TWO TYPES OF MONOAMINE OXIDASE

描述：本项目的目标是了解 单胺氧化酶A和B的调节及其在体内的作用， 这些同工酶对单胺的降解很重要 神经递质和生物胺。MAO活动的异常水平 与许多精神障碍有关。更好地理解 这些同工酶将允许更有效的发展 精神障碍的治疗。具体目标概述如下： 目的：研究MAO A和MAO B在神经递质代谢中的作用。 双基因敲除小鼠MAO A和B(A/B)双基因敲除小鼠的行为 将通过以下途径灭活MAO KO小鼠的MAO A基因而产生 同源重组。聚合酶链式反应，Southern印迹分析，MAO催化 将使用活性和蛋白质印迹分析来确保MAO A 和B在这些小鼠中是缺乏的。大脑水平(整个大脑和区域) 5-羟色胺、去甲肾上腺素和多巴胺(MAO A底物)及其 MAO A/B双KO和野生型小鼠体内代谢物的测定 高压液相色谱分析。人的大脑水平 神经调节剂B-苯乙胺(MAO B底物)将通过 气相色谱-质谱仪(GC-MS)。这些级别将是 与MAO A和MAO B的催化活性相关。攻击性行为将是 雄性MAO A/B双KO小鼠的分析及其与脑的相关性 神经递质水平。 II.探讨F、M和Sp1因子在调节细胞周期中的作用 MAO B基因表达。因子F和因子M所需的基本DNA碱基 结合将通过定点突变和每个基因的作用来确定 MAO B基因表达中的因子将通过凝胶延迟和 启动子活性分析。利用紫外光交联实验，我们将 确定因子是由单一多肽还是由多个多肽组成。 将克隆编码因子F和M的全长cDNA，并将其 功能有效性将通过凝胶滞留分析来确定 表达的蛋白质。编码因子F、M和Sp1的cDNA将是 转基因及表达因子对MAO B体内表达的影响 将通过测定启动子活性、MAO B mRNA水平、 蛋白质水平和催化活性。 这些研究对基础神经药理学和临床都具有重要意义。 研究。

DESCRIPTION: The objectives of this project are to understand the regulation and the in-vivo function of monoamine oxidase (MAO) A and B, which are isoenzymes important for the degradation of monoamine neurotransmitters and biogenic amines. Abnormal levels of MAO activity have been associated with a number of mental disorders. A better understanding of these isoenzymes will allow for the development of more effective treatments for mental disorders. The specific aims are outlined below: To study the function of both MAO A and B in neurotransmitter metabolism and behavior using double knock-out (KO) mice MAO A and B (A/B) double KO mice will be generated by inactivating the MAO A gene of MAO KO mice via homologous recombination. PCR, Southern blot analysis, MAO catalytic activity and Western blot analysis will be used to ensure that both MAO A and B are deficient in these mice. Brain levels (whole brain and regions) of serotonin, norepinephrine and dopamine (MAO A substrates) and their metabolites will be determined in MAO A/B double KO and wild type mice by high pressure liquid chromatography (HPLC). Brain levels of the neuromodulator B-phenylethylamine (MAO B substrate) will be determined by Gas Chromatography/Mass Spectrometry (GC-MS). These levels will be correlated with MAO A and B catalytic activity. Aggressive behavior will be analyzed in male MAO A/B double KO mice and correlated with brain neurotransmitter levels. II. To investigate the role of factors F, M and Sp1 in the regulation of MAO B gene expression. The essential DNA bases required for factors F and M binding will be determined by site-directed mutagenesis and the role of each factor in MAO B gene expression will be identified by gel retardation and promoter activity assays. Using UV crosslinking experiments we will determine if the factors comprise of a single or multiple polypeptides. Full-length cDNAs encoding factors F and M will be cloned and their functional validity will be determined by gel retardation assays with expressed proteins. The cDNA encoding factor F, M and Sp1 will be transfected and the effect of expressed factors on MAO B expression in vivo will be studied by determination of promoter activity, MAO B mRNA levels, protein levels and catalytic activity. These studies are important for both basic neuropharmacology and clinical research.