Synapse Remodeling and Neuronal MHC Class I

突触重塑和神经元 MHC I 类

• 批准号: 8267564
• 负责人: Carla J Shatz
• 金额: $ 40.69万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-02 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8267564
• 关键词: Accounting; Action Potentials; Address; Adult; Alleles; Alzheimer' s Disease; Antibodies; Autistic Disorder; Axon; Binding; Biochemical; Biolistics; Brain; Brain region; Cell surface; Cells; Cellular Immunity; Cerebellum; Childhood; Childhood Neurological Disorder; Communication; Complementary DNA; Development; Disease; Dyslexia; Family; Family member; Fractionation; Functional disorder; Funding; Genbank; Gene Family; Genes; Goals; H2 gene; Health; Hippocampus (Brain); Human; Image; Immune system; Immunologic Receptors; In Vitro; Individual; Inflammation; Investigation; Knock-out; Learning; Learning Disorders; Length; Lentivirus Vector; Life; MHC Class I Genes; Mediating; Memory; Memory Disorders; Modeling; Mus; Mutant Strains Mice; Nervous system structure; Neurons; Ocular Dominance; Pattern; Phenotype; Physiological; Presynaptic Receptors; Process; Proteins; Research; Resolution; Retinal Ganglion Cells; Role; Sensory; Signal Pathway; Specificity; Structure; Surface; Synapses; Synapsins; Synaptic Transmission; Synaptic plasticity; Techniques; Therapeutic; Transgenic Mice; Transgenic Organisms; Translating; Visual system structure; Western Blotting; Work; aging brain; area striata; cell type; critical period; experience; human leukocyte antigen gene; in vivo; loss of function; mutant; neural circuit; neural patterning; novel; overexpression; promoter; protein distribution; relating to nervous system; research study; retinogeniculate; tomography

DESCRIPTION (provided by applicant): The long-term goal of this research is to understand how experience during critical periods of brain development, mediated by the functioning of neural circuits, is translated into lasting structural change in synaptic connectivity. The specific hypothesis under study here is that MHC Class I genes (MHCI; HLA in human) expressed in neurons and at synapses act as a negative regulators to limit activity-dependent synaptic plasticity. The idea that there are molecules and signaling pathways normally working to oppose synaptic plasticity is novel and has significant therapeutic implications. MHCI genes are famous for their role in cell-mediated immunity, but here we study a novel role in neurons. Neuronal MHCI expression in the CNS was discovered unexpectedly in an unbiased screen for genes regulated by neural activity in development. Initial studies in mice provided indirect evidence for a role for these molecules in synaptic plasticity (Huh et al, 2000). Research during the past funding period has revealed that loss of function of just 2 of the 60+ MHCI genes, H2-Kb and/or H2-Db, alters synaptic plasticity rules, unexpectedly often enhancing plasticity and learning (McConnell et al, 2009). Three specific aims are planned: 1) Demonstrate requirement for H2- Db, H2-Kb in synaptic plasticity: Double mutant mice (KbDB-/-) will be studied to determine if these 2 genes can account for many of the changes in synaptic plasticity observed in initial studies of mice lacking surface expression of the majority of MHCI proteins. Mice overexpressing H2-Db will also be examined. Rescue experiments will be performed: Double transgenic mice (NSE-Db+/+; KbDb-/-) have been generated in which Db function is rescued only in neurons. GFP-tagged full-length cDNAs for H2-Kb or H2-Db will be expressed using Lentiviral vectors. 2) Determine if MHCI protein is located at synapses: A working model for neuronal MHCI suggests that MHCI protein located postsynaptically binds across the synapse to presynaptic receptors such as PirB. Immunostaining with MHCI antibodies will be used to examine synaptic distribution. Array Tomography (AT) will be used for higher resolution localization of MHCI protein in direct relation to multiple synaptic markers, as well as to PirB. Biochemical fractionation and Western Blotting will also be used to assess subcellular localization of MHCI and potential interacting partners. 3) Generate a conditional allele of H2-Db for studies of neuronal function: A transgenic mouse will be generated to obtain brain and neuronal cell-type specific knockouts for further study of requirement and specificity of H2-D in neurons. All experiments will make use of electrophysiological, anatomical and imaging studies of mouse visual system in vivo and in vitro to assess activity-dependent synapse development and plasticity. By studying mice with gain- or loss- of MHCI function, these experiments should help to establish whether and where H2-Kb and H2-Db are required normally in neurons and should permit a more systematic investigation of the function of specific MHCI molecules in the brain. 7. Project Narrative/Relevance 2-3 sentences Results from experiments proposed here will broaden understanding of how experience-dependent alterations at synapses, both during critical periods of learning in childhood and in memory formation throughout life, are ultimately encoded in the structure of neural circuits. Understanding molecules and mechanisms involved is crucial for addressing and ultimately curing disorders of learning and memory, from Dyslexia, Autism and other childhood neurological disorders, to Alzheimers' and other memory dysfunction in the aging brain. Moreover, neuronal MHCI is known to be modulated by inflammation and can be recognized by cells of the immune system, providing a direct means of communication between the nervous and the immune systems in both health and disease. PUBLIC HEALTH RELEVANCE: Results from experiments proposed here will broaden understanding of how experience- dependent alterations at synapses, both during critical periods of learning in childhood and in memory formation throughout life, are ultimately encoded in the structure of neural circuits. Understanding molecules and mechanisms involved is crucial for addressing and ultimately curing disorders of learning and memory, from Dyslexia, Autism and other childhood neurological disorders, to Alzheimers' and other memory dysfunction in the aging brain. Moreover, neuronal MHCI is known to be modulated by inflammation and can be recognized by cells of the immune system, providing a direct means of communication between the nervous and the immune systems in both health and disease.

描述（由申请人提供）：本研究的长期目标是了解在大脑发育的关键时期，由神经回路功能介导的经验如何转化为突触连接的持久结构变化。这里研究的具体假设是，MHC I类基因（MHCI;人类HLA）在神经元和突触中表达，作为一个负调节器，以限制活动依赖性突触可塑性。有一些分子和信号通路通常会对抗突触可塑性，这一想法是新颖的，具有重要的治疗意义。MHCI基因以其在细胞介导的免疫中的作用而闻名，但在这里，我们研究了神经元中的新作用。神经元MHCI在中枢神经系统中的表达被意外地发现在一个无偏见的筛选基因调节神经活动的发展。在小鼠中的初步研究为这些分子在突触可塑性中的作用提供了间接证据（Huh等人，2000）。在过去的资助期间的研究已经揭示了60+ MHCI基因中的仅2个H2-Kb和/或H2-Db的功能丧失改变了突触可塑性规则，出乎意料地经常增强可塑性和学习（McConnell等人，2009）。计划了三个具体目标：1）证明在突触可塑性中对H2- Db、H2-Kb的需要：将研究双突变小鼠（KbDB-/-）以确定这两个基因是否可以解释在缺乏大多数MHCI蛋白的表面表达的小鼠的初始研究中观察到的突触可塑性的许多变化。还将检查过表达H2-Db的小鼠。将进行拯救实验：已经产生了双转基因小鼠（NSE-Db+/+; KbDb-/-），其中Db功能仅在神经元中被拯救。将使用慢病毒载体表达GFP标记的H2-Kb或H2-Db的全长cDNA。2)确定MHCI蛋白是否位于突触：神经元MHCI的工作模型表明，位于突触后的MHCI蛋白跨越突触与突触前受体（如PirB）结合。将使用MHCI抗体的免疫染色来检查突触分布。阵列断层扫描（AT）将用于更高分辨率的MHCI蛋白定位，与多种突触标志物以及PirB直接相关。还将使用生化分级分离和蛋白质印迹法来评估MHCI和潜在相互作用伴侣的亚细胞定位。3)产生H2-Db的条件等位基因用于神经元功能的研究：将产生转基因小鼠以获得脑和神经元细胞类型特异性敲除，用于进一步研究神经元中H2-D的需求和特异性。所有实验将利用小鼠视觉系统在体内和体外的电生理、解剖和成像研究来评估活动依赖性突触发育和可塑性。通过研究获得或丧失MHCI功能的小鼠，这些实验应该有助于确定H2-Kb和H2-Db在神经元中是否以及在何处正常需要，并且应该允许对特定MHCI分子在大脑中的功能进行更系统的研究。7.项目叙述/相关性2-3句子本文提出的实验结果将拓宽对经验依赖性突触改变的理解，无论是在儿童学习的关键时期还是在整个生命的记忆形成中，最终都编码在神经回路的结构中。了解所涉及的分子和机制对于解决并最终治愈学习和记忆障碍至关重要，从阅读障碍，自闭症和其他儿童神经系统疾病，到老年痴呆症和衰老大脑中的其他记忆功能障碍。此外，已知神经元MHCI受炎症调节，并且可以被免疫系统的细胞识别，从而在健康和疾病中提供神经系统和免疫系统之间的直接通信手段。 公共卫生相关性：这里提出的实验结果将拓宽对经验依赖性突触改变的理解，无论是在儿童学习的关键时期还是在整个生命的记忆形成中，最终都编码在神经回路的结构中。了解所涉及的分子和机制对于解决并最终治愈学习和记忆障碍至关重要，从阅读障碍，自闭症和其他儿童神经系统疾病，到老年痴呆症和衰老大脑中的其他记忆功能障碍。此外，已知神经元MHCI受炎症调节，并且可以被免疫系统的细胞识别，从而在健康和疾病中提供神经系统和免疫系统之间的直接通信手段。