5-HT2C-receptor expression and function in depression

描述：（申请人摘要）一种亚型的表达和功能 5-羟色胺（5-HT）受体，即5-HT2C受体，受RNA编辑调控 以及其编码的前mRNA的选择性剪接。在死后前额叶 抑郁自杀受害者的皮层组织，5-HT2C的编辑模式 前mRNA与对照组大脑的编辑模式显著不同， 可能导致5-HT 2C受体-G蛋白的效率降低 交互.拟议研究计划的总体目标是 进一步证实了抑郁症和改变的 前额叶皮质5-HT2C前体mRNA的编辑模式，并定义 这种改变的编辑模式的可能功能后果，并测试 可能的潜在机制。建议实现三个具体目标 这些目标。在具体目标1中，研究人员提出了一个扩展的 复制差异5-HT2C编辑发现引导这项工作。的 5-HT 2C受体编辑模式将在男性组的脑中进行比较。 抑郁的自杀受害者和抑郁的非自杀受试者，以评估 抑郁症与自杀的相对贡献。5-HT2C的编辑模式 通过对产生的cDNA进行核苷酸测序来鉴定受体mRNA， 用rt-pcr拟议的具体目标2将评估以下方面的职能意义： 使用稳定转染的NIH3T3细胞系改变5-HT 2C受体mRNA编辑 表达三种主要改变的5-HT2C受体亚型的细胞， 抑郁症，以及更罕见的未经编辑和完全编辑的亚型。 功能测定将确定编辑的和未编辑的蛋白质的比较能力。 未编辑的同种型与G蛋白偶联并激活磷脂酶C 第二信使系统RNA编辑对鸟苷酸的影响 5-HT2C受体-G蛋白相互作用的敏感性，以及 还将初步评估不同的受体同种型。相关 功能实验将比较GTP（S）与前额叶膜的结合， 抑郁的自杀受害者和抑郁的非自杀受害者的皮层组织 受试者，以确定5-HT2C编辑改变与 5-HT 2C抑制G蛋白的基础和激动剂促进的激活 受体。具体目标3将比较机械上不同的 抗抑郁药和5-HT2A/2C受体拮抗剂酮色林， 新皮质5-HT2C受体RNA编辑模式，在野生型小鼠中。这些 研究将试图评估观察到的5-HT2C RNA的改变是否 抑郁症的编辑反映了药物作用。

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.