Cellular and Molecular Mechanisms of Behavioral Dysfunction in a Zebrafish Model of CHARGE Syndrome

电荷综合征斑马鱼模型行为障碍的细胞和分子机制

基本信息

批准号：
10372659
负责人：
Kurt C. Marsden
金额：
$ 41.12万
依托单位：
NORTH CAROLINA STATE UNIVERSITY RALEIGH
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-10 至 2024-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10372659
关键词：
Acoustic Stimulation Acoustics Affect Anxiety Area Atlases Attention deficit hyperactivity disorder Auditory Behavior Behavior Therapy Behavioral Behavioral Mechanisms Behavioral Symptoms Brain Brain region CHARGE syndrome CHD7 gene CRISPR/Cas technology Cells Choanal Atresia Chromatin Remodeling Factor Clutch Size Coloboma Computer software DNA Binding Data Defect Development Disease Ear Elements Eye Fertilization Functional disorder Genes Genetic Genetic Diseases Genetic Transcription Genital Genitalia Genitourinary system Genotype Glutamates Goals Heart Heart Abnormalities Heterozygote Human Genetics Image Impairment Informal Social Control Intellectual functioning disability Label Larva Lighting Link Live Birth Location Measures Mediating Methods Microinjections Microscopy Modeling Molecular Molecular Target Morphology Mutate Nervous system structure Neurodevelopmental Disorder Neurons Obsessive-Compulsive Disorder Optics Pain Pathway interactions Patients Pattern Pharmacology Photic Stimulation Population Proteins Proteome Proteomics Regulation Research Resources Role Sampling Sensory Shapes Site Sleep Disorders Startle Reaction Stereotyping Stimulus Structure System Techniques Therapeutic Tissues Transgenic Organisms Visual Work Zebrafish auditory processing autism spectrum disorder base behavioral phenotyping brain morphology cell type chromatin remodeling clinically relevant common symptom driving behavior human disease in vivo in vivo calcium imaging in vivo imaging loss of function mutation malformation molecular dynamics morphometry motor impairment mutant neural circuit neurobehavioral neurodevelopment neuronal circuitry novel novel diagnostics novel therapeutic intervention relating to nervous system response transcriptome transcriptome sequencing transcriptomics visual information

项目摘要

Project Summary CHARGE syndrome is a rare, multi-system disorder that affects approximately 1 in 10,000 live births. The most common symptoms include ocular coloboma, choanal atresia, heart defects, genital abnormalities, ear malformations, and an array of neuro-behavioral difficulties. Sensory under- or over-load, motor impairments, enhanced pain, sleep disorders, attention deficit hyperactivity disorder (ADHD), obsessive compulsive disorder (OCD), intellectual disability, anxiety, and autism are all frequently observed in CHARGE. Treatments for these behavioral symptoms are limited, highlighting an area of critical need in understanding the mechanisms underlying these defects in order to develop new therapeutic approaches. Two-thirds of CHARGE cases are caused by loss-of-function mutations in chd7, which encodes a DNA-binding, ATP-dependent chromatin remodeling protein. chd7 is highly expressed in the developing vertebrate brain and regulates transcription of several key neurodevelopmental genes, but direct cellular and molecular links between chd7 function and behavioral regulation have not been established. This project leverages the larval zebrafish model, which has rapidly emerged as a powerful system for investigating the development and function of behavioral circuits as well as human genetic disease. Using CRISPR/Cas9 we have established a chd7-null line and have characterized several morphological and behavioral phenotypes reflective of CHARGE. In response to acoustic stimuli, chd7 mutants perform normal short-latency startle responses (SLCs) but have impaired long-latency escape responses (LLCs). Similarly, chd7 mutants respond normally to increases in illumination but are deficient in responding to decreases in illumination. These deficits are independent of morphological defects in the eyes and ears, indicating that chd7 likely regulates specific behavioral circuits in the brain. In Aim 1 we will systematically interrogate the known circuit elements driving these behaviors using in vivo calcium imaging and cell-specific rescue to locate the sites of chd7 action. We will then comprehensively define the brain regions that are dependent on chd7 using whole-brain morphometry and activity analyses. In Aim 2 we will apply state-of- the-art proteomic and transcriptomic approaches to identify molecular pathways that link chd7 with these changes in brain structure and function. By analyzing samples from three developmental timepoints, we will also define the temporal dynamics of these changes. Finally, we will use a systematic CRISPR/Cas9 approach to validate the top proteomics- and transcriptomics-based chd7 targets in vivo by measuring brain development and behavior in mutant larvae. Overall, the results of this work will establish direct links between chd7, its molecular targets, and behavioral circuits. Furthermore, these aims will generate a powerful set of broadly useful resources for interrogating the cellular and molecular bases of chd7-dependent neural development.

项目摘要电荷综合征是一种罕见的多系统疾病，影响10,000个活产中大约1个。最多常见症状包括眼镜瘤，Choanal闭锁，心脏缺陷，生殖器异常，耳朵畸形和一系列神经行为困难。感觉不足或负载，运动障碍，增加疼痛，睡眠障碍，注意力缺陷多动症（ADHD），强迫症（强迫症）经常观察到智力残疾，焦虑和自闭症。这些治疗行为症状是有限的，突出了了解机制的关键需求这些缺陷的基础是为了开发新的治疗方法。三分之二的费用案件是由CHD7的功能丧失突变引起，该突变编码DNA结合，依赖ATP的染色质重塑蛋白。 CHD7在发育中的脊椎动物大脑中高度表达，并调节几个关键的神经发育基因，但是CHD7功能和行为调节尚未建立。该项目利用幼虫斑马鱼模型迅速成为一个有力的系统，用于研究行为回路的发展和功能以及人类遗传疾病。使用CRISPR/CAS9，我们已经建立了CHD7-NULL线并具有表征了反映电荷的几种形态和行为表型。响应声学刺激，CHD7突变体执行正常的短期惊吓反应（SLC），但长期损害逃生响应（LLCS）。同样，CHD7突变体正常对照明的增加反应，但不足在响应照明的减少时。这些缺陷与眼睛中的形态缺陷无关耳朵，表明CHD7可能调节大脑中的特定行为回路。在目标1中，我们将使用体内钙成像和特定于细胞的营救以找到CHD7动作的地点。然后，我们将全面定义大脑区域使用全脑形态法和活性分析依赖CHD7。在AIM 2中，我们将应用 ART蛋白质组学和转录组方法，以鉴定将CHD7与这些联系起来的分子途径大脑结构和功能的变化。通过分析来自三个开发时间点的样本，我们也将定义这些变化的时间动态。最后，我们将使用系统的CRISPR/CAS9方法通过测量大脑发育来验证体内基于蛋白质组学和基于转录组学的CHD7靶标和突变幼虫的行为。总体而言，这项工作的结果将建立CHD7之间的直接联系分子靶标和行为回路。此外，这些目标将产生一套强大的广泛有用的询问CHD7依赖性神经发育的细胞和分子碱基的资源。