Cell-specific Functional and Transcriptomic Analysis of Plasticity Pathways in MECP2-Duplication Syndrome

MECP2 重复综合征可塑性途径的细胞特异性功能和转录组分析

• 批准号: 10593623
• 负责人: Stelios Manolis Smirnakis
• 金额: $ 47.27万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-20 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10593623
• 关键词: Affect; Animals; Behavioral; Brain; Cells; Communities; Control Animal; Data; Dendritic Spines; Disease; Dissection; Epilepsy; Exhibits; Exposure to; Functional disorder; Future; Gene Duplication; Genes; Genotype; Image; In Situ; In Vitro; Individual; Intellectual functioning disability; Intervention; Investigation; Ion Channel; Laboratories; Last Name; Learning; MAP Kinase Gene; Maps; MeCP2 Duplication Syndrome; Measurable; Measures; Mediating; Memory; Messenger RNA; Methods; Methyl-CpG-Binding Protein 2; Modeling; Molecular; Motor Cortex; Mus; Neocortex; Neurons; Neurotransmitters; Output; Pathway interactions; Patients; Penetrance; Pharmacology; Phenotype; Process; Property; RNA; Rett Syndrome; Signal Transduction; Sleep; Synapses; Synaptic plasticity; Syndrome; Time; Tissues; Training; Transcript; Ursidae Family; Vertebral column; Visual; Visual Cortex; Visual evoked cortical potential; area V1; autism spectrum disorder; cell type; cohort; experimental study; flexibility; follow-up; hippocampal pyramidal neuron; image registration; in vivo; in vivo imaging; in vivo two-photon imaging; loss of function; male; motor disorder; mouse model; mutant; neural circuit; protein function; relating to nervous system; response; signal processing; spasticity; therapeutic target; transcriptome; transcriptomics; transmission process; visual map

MECP2-duplication syndrome is caused by duplication of the MECP2 gene leading to progressive intellectual disability, autism, motor dysfunction, spasticity, and epilepsy in males with 100% penetrance. Synaptic plasticity mechanisms are causally implicated in multiple autism spectrum disorders, including Methyl-CpG-binding-protein-2 (MECP2) duplication syndrome. We recently described an abnormal synaptic plasticity phenotype in the Tg1 mouse model of MECP2-duplication syndrome, implicating the Ras/MAPK pathway. However, cell-specific investigation of plasticity pathways involved has yet to be undertaken. Characterizing common plasticity pathway themes of dysfunction in autism spectrum disorders (ASD) is potentially of great value for identifying new targets for intervention. Until recently it has not been possible to measure the transcriptome of individual cells in situ and relate it to their functional properties. Here, we adapt a recently developed high-throughput in-situ mRNA hybridization method (MERFISH), capable of resolving hundreds to thousands of distinct mRNA transcripts per imaging session, to investigate cortical plasticity mechanisms during learning in the Tg1 mouse model of the MECP2-duplication syndrome. We use a robust visual training paradigm introduced by M Bear, consisting of multiple presentations of a high contrast flickering grating at a specific orientation. This is known to form a true memory trace in area V1+ that requires sleep for consolidation, has a behavioral correlate and shares core molecular features with LTP. Aim-1: Compare how visual cortex circuits malfunction with learning in MECP2-duplication mice vs in littermate controls. Visual response properties, intra- and inter-layer functional connectivity profiles will be measured within area V1 and compared across genotypes before and after training. Hypothesis: The capacity of cortical circuits for learning differs in mutant vs control mice in a laminar specific way. Inter-layer functional connectivity profiles will be abnormal in MECP2-duplication syndrome resulting in abnormal signal transmission and processing across cortical laminae. This will be exacerbated post-training leading to rigid, less flexible, neural responses. Aim-2: Compare single-cell resolved transcriptomic profiles of plasticity and neurotransmitter pathways in the visual cortex of MECP2-duplication vs control animals undergoing the training paradigm introduced in aim-1. Relate neuronal responses obtained in aim-1 to cell-specific transcriptomic profiles obtained from the same neurons. Hypothesis: We will acquire a more complete, cell-specific, picture of how mechanisms of plasticity fail in the neocortex of the mouse model of MECP2-duplication syndrome. Data obtained will generate hypotheses for follow up experiments, helping to identify targets for future pharmacologic interventions. IMPACT: Relate for the first time, cell-specific plasticity & neurotransmitter/ion-channel transcriptomic profiles with cell properties acquired in vivo during learning, characterizing the mechanisms underlying cortical dysfunction in MECP2-duplication syndrome. Data will be made available to the scientific community. In time, additional models of autism will be brought into this framework, yielding a roadmap for identifying appropriate therapeutic targets in ASD.

MeCP2-重复综合征是由MECP2基因复制导致智力进行性增加引起的 100%外显的男性的残疾、自闭症、运动障碍、痉挛和癫痫。突触可塑性 与多种自闭症谱系障碍有关的机制，包括甲基CpG结合蛋白-2 (MeCP2)重复综合征。我们最近在Tg1中描述了一种异常突触可塑性表型 MECP2-复制综合征小鼠模型，涉及RAS/MAPK通路。然而，特定于细胞 对所涉及的可塑性途径的调查还有待进行。常见可塑性途径的特征 自闭症谱系障碍(ASD)的功能障碍主题对于确定新的治疗靶点具有潜在的重要价值 干预。直到最近，还不可能在原位测量单个细胞的转录组并将其联系起来 它与它们的功能特性有关。在这里，我们采用了最近发展起来的高通量原位mRNA杂交 方法(MerFish)，能够在每个成像过程中解析成百上千个不同的mRNA转录本， Tg1小鼠MECP2重复模型学习过程中皮质可塑性机制的研究 综合症。我们使用由M Bear引入的健壮的视觉训练范例，包括多个演示 在特定方向上的高对比度闪烁光栅。已知这在区域V1+中形成了真实的存储器轨迹， 需要睡眠来巩固，具有行为相关性，并与LTP共享核心分子特征。 目的-1：比较MECP2重复小鼠与仔鼠视觉皮层回路功能障碍与学习的差异 控制。视觉响应属性、层内和层间功能连接配置文件将在 V1区，并比较训练前后各基因型别的差异。假说：大脑皮质环路的容量 突变小鼠和对照小鼠的学习能力以层状结构特有的方式不同。层间功能连接配置文件将为 MECP2重复综合征异常导致跨皮质信号传递和处理异常 片层。这将在训练后加剧，导致僵硬的、更不灵活的神经反应。 目的-2：比较视觉中单细胞分辨的可塑性和神经递质通路的转录图谱 MECP2的皮层-重复与对照组动物进行AIM-1中介绍的训练范式。关联 在AIM-1中获得的神经元对从相同神经元获得的细胞特异性转录图谱的反应。 假设：我们将获得一个更完整的，细胞特有的，关于可塑性机制如何在 MECP2复制综合征小鼠模型的新皮质。所获得的数据将生成以下假设 UP实验，帮助确定未来药物干预的靶点。 影响：首次将细胞特异性可塑性和神经递质/离子通道转录图谱与 在学习过程中获得的体内细胞特性，表征了导致大脑皮质功能障碍的机制 MeCP2-重复综合征。将向科学界提供数据。随着时间的推移，其他型号的 自闭症将被纳入这一框架，为确定自闭症的适当治疗靶点提供路线图。