Excitatory signaling and oxidative phosphorylation alterations in schizophrenia

精神分裂症的兴奋性信号传导和氧化磷酸化改变

• 批准号: 9108013
• 负责人: JILL RENEE' Glausier
• 金额: $ 16.01万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-23 至 2020-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9108013
• 关键词: ATP Synthesis Pathway; Address; Affect; Animal Model; Animals; Autopsy; Axon; Brain; Brain region; Cells; Characteristics; Chronic; Clinical; Clinical Research; Complex; Coupled; Data; Diagnosis; Disease; Drug Targeting; Educational process of instructing; Educational workshop; Electron Microscopy; Electrons; Encephalopathies; Energy Metabolism; Environment; Enzymes; Event; Evolution; Experimental Animal Model; Experimental Designs; Factor-42; Faculty; Financial compensation; Functional disorder; Gene Expression; Genetic Transcription; Goals; Human; Impaired cognition; Impairment; Individual; Laboratories; Laboratory Research; Lasers; Measures; Mentored Research Scientist Development Award; Mentorship; Messenger RNA; Metabolic; Microdissection; Microscopic; Mitochondria; Mitochondrial Diseases; Molecular; Morphology; Mus; Neurons; Oxidases; Oxidative Phosphorylation; Parvalbumins; Pathology; Patients; Performance; Pharmacogenetics; Pharmacological Treatment; Physiological; Population; Postdoctoral Fellow; Prefrontal Cortex; Primates; Professional Competence; Proteins; Proteomics; Psychiatry; Pyramidal Cells; RNA; Research; Research Infrastructure; Research Proposals; Research Technics; Respiration; Rodent; Sampling; Schizophrenia; Scientist; Short-Term Memory; Signal Transduction; Site; Source; Symptoms; Techniques; Technology; Testing; Therapeutic; Tissue Model; Tissues; Training; Transcript; Transgenic Mice; Translations; Universities; Viral; Viral Vector; base; brain circuitry; brain tissue; calretinin; career; career development; cell type; cognitive function; cytochrome c oxidase; density; designer receptors exclusively activated by designer drugs; effective therapy; experience; gray matter; instructor; interest; laser capture microdissection; medical schools; member; neurotransmission; next generation; nuclear respiratory factor; professor; public health relevance; student mentoring; therapeutic target; training opportunity; transcription factor

 DESCRIPTION (provided by applicant): Schizophrenia is a complex disorder lacking an effective treatment option for the pervasive and debilitating cognitive impairments experienced by patients. I have been acutely interested in identifying the underlying circuitry alterations tha contribute to these impairments, so that they may be treated, since the start of my doctoral training. Though my focus has been singular, the approaches and conceptual framework used to study this problem have evolved over my scientific career, and this K01 application represents the next major step in that evolution. This K01 application includes research, clinical, and career development, along with teaching/training opportunities to maximize my ability to successfully transition to independence. My long term career goals are to 1) identify and describe the molecular and cellular alterations that contribute to cortical dysfunction in schizophrenia, 2) determine the possible causes of these observed pathologies using animal models, 3) use these findings to develop pathophysiology-based pharmacological treatments for cognitive impairment in schizophrenia patients, 4) establish an independent research laboratory at a top-tier university to accomplish these goals, and 5) mentor students and postdoctoral fellows to contribute to the next generation of scientists and innovators. My short term career goals include 1) master the research techniques proposed, including laser capture microdissection of single cell populations from human and rodent cortex, electron microscopic mitochondrial analysis of human prefrontal cortical tissue, and shRNA and Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) paradigms for rodent studies; 2) master experimental design to determine whether disease findings are a cause, compensation, consequence or confound; and 3) successfully transition as a faculty member from Instructor to Assistant Professor at the University of Pittsburgh. Importantly, completing these short terms goals via this K01 award begins to address long term goals 1 - 3, and puts me on a trajectory to accomplish the remaining long term objectives. The current training plan is also augmented with [[formal courses that address each of the major training goals]], interactive workshops focused on scientific career development, and teaching experiences to further broaden my career skill set. The Department of Psychiatry at the University of Pittsburgh's School of Medicine is an ideal environment in which to accomplish these short and long term goals. This department is a national leader in clinical research, treatment and training. Under the primary mentorship of Dr. David Lewis, Chair of Psychiatry, I will have full access to his laboratory and all of the infrastructure support required for the current application. Working memory is a core cognitive function impaired in schizophrenia that depends upon activation of prefrontal cortex (PFC) circuitry. Accordingly, individuals diagnosed with schizophrenia show reduced PFC activation while performing working memory tasks. This lower PFC activation appears to be an integral part of the disease pathophysiology, and not simply a reflection of poor performance. Thus, the cellular and circuitry alterations that underlie lower PFC neuronal activity in schizophrenia must be determined in order to identify appropriate therapeutic targets. This research proposal focuses on determining which of two discrete possible molecular/physiological disturbances is a likely upstream event leading to PFC impairments in schizophrenia. Supporting neuronal excitation represents the largest energy-consuming activity in the brain, supplied by ATP synthesis in mitochondria via oxidative phosphorylation (OXPHOS). Accumulating evidence indicates that expression of the terminal and rate-limiting OXPHOS enzyme, cytochrome c oxidase (COX), is lower in the PFC of schizophrenia subjects. Thus, the research goal of this K01 application is to determine the underlying mechanism contributing to lower levels of COX in the PFC of schizophrenia subjects. Lower COX could be a consequence of chronic reductions in neuronal excitation that lower ATP demand in the affected neurons (Hypothesis 1), or due to deficient COX expression that impairs metabolic capacity in all neurons (Hypothesis 2). Distinguishing between these alternatives has important implications for identifying appropriate therapeutic targets for cortical dysfunction in schizophrenia, as H1 indicates targeted enhancement of excitation, whereas H2 indicates enhancing COX expression to recover mitochondrial respiration. In order to test which hypothesis is most supported by findings in affected individuals, laser microdissection is used to dissect samples of three distinct neuronal populations and quantitative PCR is used to measure the expression of COX-related transcripts (Aim 1.1), and stereological electron microscopy is used to quantify mitochondrial abundance and morphology (Aim 1.2) in the PFC of schizophrenia and healthy comparison subjects. However, because cause- and-effect relationships cannot be determined using human postmortem tissue, experimental animal models are used in Aims 2 and 3 to directly test the mechanisms of H1 and H2. In Aim 2, DREADD pharmacogenetic technique is used to induce long-term reductions in PFC pyramidal cell excitation, and each measure from Aim 1 is assessed. In Aim 3, viral delivery of shRNA approach is used to impair COX availability in the PFC, and each measure from Aim 1 is assessed. Together, these Aims provide a definitive characterization of the disease phenomenon, and extend to proof-of-concept studies in animal models to provide compelling, convergent and conclusive data regarding which mechanism is operative in the illness.

 描述（由申请人提供）：精神分裂症是一种复杂的疾病，缺乏有效的治疗选择，患者经历的广泛和衰弱的认知障碍。自从我开始博士学习以来，我一直对确定导致这些损伤的潜在电路改变非常感兴趣，以便可以治疗它们。虽然我的研究重点是单一的，但在我的科学生涯中，用于研究这个问题的方法和概念框架已经发生了变化，而K 01应用程序代表了这一演变的下一个重要步骤。此K 01应用程序包括研究，临床， 和职业发展，沿着教学/培训机会，以最大限度地提高我成功过渡到独立的能力。我的长期职业目标是1）识别和描述导致精神分裂症皮质功能障碍的分子和细胞改变，2）使用动物模型确定这些观察到的病理的可能原因，3）使用这些发现开发基于病理生理学的药物治疗精神分裂症患者的认知障碍，4）在一流大学建立一个独立的研究实验室来实现这些目标，5）指导学生和博士后研究员为下一代科学家和创新者做出贡献。我的短期职业目标包括：1）掌握所提出的研究技术，包括人类和啮齿动物皮层单细胞群的激光捕获显微切割，人类前额叶皮层组织的电子显微镜线粒体分析，以及用于啮齿动物研究的shRNA和由设计药物独家激活的设计受体（DREADDs）范例; 2）掌握实验设计，以确定疾病的发现是否是一个原因，补偿，后果或混淆;和3）成功地过渡作为教师从讲师助理教授在匹兹堡大学。重要的是，通过这个K 01奖项完成这些短期目标，开始解决长期目标1 - 3，并把我放在一个轨道上，以实现其余的长期目标。当前的培训计划还增加了[[解决每个主要培训目标的正式课程]]、专注于科学职业发展的互动研讨会以及教学经验，以进一步拓宽我的职业技能。匹兹堡大学医学院精神病学系是实现这些短期和长期目标的理想环境。该部门在临床研究、治疗和培训方面处于全国领先地位。在精神病学主席大卫刘易斯博士的主要指导下，我将完全访问他的实验室和当前应用程序所需的所有基础设施支持。工作记忆是精神分裂症患者的核心认知功能，依赖于前额叶皮层（PFC）回路的激活。因此，被诊断患有精神分裂症的个体在执行工作记忆任务时表现出减少的PFC激活。这种较低的PFC激活似乎是疾病病理生理学的一个组成部分，而不仅仅是表现不佳的反映。因此，细胞和电路的改变，在精神分裂症的PFC神经元活性较低的基础上，必须确定，以确定适当的治疗目标。这项研究计划的重点是确定两个离散的可能的分子/生理干扰是一个可能的上游事件，导致PFC功能障碍的精神分裂症。支持神经元兴奋代表大脑中最大的能量消耗活动，由线粒体中经由氧化磷酸化（OXPHOS）的ATP合成提供。越来越多的证据表明，表达的终端和限速OXPHOS酶，细胞色素c氧化酶（考克斯），是较低的精神分裂症患者的PFC。因此，该K 01应用的研究目标是确定导致精神分裂症受试者PFC中考克斯水平较低的潜在机制。较低的考克斯可能是神经元兴奋的慢性减少的结果，其降低了受影响神经元中的ATP需求（假设1），或者是由于考克斯表达不足，其损害了所有神经元中的代谢能力（假设2）。区分这些替代方案对于确定精神分裂症中皮质功能障碍的适当治疗靶点具有重要意义，因为H1表示靶向增强兴奋，而H2表示增强考克斯表达以恢复线粒体呼吸。为了检验这一假设是最支持的结果在受影响的个人，激光显微切割被用来解剖样品的三个不同的神经元群体和定量PCR被用来测量COX相关的转录本的表达（目的1.1），和体视学电子显微镜被用来量化线粒体丰度和形态（目的1.2）在PFC的精神分裂症和健康的比较科目。然而，由于无法使用人类死后组织确定因果关系，因此在目标2和3中使用实验动物模型直接测试H1和H2的机制。在目标2中，使用DREADD药物遗传学技术诱导PFC锥体细胞兴奋的长期降低，并评估目标1的每项措施。在目标3中，使用病毒递送shRNA方法来削弱PFC中的考克斯可用性，并评估来自目标1的每个测量。总之，这些目标提供了疾病现象的明确表征，并扩展到动物模型中的概念验证研究，以提供关于疾病中哪种机制起作用的令人信服的，收敛的和结论性的数据。