ANTISENSE KNOCKOUT OF CNS D2 DOPAMINE RECEPTORS

尽管多巴胺能受体激动剂和拮抗剂 一系列精神和神经疾病的标准疗法， 多巴胺受体的神经生物学研究受到严重阻碍 不能有选择地操纵个别多巴胺受体亚型。 药理工具只允许两个受体“家族”(d1和d2) 被区分开来，而最近的克隆研究证明了 至少5种多巴胺受体亚型。目前的实验 利用这些最新的分子进展利用反义 选择性阻断多巴胺产生的寡脱氧核苷酸 活体内的受体。反义基因敲除可以高度特异性地阻止 首次将信使核糖核酸翻译成功能性受体蛋白 时间，一种用于获取有关的功能信息的选择性手段 个别多巴胺受体亚型。初步数据包括在 提示中枢神经系统注射D2反义寡核苷酸可产生特异性D2多巴胺 大鼠脑内受体敲除与多巴胺介导的减少 行为和自身受体功能。我们将使用受体放射自显影 和匀浆结合以实现D2多巴胺的参数分析 受体敲除在时间进程、剂量相关性和受体方面的作用 专一性。反义寡核苷酸Will的局部脑内应用 允许选择性地敲除突触后和突触前的D2受体。 D2多巴胺受体基因敲除的功能方面将由 自发和药物诱发的运动活动分析。活体内 微透析将用于评估特定D2基因敲除对 纹状体乙酰胆碱的释放及其自身调节 纹状体多巴胺的合成。活体电生理实验 将用于确定多巴胺受体亚型负责 自身受体介导的多巴胺末端兴奋性的变化，并 监测反义治疗对自发性多巴胺神经元的影响 活动。最后，我们建议将体内反义技术应用于 其他多巴胺受体亚型的研究，从D3开始 为其提供导频数据的接收器。有关的唯一信息 特定多巴胺受体亚型在中枢作用中的作用(S) 多巴胺将从这些研究中获得。数据与以下内容相关 了解多巴胺在正常和异常行为中的作用 发展更具体的方式来影响这些过程。

Despite the fact that dopaminergic receptor agonists and antagonists are standard therapy for a number of psychiatric and neurological disorders, study of the neurobiology of dopamine receptors is severely hampered by an inability to selectively manipulate individual dopamine receptor subtypes. Pharmacological tools permit only two receptor "families" (D1 and D2) to be distinguished whereas recent cloning studies demonstrate the existence of at least 5 dopamine receptor subtypes. The present experiments capitalize on these recent molecular advances by utilizing antisense oligodeoxynucleotides to selectively block the production of dopamine receptors in vivo. Antisense knockout allows the highly specific arrest of mRNA translation into functional receptor protein providing, for the first time, a selective means for acquiring functional information regarding individual dopamine receptor subtypes. Preliminary data are included to show that CNS infusion of D2 antisense oligo produces specific D2 dopamine receptor knockout in rat brain along with decreases in dopamine-mediated behaviors and autoreceptor function. We will use receptor autoradiography and homogenate binding to effect a parametric analysis of D2 dopamine receptor knockout in terms of time course, dose-relatedness, and receptor specificity. Local, intracerebral application of the antisense oligo will allow for selective knockout of post-versus pre-synaptic D2 receptors. Functional aspects of D2 dopamine receptor knockout will be determined by analysis of spontaneous and drug-induced motor activity. In vivo microdialysis will be used to assess the impact of specific D2 knockout on striatal acetylcholine release and on autoregulation of the release and synthesis of striatal dopamine. In vivo electrophysiological experiments will be used to determine the dopamine receptor subtype responsible for autoreceptor-mediated changes in dopamine terminal excitability, and to monitor any effects of antisense treatment on dopamine neuron spontaneous activity. Finally, we propose to apply the in vivo antisense technique to the study of other dopamine receptor subtypes, beginning with the D3 receptor for which pilot data is presented. Unique information on the role(s) of specific dopamine receptor subtypes in the central actions of dopamine will be obtained from these studies. The data are relevant to understanding the role of dopamine in normal and abnormal behavior and to the development of more specific ways to influence these processes.