DOPAMINE D2-RECEPTOR MEDIATED SIGNAL TRANSDUCTION

本项目的总体目标是探索信号转导 多巴胺通过D2受体起作用的机制，相信 这可能阐明了这些受体在调节 神经元活动 因此，目前的建议需要两个 方法-首先，建立一个模型细胞系统，其中多巴胺的 调节细胞内第二信使cAMP和[Ca 2 +] i的作用 其次，研究电生理学 D2受体介导信号的生化扰动的后果 纹状体中的转导。 待评估的模型系统将是一个 神经母细胞瘤×中国仓鼠胚脑移植杂种细胞系 （NCB-20），其表达多巴胺、5-羟色胺、缓激肽 鸦片制剂和乙酰胆碱。 第一组实验将决定 D2受体是否抑制NCB-20细胞中的腺苷酸环化酶， 这些受体是否也可能干扰[Ca2 +] i动员在这些 细胞，通过测量D2-受体对磷脂酰肌醇的作用 代谢和[Ca2 +] i动员。 在第二组实验中， 条件将被优化用于纹状体内注射百日咳 毒素，使得百日咳毒素底物的显著部分 是ADP核糖基化的;将确定 这种治疗消除了受体介导的抑制， 腺苷酸环化酶;随后，受体介导的改变， 将在这些动物中检查电生理特性 与假注射对照相比。

The overall goal of this project is to explore signal transduction mechanisms for dopamine acting through D2-receptors, in the belief that this may elucidate the role played by these receptors in regulating neuronal activity. Consequently, the present proposal takes two approaches - first, to establish a model cell system in which dopamine's role in modulating the intracellular second messengers, cAMP and [Ca2+]i can be delineated and secondly, to investigate the electrophysiological consequences of a biochemical perturbation of D2-receptor mediated signal transduction in striatum. The model system to be evaluated will be a neuroblastoma x Chinese hamster embryonic brain explant hybrid cell line (NCB-20), which expresses receptors for dopamine, serotonin, bradykinin, opiates and acetylcholine. The first group of experiments will determine whether D2-receptors inhibit adenylate cyclase in NCB-20 cells and whether these receptors also may perturb [Ca2+]i mobilization in these cells, by measuring D2-receptor effects on phosphatidylinositol metabolism and [Ca2+]i mobilization. In the second group of experiments, conditions will be optimized for the intrastriatal injection of pertussis toxin, so that a significant portion of the pertussis toxin substrates are ADP-ribosylated; a determination will be made on the effectiveness with which this treatment eliminates receptor-mediated inhibition of adenylate cyclase; subsequently, receptor-mediated alterations in electrophysiological properties will be examined in these animals compared to sham-injected controls.