5-HT2C-receptor expression and function in depression

描述：(申请人的摘要)一个子类型的表达和功能 在5-羟色胺(5-HT)受体中，5-HT2C受体受RNA编辑的调节 以及其编码的Pre-mRNA的选择性剪接。在死后前额叶 抑郁症自杀者的皮质组织，5-HT2C的编辑模式 Pre-mRNA与对照组大脑的编辑模式有很大不同 可能导致5-HT2C受体-G蛋白的效率降低 互动。拟议研究计划的总体目标是 进一步区分抑郁症和更年期抑郁症之间的可能关系 前额叶皮质5-HT2C前体mRNA的编辑模式，并定义 此更改的编辑模式可能产生的功能后果并进行测试 可能的潜在机制。提出了三个具体目标，以实现 这些目标。在具体目标1中，调查员提出了一个扩展的 复制差异5-HT2C编辑发现，试点这项工作。这个 5-HT2C受体的编辑模式将在男性组的大脑中进行比较 抑郁症自杀受害者和抑郁症非自杀受试者评估 抑郁症与自杀的相对贡献。5-HT2C的编辑模式 受体mRNAs将通过产生的cDNAs的核苷酸序列来识别 用RT-PCR方法检测。拟议的具体目标2将评估 稳定表达NIH3T3的NIH3T3细胞对5-HT2C受体基因的编辑 表达相关的三种主要改变的5-HT2C受体亚型的细胞 抑郁症，以及更罕见的未编辑和完全编辑的亚型。 功能分析将确定编辑和编辑的相对能力 未编辑的与G蛋白偶联并激活磷脂酶C的异构体 第二信使系统。RNA编辑对鸟苷核苷酸的影响 5-HT2C受体与G蛋白相互作用的敏感性及其相互作用 不同的受体亚型也将被初步评估。相关 功能实验将比较GTP(S)与前额叶膜的结合 抑郁症自杀者和抑郁症非自杀者的皮质组织 受试者以确定5-HT2C编辑改变的效果 5-HT2C对基础和激动剂促进的G蛋白活化的抑制作用 感受器。具体目标3将比较机械上截然不同的 抗抑郁药和5-HT2A/2C受体拮抗剂酮丝林 新皮质5-HT2C受体RNA编辑模式，野生型小鼠。这些 研究将试图评估观察到的5-HT2C RNA的变化 对抑郁症的编辑反映了药物的效果。

DESCRIPTION: (Applicant's abstract) The expression and function of one subtype of serotonin (5-HT) receptors, the 5-HT2C receptor, is regulated by RNA editing and alternative splicing of its encoded pre-mRNA. In postmortem prefrontal cortical tissue from depressed suicide victims, the editing pattern of 5-HT2C pre-mRNA differs significantly from the editing pattern of control brains that could potentially result in a decreased efficiency of 5-HT2C receptor-G protein interactions. The overall objectives of the proposed research plan are to varify further a possible relationship between depression and an altered editing pattern of prefrontal cortical 5-HT2C pre-mRNA, and to define the possible functional consequences of this altered editing pattern and to test possible underlying mechanisms. Three specific aims are proposed to attain these objectives. In specific aim 1, the investigator proposes an extended replication of the differential 5-HT2C editing finding piloting this work. The 5-HT2C receptor editing pattern would be compared in brains of groups of male depressed suicide victims and depressed non-suicide subjects to assess the relative contribution of depression versus suicide. Editing patterns of 5-HT2C receptor mRNAs would be identified by nucleotide sequencing of cDNAs generated by RT-PCR. Proposed specific aim 2 would assess the functional significance of altered 5 HT2C receptor mRNA editing using stably transfected lines of NIH3T3 cells that express the three major altered 5-HT2C receptor isoforms associated with depression, as well as the more rare nonedited and fully edited isoforms. Functional assays would determine the comparative ability of the edited and nonedited isoforms to couple to G protein and activate the phospholipase C second messenger system. The effects of RNA editing on the guanyl nucleotide sensitivity of 5-HT2C receptor-G protein interactions, and the interaction of different receptor isoforms would also be preliminarily assessed. Related functional experiments would compare GTP(S binding to membranes of prefrontal cortical tissues of depressed suicide victims and depressed non-suicide subjects to determine the effect of 5-HT2C editing alterations associated with depression on basal and agonist-promoted activation of G proteins by 5-HT2C receptors. Specific aim 3 would compare the effects of mechanistically distinct antidepressants, and the 5-HT2A/2C receptor antagonist ketanserin, on the neocortical 5-HT2C receptor RNA editing pattern, in wild-type mice. These studies would attempt to assess whether the observed alteration of 5-HT2C RNA editing in depression reflects medication effects.