Optogenetic approaches to study post-stroke recovery mechanisms

研究中风后恢复机制的光遗传学方法

• 批准号: 10211210
• 负责人: GARY K STEINBERG
• 金额: $ 60.44万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2025-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10211210
• 关键词: Address; Affect; Animals; Area; Automobile Driving; Biological; Brain; Brain Diseases; Brain region; Calcium; Cause of Death; Cell Count; Characteristics; Cholesterol; Clinical Research; Corpus striatum structure; Data; Event; Female; Frequencies; Functional Magnetic Resonance Imaging; Future; G Protein-Coupled Receptor Signaling; Genes; Goals; Grant; High-Throughput RNA Sequencing; Image; Imaging Techniques; Individual; Infarction; Intervention; Location; Maps; Medical; Metabolism; Microscope; Molecular; Molecular Profiling; Molecular Target; Motor Cortex; Mus; Neurons; Parvalbumins; Pathway interactions; Population; Recovery; Recovery of Function; Reporting; Resolution; Role; Sex Differences; Site; Somatosensory Cortex; Stains; Stroke; Techniques; Testing; Thalamic structure; Therapeutic; Therapeutic Intervention; Time; Translating; Validation; behavior test; brain remodeling; calmodulin-dependent protein kinase II; cell type; cholesterol biosynthesis; design; disability; effective therapy; functional outcomes; imaging system; improved; insight; male; multimodality; nervous system disorder; neural circuit; neuroimaging; neuroregulation; new therapeutic target; optogenetics; partial recovery; portability; post stroke; promoter; relating to nervous system; sex; stroke outcome; stroke recovery; tool; transcriptome; transcriptome sequencing

PROJECT SUMMARY Stroke is the leading cause of death with very limited treatment options. This devastating neurological disease is increasingly viewed as a disease of brain connectivity as a damaged stroke area can affect both local and connected brain regions, causing disruptions in neuronal activity and metabolism network-wide. Recovery of lost function can occur after stroke and is attributed to brain remodeling in areas adjacent to or connected to the infarct. In this proposal, we aim to investigate the role of key brain circuits in post-stroke recovery at the functional, cellular and molecular level, using optogenetics, advanced live imaging and high throughput RNA sequencing techniques. Previously our lab has demonstrated that selective optogenetic neuronal stimulation in the ipsilesional motor cortex (iM1) can activate plasticity mechanisms and promote recovery. Recently we have employed the optogenetic functional MRI technique to systematically map brain-wide changes in neural circuits after stroke. We have identified key circuits altered by stroke and demonstrated two key circuits restored by iM1 stimulations. Our map data also revealed two candidate circuits that were not restored by iM1 stimulations, suggesting that greater recovery could be achieved if we can rescue these circuits by directly stimulating them. In this proposal we aim to investigate key neural circuits we identified from our activation maps and elucidate their role in post-stroke recovery. In Aim1 we will use circuit-specific optogenetic tools and functional behavior tests to interrogate the role of key circuits in post-stroke recovery. This aim will address whether these circuits have beneficial or maladaptive role during post-stroke recovery. In Aim2 we will examine cellular resolution of real-time neuronal activity dynamics in key circuits after stroke using a portable live calcium imaging system. This will elucidate the neural activity dynamics (excitatory and inhibitory) of key circuits at the cellular level, allowing us to identify the temporal profile and the key neuronal populations altered by stroke, and how iM1 stimulations affect these characteristics to enhance recovery. In Aim3 we will investigate the transcriptome of key circuit areas using RNAseq, in order to identify key molecular targets and pathways altered by stroke and by iM1 stimulations. Preliminary RNAseq analysis revealed distinct pathways altered by iM1 stimulations. We aim to perform RNAseq in multiple regions including iM1 (stimulation site) and ipsilesional thalamus (iM1- connected region) to elucidate whether similar pathways are involved, and if we can identify a common molecular signature that drive recovery. We will also perform RNAseq in both sexes in order to ascertain any sex-specific differences that may be present in post-stroke recovery. Together these results will 1) advance the understanding of neural circuit dynamics during post-stroke recovery; and 2) identify key neural circuits/cell types/molecular targets and optimal time window for designing brain stimulation strategies and other therapeutic interventions in future clinical studies.

项目概要 中风是导致死亡的主要原因，治疗选择非常有限。这种毁灭性的神经系统疾病 越来越多地被视为一种大脑连接疾病，因为受损的中风区域会影响局部和 连接大脑区域，导致整个网络的神经元活动和新陈代谢中断。恢复 中风后可能会发生功能丧失，这是由于邻近或连接区域的大脑重塑所致 梗塞。在本提案中，我们的目标是研究关键脑回路在中风后恢复中的作用 使用光遗传学、先进的实时成像和高通量 RNA 进行功能、细胞和分子水平 测序技术。此前我们的实验室已经证明选择性光遗传学神经元刺激 同侧运动皮层（iM1）可以激活可塑性机制并促进恢复。最近我们有 采用光遗传学功能磁共振成像技术来系统地绘制全脑神经回路的变化 中风后。我们已经确定了因中风而改变的关键电路，并演示了通过中风恢复的两个关键电路 iM1 刺激。我们的地图数据还揭示了两个未通过 iM1 刺激恢复的候选回路， 这表明，如果我们能够通过直接刺激这些回路来挽救它们，则可以实现更大的恢复。 在这个提案中，我们的目标是研究我们从激活图中识别出的关键神经回路并阐明 他们在中风后恢复中的作用。在 Aim1 中，我们将使用电路特定的光遗传学工具和功能行为 进行测试以探究关键回路在中风后恢复中的作用。该目标将解决这些电路是否 在中风后恢复过程中具有有益或适应不良的作用。在 Aim2 中，我们将检查细胞分辨率 使用便携式实时钙成像系统来测量中风后关键回路中的实时神经元活动动态。 这将阐明细胞水平上关键回路的神经活动动力学（兴奋性和抑制性）， 使我们能够识别因中风而改变的时间分布和关键神经元群，以及 iM1 如何改变 刺激会影响这些特征以促进恢复。在 Aim3 中，我们将研究以下转录组 使用 RNAseq 分析关键电路区域，以确定因中风和中风而改变的关键分子靶点和通路 通过 iM1 刺激。初步 RNAseq 分析揭示了 iM1 刺激改变的不同途径。我们 旨在在多个区域进行 RNAseq，包括 iM1（刺激位点）和同侧丘脑 (iM1- 连接区域）来阐明是否涉及相似的路径，以及我们是否可以识别出一个共同的路径 驱动恢复的分子特征。我们还将对两性进行 RNAseq，以确定任何 中风后恢复中可能存在的性别差异。这些结果共同将 1) 推进 了解中风后恢复期间的神经回路动力学； 2) 识别关键神经回路/细胞 用于设计大脑刺激策略等的类型/分子靶点和最佳时间窗口 未来临床研究中的治疗干预。