GENERAL ANESTHETICS AND CLONED VOLTAGE-GATED CHANNELS

普通麻醉剂和克隆电压门控通道

• 批准号: 2685091
• 负责人: ANA M CORREA
• 金额: $ 10.27万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-04-01 至 2001-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2685091
• 关键词: Xenopus; Xenopus oocyte; anesthetics; calcium channel; chimeric proteins; complementary RNA; drug adverse effect; electrical conductance; general anesthesia; inhalation anesthesia; intravenous anesthesia; membrane transport proteins; molecular cloning; muscle pharmacology; neuropharmacology; potassium channel; sodium channel; voltage /patch clamp; voltage gated channel

General anesthesia is a condition in which loss of memory and consciousness is accompanied by loss of pain sensation. Inhalation general anesthetics (GA), fundamental aides in surgical anesthesia, induce both but not without displaying some serious side effects. Because of the relevance of neurotransmission in hypnotic and analgesic effects, a substantial effort has been placed in the study of the action of GA at the synapse, with emphasis on effects on ligand-gated receptors and channels. The effects of GA on voltage-gated ion channels are of no less importance in neuronal transmission. Voltage-gated ion channels play an added crucial role in general anesthesia in that many side effects of GA have been traced to impaired function of these channels in various tissues. A well-documented example of GA induced side effects involving voltage-gated channels is that of cardiac depression, which puts patients under considerable risk. The design of 'ideal' general anesthetics will emerge from the knowledge of the specific molecular mechanisms involved in GA effects. It is the long-term goal of this project to contribute to the understanding of the mechanisms of action of GA by providing novel information on G-channel interactions. In this proposal the effects of volatile anesthetics an of a very promising intravenous anesthetic, Propofol, will be studied in detail by taking advantage of the tools provided by molecular biology, by a high expression system (Xenopus oocytes) and by high resolution electrophysiological recording techniques. With this approach, new detailed studies of the function of single species of channels can be performed with essentially no contamination from other membrane proteins. The specific aims are: 1) To characterize the action of general inhalation anesthetics on the macroscopic gating and conductance properties of cloned voltage-gated ion channels. The cut-open oocyte voltage-clamp technique will be used to record ionic and gating currents from oocytes expressing K+, Na+ or Ca2+ channels from injected cRNA. 2) To characterize general anesthetic effects on the function of the individual channel proteins by recording at the single-channel level. Single-channel activity will be recorded using the patch-clamp technique, 3) To determine, from comparative studies, regions of the structure that interact with the anesthetics and/or that are directly involved in the affected functions. In these studies, different combinations of subunits and channel subtypes., chimeras and channel mutants will be examined. The goal is to identify regions in the proteins involved in specific interactions with the anesthetics. The roles played by auxiliary subunits will be investigated. (4) To initiate studies on the action of intravenous anesthetics on voltage-gated ion channels. These studies will concentrate on the effects of Propofol on macroscopic, gating and single-channel currents. The outcome of this project should provide important information on relevant structural features intervening in GA- channel interactions.

全身麻醉是一种记忆和意识丧失的情况 伴随着痛觉丧失。 吸入性全身麻醉药 (GA)，手术麻醉的基本助手，诱导两者，但不是没有 出现了严重的副作用 由于相关的 催眠和镇痛作用的神经传递，一个实质性的努力， 已经被放置在GA在突触的作用的研究中， 强调对配体门控受体和通道的影响。 的影响 GA对电压门控离子通道的作用在神经元细胞中同样重要。 传输 电压门控离子通道在细胞凋亡中起着额外的关键作用。 全身麻醉，因为GA的许多副作用已被追溯到 这些通道在各种组织中的功能受损。 一个证据充分的 GA诱导的涉及电压门控通道的副作用的例子是， 心源性抑郁，这会让患者面临相当大的风险 的 “理想的”全身麻醉剂的设计将从对麻醉剂的认识中产生。 GA效应的具体分子机制。 这是长期的 本项目的目标是促进对机制的理解， GA的行动，提供新的信息G通道的相互作用。 在这个提议中，挥发性麻醉剂的作用是一种非常有前途的方法。 静脉麻醉剂，异丙酚，将详细研究，采取 分子生物学提供的工具的优势，通过高表达 系统（非洲爪蟾卵母细胞）和高分辨率电生理 记录技术。 通过这种方法，新的详细研究， 单一种类的通道的功能可以在基本上没有 其他膜蛋白的污染。 具体目标是：（1） 描述全身吸入麻醉剂对 克隆电压门控离子的宏观门控和电导特性 渠道 切开卵母细胞电压钳技术将用于 记录来自表达K+、Na+或Ca 2+的卵母细胞的离子和门控电流 来自注射的cRNA的通道。2)描述全身麻醉效果 通过在细胞膜上记录单个通道蛋白的功能， 单通道水平。 单通道活动将使用 膜片钳技术，3）从比较研究中确定区域， 与麻醉剂相互作用和/或 直接参与受影响的功能。 在这些研究中， 亚基和通道亚型的组合，嵌合体和通道 将对突变体进行检查。 目标是确定蛋白质中的区域 参与与麻醉剂的特定相互作用。 发挥的作用 将研究辅助亚单位。 (4)启动关于 静脉麻醉剂对电压门控离子通道的作用。 这些 研究将集中在异丙酚对宏观，门控， 和单通道电流。 该项目的成果应提供 关于干预GA的相关结构特征的重要信息- 渠道互动。