Trace charactethation of neurosteroids to develop a new antipsychotic agent targeting neurosteroidogenesis

神经甾体与γ-氨基丁酸A型（GABA_A）受体具有高亲和力结合，正性调节GABA对这些受体的作用，从而产生显著的麻醉、镇静和抗焦虑作用。脑和血清中内源性神经甾体的定量分析对于表征其生理功能和影响脑功能的机制具有重要意义。脑和血清中的神经甾体测定也有助于开发新的抗精神病药物靶向神经甾体合成。基于此背景信息，开发并验证了灵敏的液相色谱-串联质谱方法，该方法能够定量测定大鼠脑和血清中的痕量神经甾体（5α-还原型甾烷神经甾体、3-氧代-4-烯-神经甾体、神经活性雄激素和四氢皮质酮异构体）。有些方法采用衍生化程序，这是非常有效的增加， ...更多信息 使用所开发的方法从动物研究中获得的新发现如下。1)大脑别孕烯醇酮、表别孕烯醇酮和5α-二氢孕酮水平通过制动应激和乙醇给药迅速升高。Epiallopregnanolone是脑特异性产物。2)压力引起的脑睾酮和雄烯二酮（雄甾烷类类固醇）水平的变化远低于孕酮及其代谢产物（甾烷类类固醇）。3)大部分脑睾酮来自外周器官，而相当数量的5α-二氢睾酮在脑中持续合成。GABA能神经活性雄激素5α-雄甾烷-3 α，17 β-二醇（5α-androstane-3α，17 β-diol）始终存在于雄性大鼠脑内，其水平不受应激的影响。4)束缚应激后，雌雄大鼠脑内均存在3α，5 α-四氢皮质酮，但其脑内含量存在性别差异。少

Neurosteroids bind to the γ-aminobutyric acid type A (GABA_A) receptors with a high affinity, positively modulate the action of GABA at these receptors and thus elicit marked anesthetic, sedative and anxiolytic effects. The quantitative analysis of the endogenous neurosteroids in the brain and serum is very important for characterization of their physiological functions and the mechanism by which they affect the brain functions. The brain and serum assays of the neurosteroids can also contribute to the development of new antipsychotic agents targeting neurosteroidogenesis. Based on this background information, sensitive liquid chromatography-tandem mass spectrometric methods that enable the quantification of trace neurosteroids (5α-redduced pregnane neurosteroids, 3-oxo-4-ene-neurosteroids, neuroavtive androgens and tetrahydrocorticosterone isomers) in rat brain and serum were developed and validated. Some methods employed derivatization procedures, which were very effective in increas … More ing the detection responses of steroids.New discoveries from the animal studies using the developed methods were as follows. 1) The brain allopregnanolone, epiallopregnanolone and 5α-dihydroprogesterone levels are rapidly elevated by immobilization stress and ethanol administration. Epiallopregnanolone is the brain-specific product. 2) The stress-induced level changes in the brain testosterone and androstenedione (androstane steroids) are much lower than those of progesterone and its metabolites (pregnane steroids). 3) Most of the brain testosterone is derived from the peripheral organs, while a fair amount of 5α-dihydrotestosterone is continuously synthesized in the brain. The GABAergic neuroactive androgen, 5α-androstane-3α,17β-diol, is always present in the male rat brain and its level is not influenced by the stress. 4) 3α,5α-Tetrahydrocorticosterone is present in the brain of both male and female rats after being subjected to the immobilization stress, but there is a sex difference in its brain level. Less