Tsutsui K, Yamazaki T
Faculty of Integrated Arts and Sciences, Hiroshima University, Japan.
Brain Res. 1995 Apr 24;678(1-2):1-9. doi: 10.1016/0006-8993(95)00116-8.
The present study was carried out to determine steroid biosynthesis from cholesterol in the brain of adult male Japanese quails. As an initial step of the experiments, the concentrations of pregnenolone, dehydroepiandrosterone and their sulfate esters in the brain and plasma extracts were measured by specific radioimmunoassays (RIAs). To exclude the possibility that these steroids in the brain are derived from peripheral steroidogenic glands, hypophysectomized and sham-operated birds were used in this experiment. The pregnenolone concentration was much larger in the brain than in the plasma in these two groups. Hypophysectomy led to decreases in the plasma and brain pregnenolone concentrations, but the change in the brain was less pronounced than that in the plasma. Although pregnenolone sulfate ester, dehydroepiandrosterone and its sulfate ester were also detected in brain extracts, those levels were low in both hypophysectomized and sham-operated birds. The biochemical demonstration of cholesterol metabolism was further conducted in intact mitochondria. When mitochondrial fractions obtained from the whole brain were incubated with cholesterol at 37 degrees C, the pregnenolone level in mitochondria increased as a function of incubation time. Finally, Western immunoblot analysis using a purified IgG with polyclonal antibodies against the purified bovine adrenal cytochrome P450scc was performed after SDS-gel electrophoresis of homogenates of the hypothalamus and preoptic area. A protein recognized the antibody as a band of electrophoretic mobility in the proximity of reference bovine P450scc. These results suggest that the brain produces pregnenolone from cholesterol in adult male Japanese quails. Most accumulation of pregnenolone in the brain may be independent of the peripheral endocrine gland system.
本研究旨在确定成年雄性日本鹌鹑大脑中胆固醇的类固醇生物合成。作为实验的第一步，通过特异性放射免疫分析（RIA）测量大脑和血浆提取物中孕烯醇酮、脱氢表雄酮及其硫酸酯的浓度。为排除大脑中这些类固醇来自外周类固醇生成腺体的可能性，本实验使用了垂体切除和假手术的鸟类。在这两组中，大脑中的孕烯醇酮浓度远高于血浆中的浓度。垂体切除导致血浆和大脑中孕烯醇酮浓度降低，但大脑中的变化不如血浆中明显。虽然在大脑提取物中也检测到了硫酸孕烯醇酮、脱氢表雄酮及其硫酸酯，但在垂体切除和假手术的鸟类中，这些水平都很低。在完整的线粒体中进一步进行了胆固醇代谢的生化证明。当将从全脑获得的线粒体部分与胆固醇在37℃下孵育时，线粒体中的孕烯醇酮水平随孵育时间而增加。最后，在下丘脑和视前区匀浆进行SDS凝胶电泳后，使用针对纯化的牛肾上腺细胞色素P450scc的多克隆抗体的纯化IgG进行Western免疫印迹分析。一种蛋白质被该抗体识别为电泳迁移率与参考牛P450scc接近的条带。这些结果表明，成年雄性日本鹌鹑的大脑可从胆固醇产生孕烯醇酮。大脑中孕烯醇酮的大部分积累可能独立于外周内分泌腺系统。
