脳におけるホルモン代謝 第1編 ラット脳における⊿(4)-3-ketosteroid 5α-oxidoreductaseの存在とそれに及ぼす中枢作用薬の影響
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It is well known that testosterone, estradiol and progesterone have an influence on sexual behaviors in mammals. On the other hand, sex hormones may be related to mental excitation in men, and also psychic aberration is often occur during puberty, menstrual cycles and pregnancy period. In order to pursue the role of sex hormones in mental function, I tried first to examine the metabolism of testosterone in the central nervous system and the effect of central acting drugs on this metabolism. Methods: 0.2μCi of 4-(14)C-testosterone (specific activity 60mCi/mM), 1 ml of rat diencephalon homogenate (about 8 mg protein) and NADPH (95μM/L) were mixed together and the total volume was adjusted to 2.5 ml with phosphate buffer (pH 6.8). The reaction mixture was incubated at 38℃ for 120 min with continual shaking; the reaction contents were exposed to air. A sample of enzyme preparation placed in a boiling water bath for 5 min was used as blank. Following incubation, the metabolite formed was extracted twice with 10 ml of chloroform: ether mixture (3:1), and the pooled extract was washed once with distilled water, being dehydrated with anhydrus sodium sulphate and evaporated to dryness. Following florisil column chromatography, the steroids absorbed were eluted with 25 ml of 2 % methanol-chloroform and evaporated to dryness. This fraction was spotted on a TLC plate (silicage G) and developed in chloroform: ether (3:1). Each fractions was identified by autoradiography. The fractions corresponding to DHT and 5α-androstan-3α, 17β-diol were eluted and the solvent was removed off under N(2) gas. These were used as the samples for identification of the metabolites. Results : 1) Identification of the metabolites was performed by TLC, GLC and estimation of specific activity. DHT and 5α-androstan-3α, 17β-diol were confirmed as principal metabolites. 2) The occurrance of ⊿(4)-3-ketosteroid 5α-oxidoreductase was confirmed in rat diencephalon. The characteristics of the enzyme were as follows; optimal pH is 6.8 and apparent Km value is 2.2 x 10(-6)M. The enzyme showed the highest activity in crude microsomal fraction from rat diencephalon, and the enzyme activity was inhibited by progesterone but was unaffected by estradiol. 3) Male Wistar rats weighing about 100 g were divided into four groups. Each group received chlorpromazine HCl (0.4mg/0.1ml/100g body weight), sodium diphenylhydantoin (0.5mg/0.1ml/100g body weight), imipramine HCl (0.1mg/0.1ml/100g body weight) or normal saline (0.1ml/100g body weight) intraperitoneally for 14 consecutive days respectively. 24 hours following the last administration of the drug, the rats were killed by decapitation. Diencephalon was removed and homogenated with 9 volumes of phosphate buffer (pH 6.8). Homogenates were incubated with testosterone and NADPH for the estimation of the ⊿4 -3-ketosteroid 5α-oxidoreductase activity. All the drugs examined in this study decreased the enzyme activity significantly.
- 1977-02-28
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