A mouse model linking anesthetic sensitivity to mitochondrial function

将麻醉敏感性与线粒体功能联系起来的小鼠模型

基本信息

批准号：
8628393
负责人：
Margaret Mary Sedensky
金额：
$ 54.14万
依托单位：
SEATTLE CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-05-01 至 2018-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8628393
关键词：
A Mouse Absence of pain sensation Adverse effects Affect Anesthesia procedures Anesthetics Animal Model Animals Area Astrocytes Behavior Biology Brain region Caenorhabditis elegans Cells Child Complex Conscious Defect Dose Drug Design Electron Transport Functional disorder Future Genetic Glutamates Goals Halothane Health Human Hypersensitivity Invertebrates Investigation Isoflurane Ketamine Knock-out Knockout Mice Knowledge Laboratories Lead Link Mammals Measurement Measures Mediating Memory Mental Depression Mitochondria Mitochondrial Diseases Modeling Modern Medicine Molecular Monitor Mus Mutant Strains Mice Mutation Nematoda Neonatal Nervous system structure Neurons Newborn Animals Operative Surgical Procedures Organism Patients Pharmaceutical Preparations Phenotype Phylogeny Physiology Play Primates Reflex action Resistance Rodent Role Safety Supporting Cell Surgical incisions Synapses Synaptic Transmission System Tail Testing Unconscious State Wild Type Mouse Work animal data behavior change cell type cholinergic improved insight interest knockout animal man mitochondrial dysfunction mouse model neurotoxicity neurotransmission novel response synaptic function tool

项目摘要

DESCRIPTION (provided by applicant): A true enigma of modern medicine has persisted for over 150 years. How can volatile anesthetics, as a class of drugs, simultaneously cause a patient to reversibly lose consciousness, feel no pain, and have no memory of surgery? The mechanism by which volatile anesthetics (VAs) produce reversible loss of consciousness and render an organism insensate to a surgical incision remains an unsolved mystery. Our laboratory has exploited a very simple animal model, the nematode C. elegans, to investigate the molecular mechanisms of volatile anesthetic action. Previously, we demonstrated that mitochondrial complex I, an entry point of the electron transport chain, specifically controls the sensitivity of C. elegans to volatile anesthetics. We also found that complex I defects control anesthetic sensitivity in humans. We now have an exciting opportunity to extend those findings into mice using the knockout animal, Ndufs4. The Ndufs4 KO mice lack a subunit of complex I, which results in complex I dysfunction. We anesthetized young (PN23-27) Ndusf4 and wild-type (WT) mice with isoflurane or halothane. For either VA, the KO mice became unresponsive to a tail pinch at a dose 2.8-fold lower than for WT controls. These KO mice display the greatest change in VA sensitivity ever described in a mammal. We also measured the EC50s for loss of righting reflex (LORR) of the KO mice for anesthetics whose targets are well characterized. Surprisingly, the animals were actually resistant to the effects of ketamine. The effects of the complex I KO are specific in terms of anesthetic and not simply the result of generalized CNS depression. Complex I dysfunction causes hypersensitivity to volatile anesthetics across phylogeny. However, the question remains, how do complex I defects affect VA sensitivity. We hypothesize that complex I depression (i.e. energy depletion) leads to defective synaptic function in specific cell types, making them more susceptible to neuronal silencing by VAs. We will knock out Ndufs4 in GABAergic, cholinergic or glutamatergic neurons, in neuronal support cells known as astrocytes, as well as in specific regions of the brain. We will also perform electrophysiologic measurements of WT and mutant mice with and without volatile anesthetics. Our initial results indicate that the VA hypersensitivity in Ndufs4 mice is mediated through glutamatergic neurons. Our aims are to characterize which neurons and regions of the brain are important for the anesthetic phenotype of these animals, as well as to discover how basic neuronal physiology is altered in the KO animals. Our overarching goal is to understand how the VAs functions. We have linked mitochondrial function to behavior in VAs in worms, mice, and man, a finding the field must consider. Our proposed studies will tease out which cell types mediate the anesthetic response, and will give important insights into the basic mechanisms of action of VAs.

描述（由申请人提供）：现代医学的一个真正的谜团已经持续了 150 多年。挥发性麻醉药作为一类药物，如何能够同时导致患者可逆性失去意识、感觉不到疼痛、对手术失去记忆？挥发性麻醉剂（VA）产生可逆性意识丧失并使有机体对手术切口失去知觉的机制仍然是一个未解之谜。我们的实验室利用了一种非常简单的动物模型，即线虫线虫，来研究挥发性麻醉作用的分子机制。此前，我们证明线粒体复合物 I（电子传递链的入口点）特异性控制线虫对挥发性麻醉剂的敏感性。我们还发现复合体 I 缺陷控制人类的麻醉敏感性。我们现在有一个令人兴奋的机会，可以使用基因敲除动物 Ndufs4 将这些发现扩展到小鼠身上。 Ndufs4 KO 小鼠缺乏复合物 I 的亚基，导致复合物 I 功能障碍。我们用异氟烷或氟烷麻醉年轻 (PN23-27) Ndusf4 和野生型 (WT) 小鼠。对于任一 VA，KO 小鼠对尾部捏捏均无反应，剂量比 WT 对照低 2.8 倍。这些 KO 小鼠在 VA 敏感性方面表现出哺乳动物有史以来最大的变化。我们还测量了 KO 小鼠翻正反射丧失 (LORR) 的 EC50，以使用其目标已明确表征的麻醉剂。令人惊讶的是，这些动物实际上对氯胺酮的作用具有抵抗力。复合物 I KO 的作用在麻醉方面具有特异性，而不仅仅是全身中枢神经系统抑制的结果。复合物 I 功能障碍会导致整个系统发育过程中对挥发性麻醉剂的过敏。然而，问题仍然存在，复合体 I 缺陷如何影响 VA 敏感性。我们假设复合物 I 抑制（即能量消耗）会导致特定细胞类型的突触功能缺陷，使它们更容易受到 VA 的神经元沉默影响。我们将敲除 GABA 能、胆碱能或谷氨酸能神经元、称为星形胶质细胞的神经元支持细胞以及大脑特定区域中的 Ndufs4。我们还将在使用或不使用挥发性麻醉剂的情况下对 WT 和突变小鼠进行电生理学测量。我们的初步结果表明，Ndufs4 小鼠的 VA 超敏反应是通过谷氨酸能神经元介导的。我们的目标是确定哪些神经元和大脑区域对这些动物的麻醉表型很重要，并发现 KO 动物的基本神经元生理学是如何改变的。我们的首要目标是了解 VA 的运作方式。我们已经将线粒体功能与线虫、小鼠和人类的 VA 行为联系起来，这一发现值得该领域考虑。我们提出的研究将弄清楚哪些细胞类型介导麻醉反应，并将为 VA 的基本作用机制提供重要的见解。