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Testing the role of intracellular vs. cell surface mGlu5 in models of synaptic plasticity using CRISPR-modified mice

使用 CRISPR 修饰的小鼠测试细胞内与细胞表面 mGlu5 在突触可塑性模型中的作用

基本信息

批准号：
10372104
负责人：
KAREN L O'MALLEY
金额：
$ 31.5万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-01 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10372104
关键词：
Acoustics Address Agonist Animal Behavior Animal Disease Models Animal Model Animals Anxiety Behavior Behavioral Biological Assay Biological Models Biology Cancer Therapy Evaluation Program Candidate Disease Gene Cell surface Cells Clinical Clinical Trials Clustered Regularly Interspaced Short Palindromic Repeats Corpus striatum structure Couples Data Development Disease Disease model Drug Targeting Exhibits Frequencies Future G-Protein-Coupled Receptors GRM5 gene Genetic Models Glutamate Transporter Glutamates Goals Human In Vitro Intellectual functioning disability Intracellular Membranes Knowledge Lead Learning Link Location Long-Term Depression MAPK3 gene Mediating Medical Economics Membrane Memory Mental Depression Metabotropic Glutamate Receptors Modeling Motor Mus Neurons Neurotransmitters Outcome Pathway interactions Persons Pharmaceutical Preparations Pharmacology Phenocopy Phenotype Phosphorylation Physiological Play Preparation Process Receptor Signaling Regulation Reporting Research Rewards Role Signal Pathway Signal Transduction Slice Swimming Synapses Synaptic plasticity System Testing Therapeutic Time Up-Regulation Variant Western Blotting Work antagonist autism spectrum disorder base cell behavior chronic pain cost depressive symptoms expectation experimental study genetic approach human disease improved improved outcome in vivo insight metabotropic glutamate receptor 5 neuropsychiatric disorder new therapeutic target novel therapeutics pain behavior quisqualate receptor receptor-mediated signaling response social success therapeutic target tool transcriptome transcriptome sequencing

项目摘要

ABSTRACT Neuropsychiatric disorders such as autism spectrum disorder (ASD), anxiety and depression cost hundreds of billions of dollars each year in medical, economic and social costs. Despite progress, treatments for these conditions remain limited. Hence safer, more specific drugs would have high clinical utility. One critical, drug-able target is the metabotropic glutamate receptor, mGlu5, antagonists of which robustly improve animal models of these disorders; however the results of initial clinical trials have been mixed. Rather than invalidating mGlu5 as a therapeutic target, such results highlight the need for a better understanding of mGlu5 function, including its subcellular localization. For example, we have shown that 70-90% of mGlu5 is located on intracellular membranes, where it couples to signaling systems distinct from those of its cell surface counterpart. Importantly, intracellular mGu5 is sufficient for establishing long term depression (LTD), a form of synaptic plasticity that is dysfunctional in ASD, anxiety and depression. The objective of the proposed research is to establish the role that intracellular mGlu5 plays by identifying differences in its function on the cell surface vs. inside the cell, in vitro, ex vivo and in vivo. Our central hypothesis is that mGlu5 interacts with different signaling pathways depending on where it is located and that this results in distinct regulation of synaptic processes and related behaviors. To test this hypothesis we have developed a genetically restricted mouse line in which mGlu5 is present only on intracellular membranes (mGlu5IM). We will use this newly derived animal line in the following aims. In Aim 1, we will determine whether mGlu5IM vs. mGlu5WT or mGlu5KO has different effects on signaling pathways in vitro (cultured neurons) by testing proposed candidate genes and by using unbiased transcriptome profiling (RNA-Seq) to identify new pathways. In Aim 2, we will use ex vivo slice preparations to determine whether mGlu5IM vs. mGlu5WT or mGlu5KO variants play unique roles and signal through distinctive pathways in synaptic processes such as LTD. In Aim 3, we will determine whether the variant mice exhibit unique effects on sensorimotor, anxiety-like and depressive-like behaviors as well as on operant models of reward. This would be the first report of location- specific functions of intracellular mGlu5 in vivo. As mGlu5 is one of a growing number of receptors that signal from inside the cell, the proposed experiments will also enhance knowledge of other intracellular receptors. Future studies targeting drugs to intracellular vs. cell- surface-localized receptors are expected to lead to the development of better drugs for mGlu5- modulated disorders such as ASD and depression,

摘要神经精神障碍，如自闭症谱系障碍(ASD)、焦虑和抑郁每年造成数千亿美元的医疗、经济和社会成本。尽管尽管取得了进展，但对这些疾病的治疗仍然有限。因此，更安全、更有针对性的药物会有很高的临床实用价值。一个关键的、可用药的靶点是代谢性谷氨酸受体mGlu5，其拮抗剂有力地改善了这些疾病的动物模型；然而，初步临床试验的结果好坏参半。而不是使mGlu5作为治疗靶点，这样的结果强调了更好地了解mGlu5功能的必要性，包括它的亚细胞定位。例如，我们已经证明，70%-90%的mGlu5是位于细胞内膜上，在那里它耦合到与那些不同于它的细胞表面对应物。重要的是，细胞内的mGu5足以建立长期的抑郁症(LTD)，一种突触可塑性的形式，在ASD、焦虑和抑郁症。这项拟议研究的目标是确定细胞内 MGlu5通过识别其在细胞表面和细胞内的功能差异来发挥作用体外、体外和体内。我们的中心假设是mGlu5与不同的信号相互作用路径取决于它所在的位置，这导致对突触的不同调节流程和相关行为。为了验证这一假设，我们开发了一种从基因上 MGlu5仅存在于细胞膜上的限制性小鼠品系(MGlu5IM)。我们将在以下目标中使用这一新衍生的动物品系。在目标1中，我们将确定是否 MGlu5IM与mGlu5WT或mGlu5KO对体外培养的信号通路的影响不同神经元)通过测试建议的候选基因和使用无偏转录组图谱 (RNA-Seq)以确定新的途径。在目标2中，我们将使用体外切片制剂来确定mGlu5IM与mGlu5WT或mGlu5KO变体是否发挥独特的作用和信号通过突触过程中独特的途径，如LTD。在目标3中，我们将确定变异小鼠是否对感觉运动、焦虑样和抑郁样表现出独特的影响行为以及奖励的可操作模型。这将是第一份关于位置的报告- 细胞内mGlu5在体内的特殊功能。因为mGlu5是越来越多的从细胞内部发出信号的受体，拟议中的实验也将增强其他细胞内受体的知识。未来针对细胞内药物与细胞药物的研究- 表面定位的受体有望导致更好的mGlu5-药物的开发调节性疾病，如自闭症和抑郁症，