A systematic test of the relation of ASD heterogeneity to synaptic function

描述（由申请人提供）：尽管自闭症谱系障碍（asd）具有高度遗传性，但asd是异质性的，在很大一部分患者中，没有单一的遗传原因导致asd。相反，在asd中发现了异质遗传变化，包括许多单基因突变和拷贝数变异（CNVs）。因此，一个关键的问题是，不同的遗传变化是通过多个独立的致病途径导致asd，还是asd的各种遗传变化汇聚到一个单一的致病途径上。编码突触蛋白的几个独立基因突变，如神经素-1、神经素和SHANK3，表明自闭症可能通常涉及神经元之间突触通讯的损伤。然而，在自闭症谱系障碍中观察到的大多数其他遗传变化对突触没有已知的影响事实上，对任何大脑功能都没有已知的影响。因此，本研究的主要目标是对asd遗传变化的突触效应进行大规模、系统的分析。该方法将是过表达（模拟基因复制）或敲低（模拟基因失活）与81个ASD候选基因相对应的mrna，并使用标准化分析来测试这些操作对突触的影响。细胞活力、神经元发育、突触密度和突触功能将在培养的小鼠神经元中进行评估，使用PI实验室中建立的光学和电生理分析。在小鼠海马体中立体定向注射慢病毒或在子宫内电穿孔后，将通过相同的操作在体内研究在培养神经元中发现的影响神经元发育、突触形成或突触功能的基因。然后将使用PI实验室建立的标准电生理技术，在这些小鼠的急性切片中检查突触功能和可塑性的变化。该项目的所有结果将发布在一个专门的公共网站上，所有生成的试剂将随时提供给科学界。该项目的结果将为ASD候选基因的功能提供一个标准化的参考点，并为以下假设提供初步测试：尽管ASD的临床和遗传异质性，但它涉及一种共同的（如果不同的话）作用于大脑突触通信的途径。

DESCRIPTION (provided by applicant): Although autism spectrum disorders (ASDs) are highly heritable, ASDs are heterogeneous, and no single genetic cause contributes to ASDs in a large proportion of patients. Instead, heterogeneous genetic changes, including many single gene mutations and copy-number variations (CNVs) are found in ASDs. Thus, a key question is whether different genetic changes contribute to ASDs via multiple, independent, pathogenic pathways, or whether the various genetic changes in ASDs converge onto a single pathogenic pathway. Several independent mutations in genes encoding synaptic proteins, such as neurexin-1, neuroligins, and SHANK3, suggested that ASDs may generally involve an impairment of synaptic communication between neurons. However, most of the other genetic changes observed in ASDs have no known effect on synapses in fact, have no known effect on any brain function. Thus, the major goal of the present proposal is to conduct a large scale, systematic analysis of the synaptic effects of genetic changes in ASDs. The approach will be to over express (to mimic gene duplications) or knock down (to mimic gene inactivations) mRNAs corresponding to 81 ASD candidate genes, and to test the effect of these manipulations on synapses using standardized assays. Cell viability, neuronal development, synapse density and synapse function will be assessed in cultured mouse neurons using optical and electro-physiological assays that are well established in the PI's laboratories. Genes that were found to affect neuronal development, synapse formation, or synapse function in cultured neurons will be studied by the same manipulations in vivo after stereotaxic injection of lentiviruses into the mouse hippocampus, or after in utero electroporation. Changes in synapse function and plasticity will then be examined in acute slices from these mice using standard electrophysiological techniques well established in the PI's laboratories. All results from this project will be posted on a dedicated public website, and all reagents generated will be made readily available to the scientific community. The results of this project will provide a standardized reference point for the function of ASD candidate genes, and provide an initial test of the hypothesis that despite their clinical and genetic heterogenity, ASDs involve a common, if diverse, pathway acting on synaptic communication in the brain. PUBLIC HEALTH RELEVANCE: Autism spectrum disorders are known to be clinically and genetically heterogeneous, but it is unclear whether these two types of heterogeneity are related, and how specifically the various genetic changes affect brain function. This project will address these issues by studying the changes in neuron-to-neuron communication caused by the genes associated with autism.