3α,5α- and 3α,5β-reduced neuroactive steroids, indicating differential regulation of their biosynthetic pathways. Pregnenolone administration increased serum levels of 3α,5α-THP (+1488 % , p < 0.001), (3α,5α)-3,21-dihydroxypregnan-20-one (3α,5α-THDOC, +205 % , p < 0.01), (3α,5α)-3-hydroxyandrostan-17-one (3α,5α-A, +216 % , p < 0.001), (3α,5α,17β)-androstane-3,17-diol (3α,5α-A-diol, +190 % , p < 0.01). (3α,5β)-3-hydroxypregnan-20-one (3α,5β-THP) and (3α,5β)-3-hydroxyandrostan-17-one (3α,5β-A) were not altered, while (3α,5β)-3,21-dihydroxypregnan-20-one (3α,5β-THDOC) and (3α,5β,17β)-androstane-3,17-diol (3α,5β-A-diol) were increased from undetectable levels to 271 ± 100 and 2.4 ± 0.9 pg ± SEM, respectively (5/8 rats). Progesterone administration increased serum levels of 3α,5α-THP (+1806 % , p < 0.0001), 3α,5β-THP (+575 % , p < 0.001), 3α,5α-THDOC (+309 % , p < 0.001). 3α,5β-THDOC levels were increased by 307 % , although this increase was not significant because this steroid was detected only in 3/16 control subjects. Levels of 3α,5α-A, 3α,5β-A and pregnenolone were not altered. This method can be used to investigate the physiological and pathological role of neuroactive steroids and to develop biomarkers and new therapeutics for neurological and psychiatric disorders.