Role of ASTN2 in cerebellar circuit function and ASD-related behaviors

ASTN2 在小脑回路功能和 ASD 相关行为中的作用

• 批准号: 10524768
• 负责人: COURT A HULL
• 金额: $ 62.39万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-12-15 至 2025-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10524768
• 关键词: Acute; Address; Adhesions; Animals; Behavioral; Binding; Biological Assay; Brain; CRISPR/Cas technology; Cell membrane; Cells; Cerebellar Cortex; Cerebellum; Copy Number Polymorphism; Defect; Dendritic Spines; Development; Disease model; Distal; Electrophysiology (science); Endocytosis; Family; Fiber; Frequencies; Genes; Genetic Models; Grooming; Heterozygote; Image; In Vitro; Inherited; Interneurons; Ion Channel; Language Delays; Lead; Learning; Lifestyle-related condition; Link; Loss of Heterozygosity; LoxP-flanked allele; Mass Spectrum Analysis; Measurement; Measures; Membrane Proteins; Molecular; Mus; Mutation; Neurodevelopmental Disorder; Pathway interactions; Patients; Physiological; Protein Dynamics; Proteins; Proteome; Proteomics; Purkinje Cells; Reporting; Research; Reversal Learning; Risk Factors; Role; Sensory; Slice; Smell Perception; Social Behavior; Speech Delay; Synapses; Testing; Ultrasonics; Vertebral column; Vesicle; Work; adhesion receptor; autism spectrum disorder; behavior test; conditional knockout; density; design; experimental study; granule cell; insight; loss of function; motor deficit; mouse model; mutant; neurogenetics; novel; open field behavior; overexpression; postsynaptic; presynaptic; protein transport; receptor; social; social communication; synaptic function; trafficking; vocalization

PROJECT SUMMARY The proposed research addresses a critically important question in autism spectrum disorder (ASD) research: how defects in cerebellar circuits contribute to ASD. In particular, it examines the role of the predominantly cerebellar gene ASTN2 in cerebellar circuit function and ASD-related behaviors. Copy number variations (CNVs) in ASTN2 have been identified as a significant risk factor for ASD (Lionel et al, 2014), suggesting that ASTN2 mutations such as those found in patients with ASTN2 CNVs, lead to altered cerebellar synaptic function. In addition, we recently reported a family with a paternally inherited intragenic ASTN2 duplication, which caused a heterozygous loss of function of ASTN2. The family manifested a range of neurodevelopmental disorders, including ASD, learning difficulties and speech and language delay (Behesti et al, 2018). Our cellular and molecular studies on mouse cerebellum show that ASTN2 binds to and regulates the trafficking of multiple synaptic proteins, including Neuroligins, which have been genetically linked to ASDs, and modulates cerebellar Purkinje cell (PC) synaptic activity (Behesti et al, 2018). To provide a genetic model to study cerebellar circuit function, we generated both a global loss of function Astn2 line and a floxed Astn2 line for conditional knockout experiments. New, preliminary evidence indicates that PCs in mice lacking Astn2 have a decrease in evoked excitation relative to inhibition in PCs and reduced PC dendritic spine density, suggesting specific cerebellar circuit defects. In addition, preliminary evidence shows mild motor deficits and defects in USVs and an open field assay, ASD-related behaviors. As other preliminary findings do not indicate major defects in cerebellar development, we hypothesize that the behavioral defects we observed relate to defects in the cerebellar circuitry with underlying changes in receptor trafficking. In the proposed research, we will 1) test how loss of Astn2 alters intrinsic excitability in PCs and the synaptic efficacy of their presynaptic inputs from GCs and molecular layer interneurons, 2) use proteomics to identify changes in the levels of synaptic proteins and live imaging to assess whether such changes relate to changes in the rate of endocytosis, 3) compare changes in PC dendritic branching as well as the regional distribution of PC spines in wild type and mutant animals to provide insight on whether there are changes in the organization of PC inputs during the establishment of the cerebellar circuitry, and 4) analyze changes in social behavior and ultrasonic vocalization in Astn2 wild type, heterozygous and mutant animals. Taken together, the proposed research will provide a new mouse model that allows us to link an ASD-related gene that is predominantly expressed in the cerebellum with specific cerebellar circuit function and molecular pathways.

项目摘要 这项研究提出了自闭症谱系障碍（ASD）中一个至关重要的问题 研究：小脑回路的缺陷如何导致ASD。特别是，它审查了 主要是小脑基因ASTN 2在小脑电路功能和ASD相关行为。副本 ASTN 2中的CNV已经被确定为ASD的重要危险因素（莱昂内尔et 等人，2014年），这表明ASTN 2突变，如在ASTN 2 CNV患者中发现的那些，导致 小脑突触功能的改变此外，我们最近报道了一个家庭， 遗传性基因内ASTN 2重复，导致ASTN 2功能杂合缺失。的 家庭表现出一系列神经发育障碍，包括ASD，学习困难， 言语和语言延迟（Behesti et al，2018）。我们对小鼠的细胞和分子研究 小脑显示ASTN 2结合并调节多种突触蛋白的运输， 包括神经配素，它与自闭症有遗传联系，并调节小脑浦肯野氏 细胞（PC）突触活动（Behesti et al，2018）。为研究小脑神经回路提供遗传模型 函数，我们生成了函数Astn2行的全局丢失和条件Astn2行的floxed 淘汰实验新的初步证据表明，缺乏Astn2的小鼠中的PC具有明显的免疫抑制作用。 PC中诱发兴奋相对于抑制的降低和PC树突棘密度的降低， 说明小脑回路有缺陷此外，初步证据显示， 缺陷和缺陷的USV和一个开放的领域分析，ASD相关的行为。其他初步 研究结果并没有表明小脑发育的主要缺陷，我们假设行为缺陷是由小脑发育的缺陷引起的。 我们观察到的缺陷与小脑回路中的缺陷有关， 贩卖人口 在拟议的研究中，我们将1）测试Astn2的缺失如何改变PC的内在兴奋性， 它们来自GC和分子层中间神经元的突触前输入的突触功效，2）使用 蛋白质组学以确定突触蛋白水平的变化，以及实时成像以评估 这种变化与内吞率的变化有关，3）比较PC树突状细胞的变化， 分支以及PC棘在野生型和突变动物中的区域分布，以提供 了解在建立过程中PC输入的组织是否发生了变化， 小脑回路，以及4）分析Astn 2野生型中社会行为和超声发声的变化 型、杂合子和突变动物。总的来说，拟议的研究将提供一个新的 小鼠模型，使我们能够连接一个ASD相关基因，主要表达在 小脑具有特定的小脑回路功能和分子通路。