How do neurons in the brain decide to refine their synaptic connections in vivo?

大脑中的神经元如何决定在体内完善其突触连接？

• 批准号: 9383862
• 负责人: Hisashi Umemori
• 金额: $ 68.63万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-16 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9383862
• 关键词: Alpha Cell; Autistic Disorder; Axon; Brain; CD47 gene; Cell Adhesion Molecules; Cerebral hemisphere; Complement; Cues; Data; Defect; Development; Disease; Dominant-Negative Mutation; Electrophysiology (science); Electroporation; Emotional; Etiology; Functional disorder; Gene Expression; Genetic; Goals; Hippocampus (Brain); Image; JAK2 gene; Knock-out; Knockout Mice; Limbic System; Maintenance; Mediating; Memory; Mental disorders; Microglia; Molecular; Mus; Neurons; Pathway interactions; Phagocytosis; Phosphotransferases; Physiological; Play; Population; Presynaptic Receptors; Process; Protein Tyrosine Kinase; Regulation; Role; Schizophrenia; Signal Transduction; Signaling Molecule; Social Behavior; Synapses; System; Testing; Time; Work; cingulate cortex; design; gene complementation; in utero; in vivo; insight; mutant; nervous system disorder; neural circuit; neuropsychiatric disorder; neurotransmitter release; novel; optogenetics; overexpression; postsynaptic neurons; prevent; relating to nervous system; response; transgene expression

Formation of functional neural circuits is critical for proper functioning of the brain. To establish the most efficient synaptic circuits, synaptic connections must be refined by neural activity during development. In this proposal, we will determine the molecules and manner by which functional circuits are established by neural activity, focusing on the limbic system (including the hippocampus and cingulate cortex), which is implicated in emotional processing, memory formation and social behavior. Using a mouse genetic system in which restricted populations of hippocampal neurons can be conditionally inactivated, we found that hippocampal axons are refined through activity-dependent competition, where active neuronal connections stay (maintained) while inactive ones leave (eliminated). We further found that a cell adhesion molecule SIRP from postsynaptic neurons stabilizes active synapses through its presynaptic receptor CD47, serving as a "Stay" signal. To identify the signaling molecules that play critical roles in inactive axon elimination ("Go" signal), we generated a new system in which neural activity and gene expression can be conditionally controlled in vivo, using in utero electroporation. When neurotransmitter release is blocked in a subset of neurons in the cingulate cortex, their callosal projections (the major connections between the cerebral hemispheres) are eliminated during development. Using this system, we screened for signaling molecules that are upregulated in inactive neurons right before their axons start to leave and identified the Ca2+-dependent tyrosine kinase Pyk2. Inactive axons were not eliminated when a kinase-dead mutant of Pyk2 was expressed, indicating that Pyk2 activity is necessary for inactive axons to leave. We further identified that a Pyk2-interacting kinase, JAK2, is also necessary for inactive axon elimination. Consistently, Pyk2 and JAK2 are activated in inactive neurons. Finally, we found that overexpression of Pyk2 or JAK2 induces axonal elimination even when axons are active. We propose that the Pyk2-JAK2 pathway is the "Go" signal and serves as the determinant of axon refinement. To further characterize this pathway and to understand how the "Go" and "Stay" pathways regulate activity- dependent axon/synapse refinement, we propose to: Aim 1: Investigate the role of Pyk2 and JAK2 for axon/synapse refinement in physiological conditions. Aim 2: Analyze the electrophysiological consequences of Pyk2/JAK2 inactivation during synapse refinement using conditional KO mice. Aim 3: Examine whether the Pyk2-JAK2 pathway provides cues for microglial clearance of inactive axons in vivo. Aim 4: Investigate the interaction between the Stay (SIRP-CD47) and Go (Pyk2-JAK2) pathways in axon/synapse refinement in vivo. Our project will molecularly delineate how neurons decide to establish functional synaptic connections in the mammalian brain. Pyk2 and JAK2 are associated with various neuropsychiatric disorders. Many forms of mental illness including autism and schizophrenia are associated with abnormal alterations in the limbic circuitry. Thus, our studies should also yield novel insights into the etiology and treatment of such disorders.

功能性神经回路的形成对于大脑的正常运作至关重要。建立了世界上 有效的突触回路，突触连接必须在发育过程中通过神经活动来完善。在这 建议，我们将确定分子和方式，功能电路是由神经网络建立的。 活动，集中在边缘系统（包括海马和扣带皮层），这是牵连 情绪处理、记忆形成和社会行为。利用老鼠遗传系统， 海马神经元的限制性群体可以条件性失活，我们发现海马神经元 轴突是通过活动依赖性竞争，其中活跃的神经元连接保持（维持） 而不活跃的人则离开（被淘汰）。我们进一步发现，细胞粘附分子SIRP α在细胞内表达， 突触后神经元通过其突触前受体CD 47稳定活跃的突触，作为“停留” 信号了为了鉴定在非活性轴突消除（“Go”信号）中起关键作用的信号分子，我们 产生了一个新的系统，其中神经活动和基因表达可以在体内有条件地控制， 使用子宫内电穿孔。当神经递质释放在扣带回的一部分神经元中被阻断时， 皮质，它们的胼胝体投射（大脑两半球之间的主要连接）被消除 在发展过程中。使用这个系统，我们筛选了在失活状态下上调的信号分子 神经元的轴突开始离开之前，并确定了钙依赖性酪氨酸激酶Pyk 2。非活动 当Pyk 2的激酶死亡突变体表达时，轴突没有被消除，这表明Pyk 2的活性是 不活跃的轴突离开所必需的。我们进一步确定了Pyk 2相互作用激酶JAK 2也是一种 对于消除不活跃的轴突是必要的。Pyk 2和JAK 2在非活动神经元中被激活。最后， 我们发现Pyk 2或JAK 2的过表达诱导轴突消除，即使轴突是活跃的。我们 提出Pyk 2-JAK 2通路是“Go”信号，并作为轴突细化的决定因素。到 进一步描述这一通路，并了解“走”和“留”通路如何调节活动- 依赖性轴突/突触细化，我们提出：目的1：研究Pyk 2和JAK 2的作用， 在生理条件下的轴突/突触细化。目的2：分析电生理学后果 使用条件性KO小鼠在突触细化期间Pyk 2/JAK 2失活。目标3：审查 Pyk 2-JAK 2通路提供了体内小胶质细胞清除非活性轴突的线索。目标4：调查 Stay（SIRP β-CD 47）和Go（Pyk 2-JAK 2）通路在轴突/突触细化中的相互作用 vivo.我们的项目将从分子上描述神经元如何决定在大脑中建立功能性突触连接。 哺乳动物的大脑Pyk 2和JAK 2与多种神经精神疾病相关。许多形式的 包括自闭症和精神分裂症在内的精神疾病与边缘系统的异常改变有关 电路因此，我们的研究也应该产生新的见解的病因和治疗这类疾病。