Isoflurane: Identification of Key New Targets

异氟烷：关键新靶点的识别

• 批准号: 7647410
• 负责人: AARON P. FOX
• 金额: $ 48.43万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7647410
• 关键词: Affinity; Anesthesia procedures; Anesthetics; Animals; Base Pairing; Biological Assay; Catecholamines; Cell Line; Cells; Chromaffin Cells; Data; Data Set; Dose; Drug effect disorder; Electric Capacitance; Etomidate; Exhibits; Future; General anesthetic drugs; Generations; Genes; Glutamates; Goals; Hippocampus (Brain); Human; Isoflurane; Knock-in Mouse; Membrane Lipids; Membrane Potentials; Membrane Proteins; Modeling; Molecular; Monitor; Mus; Mutant Strains Mice; Mutate; Mutation; Nervous System Physiology; Neuraxis; Neurons; Operative Surgical Procedures; PC12 Cells; Pain; Pharmaceutical Preparations; Physiological; Play; Property; Propofol; Proteins; RNA; RNA Interference; Rattus; Reflex action; Research; Resistance; Role; S-nitro-N-acetylpenicillamine; Site; Structure of superior cervical ganglion; Synapses; Techniques; Testing; behavior test; clinically relevant; clinically significant; depressed; design; experience; fluidity; insight; knock-down; neurotransmitter release; receptor; research study; response; stable cell line; success; synaptotagmin I; syntaxin 1A; vesicle-associated membrane protein

DESCRIPTION (provided by applicant): Despite a great deal of research, a complete understanding of the actions of general anesthetics is still not available. The objective of this research is to advance our understanding of isoflurane, with the hope of gaining insights into all anesthetics. Isoflurane, a halogenated volatile anesthetic, is thought to produce anesthesia by depressing central nervous system function. Many anesthetics, including isoflurane, are thought to modulate and/ or directly activate GABAA receptors. Isoflurane is also known to have effects on other channels and receptors. In a set of preliminary studies we observed that isoflurane, at clinically relevant concentrations, dramatically inhibited the neurotransmitter release machinery in PC12 cells. Etomidate and propofol also inhibited the release machinery in PC12 cells, suggesting that inhibition of neurotransmitter release may be an important general action of anesthetics. Because neurotransmitter release mechanisms are strongly conserved between PC12 and neurons we hypothesize that anesthetics interfere with the neuronal neurotransmitter release machinery and this may represent an important site of action for anesthesia in intact animals. The experiments outlined in this application will be carried out in cultured hippocampal neurons, PC12 cells and mutant mice. This proposal has three goals. First, we will determine whether isoflurane interferes with neurotransmitter release in hippocampal neurons at clinically relevant concentrations. Next, the anesthetic site of action will be identified. We start by examining syntaxin 1A and unc-13, but other sites including SNAP- 25, synaptobrevin and synaptotagmin 1, will be investigated as well. Details about why these sites were selected are provided in this application. Once neurotransmitter release machinery targets are identified they will be mutated in order to suppress the interaction between isoflurane and the target. This information will be used to design knock-in mice with the same mutations, which will permit us to determine, using a battery of physiological and behavioral tests, whether these animals still respond to isoflurane (or other anesthetics) or whether their responses to the anesthetics are altered.Project Narrative: Isoflurane is a widely used volatile anesthetic. Our goal is to find out how isoflurane produces anesthesia. In particular, we will explore the effect on the neurotransmitter release machinery, as our preliminary data suggests this may be a critical site of action for isoflurane and other anesthetics. This may allow for the design of new more beneficial drugs in the future.

描述（由申请人提供）：尽管进行了大量研究，但仍无法完全了解一般麻醉药的行为。这项研究的目的是促进我们对异氟烷的理解，希望能够深入了解所有麻醉剂。异氟烷是一种卤代挥发性麻醉，被认为通过降低中枢神经系统功能会产生麻醉。许多麻醉药，包括异氟烷，都被认为可以调节和/或直接激活GABAA受体。众所周知，异氟烷对其他通道和受体具有影响。在一系列初步研究中，我们观察到，在临床相关的浓度下，异氟烷会极大地抑制PC12细胞中的神经递质释放机械。依托匹马甲酸和丙泊酚还抑制了PC12细胞中的释放机械，这表明抑制神经递质释放可能是麻醉剂的重要一般作用。由于神经递质释放机制在PC12和神经元之间得到了强烈的保守，因此我们假设麻醉药会干扰神经元神经递质释放机制，这可能代表了完整动物麻醉的重要作用部位。该应用中概述的实验将在培养的海马神经元，PC12细胞和突变小鼠中进行。该提议有三个目标。首先，我们将确定异氟烷是否会在临床相关浓度下干扰海马神经元中神经递质的释放。接下来，将确定动作的麻醉部位。我们首先检查语法1A和UNC-13，但也将研究包括SNAP-25，SNAPOLOVIN和SynaptoTagmin 1的其他站点。此应用程序中提供了有关为什么选择这些站点的详细信息。一旦确定了神经递质释放机械靶标，它们将被突变以抑制异氟烷与靶标之间的相互作用。该信息将用于设计具有相同突变的敲门鼠，这将使我们能够使用一系列的生理和行为测试来确定这些动物是否仍然对异氟烷（或其他麻醉药）做出反应，或者对麻醉剂的反应是否改变了。我们的目标是找出异氟烷如何产生麻醉。特别是，我们将探讨对神经递质释放机械的影响，因为我们的初步数据表明，这可能是异氟烷和其他麻醉剂的关键作用部位。这可能允许将来设计新的更有益的药物。