Molecular and circuit defects underlying different SCN2A mutations and ASDs

不同 SCN2A 突变和自闭症谱系障碍 (ASD) 背后的分子和电路缺陷

• 批准号: 10596085
• 负责人: Geoffrey S Pitt
• 金额: $ 64.34万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-07 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10596085
• 关键词: Action Potentials; Adult; Affect; Amygdaloid structure; Axon; Behavior; Behavioral; Biological Assay; Brain; C-terminal; CRISPR/Cas technology; Calmodulin; Cells; Characteristics; Clinical; Compensation; Data; Defect; Detection; Development; Diagnosis; Disease model; Distal; Electrophysiology (science); Equilibrium; Fiber; Functional disorder; Funding; Genes; Genetic; Goals; Impairment; Interneurons; Link; Modeling; Molecular; Mus; Mutation; Neocortex; Neurons; Outcome; Output; Photometry; Point Mutation; Property; Protein Truncation; Reporting; Roentgen Rays; SCN2A protein; SCN8A gene; Social Behavior; Social Interaction; Sodium Channel; Structure; Synapses; Techniques; Testing; Therapeutic; Variant; Weaning; anxiety-like behavior; autism spectrum disorder; behavioral phenotyping; designer receptors exclusively activated by designer drugs; effective therapy; endophenotype; excitatory neuron; exome sequencing; genomic locus; in vivo; insight; loss of function; loss of function mutation; mouse model; mutant; neocortical; neural circuit; neuronal excitability; novel; restoration; social deficits; tool; voltage

ABSTRACT The recent wave of whole exome sequencing studies places SCN2A, which encodes the neuronal voltage- gated Na+ channel pore-forming α subunit NaV1.2, near top of the list of genetic loci linked to autism spectrum disorders (ASDs). On the one hand, that NaV1.2 is an essential Na+ channel responsible for initiating action potentials within excitatory neurons in the developing brain provides a rationale for the prominence of SCN2A. On the other, most SCN2A mutations associated with ASDs are loss-of-function and predicted to decrease neuronal excitability, an outcome that would lower the neocortical excitation/inhibition (E/I) balance and thus contrast with the generally accepted model that behavior defects in ASDs, such as social dysfunction, result from an increased E/I balance. This conundrum persists because of the absence of Scn2a mouse models that reveal ASD-associated endophenotypes, thus limiting our ability to dissect the cellular electrophysiological defects associated with Scn2a loss-of-function mutations and the consequent circuit level dysfunctions that lead to ASD-associated behaviors. Building on x-ray crystal structures of key regulatory components of NaV1.2 that we solved and analyzed during the previous funding period, we obtained specific insights into how ASD- associated mutations in NaV1.2 perturb channel function and alter E/I balance. Further, we generated two novel Scn2a mouse models by CRISPR/Cas9 to test the specific contribution of Scn2a mutations in vivo. Initial analyses of these models reveal abnormal Na+ channel function, decreased cortical neuron excitability, and dysfunctional behaviors consistent with ASDs, while simultaneously demonstrating informative differences between the two models. These models provide a unique set of tools that will allow us to trace abnormal channel function through altered neuronal electrical activity to the consequent circuit-level dysfunction and the resulting ASD endophenotypes. We propose to exploit these novel Scn2a mutant models for the following Aims: 1) We will obtain detailed information about their neuronal electrophysiological characteristics and synaptic properties, thereby defining how Scn2a mutations perturb neuronal function. 2) We will employ fiber photometry and chemogenetic tools (DREADDs) to test whether the Scn2a mutations decrease excitatory drive to the basolateral amygdala and thereby produce the social dysfunction and impaired danger detection observed in our Scn2a mouse models. 3) We will exploit our initial electrophysiological findings to test a potential therapeutic strategy in which we aim to counteract the reduced Na+ current associated with ASD-associated SCN2A loss-of-function mutations. Our overall goals are to define the range of cellular dysfunction that results from Scn2a mutations and trace those abnormalities through the circuit level to behavioral manifestations.

抽象的 最近的整个外显子组测序研究的浪潮使SCN2A编码了神经元电压 - 门控na+通道孔形成α亚基NAV1.2，靠近与自闭症谱相关的遗传局部列表的顶部 疾病（ASD）。一方面，NAV1.2是负责启动动作的必不可少的NA+渠道 发育中的大脑兴奋性神经元内的电势为SCN2A突出的理由提供了理由。 另一方面，与ASD相关的大多数SCN2A突变是功能丧失，预计会减少 神经元的兴奋，一种将降低新皮质兴奋/抑制（E/I）平衡的结果，因此 与ASD中的行为缺陷（例如社交功能障碍）的公认模型形成对比，结果 从增加的E/I平衡。由于没有SCN2A鼠标模型，这种难题仍然存在 揭示与ASD相关的内表型，从而限制了我们剖析细胞生理学的能力 与SCN2A功能丧失突变相关的缺陷以及随之而来的电路水平功能障碍 导致与ASD相关的行为。基于NAV1.2关键调节组件的X射线晶体结构 我们在上一个资金期间解决和分析了，我们获得了有关如何ASD-的特定见解 NAV1.2扰动通道功能和更改E/I平衡中的相关突变。此外，我们产生了两个 CRISPR/CAS9的新型SCN2A小鼠模型测试体内SCN2A突变的特定贡献。最初的 对这些模型的分析表明，Na+通道功能异常，令人兴奋的皮质神经元降低，并且 功能失调的行为与ASD一致，同时证明有益的差异 在两个模型之间。这些模型提供了一套独特的工具，使我们能够追踪异常 通过改变神经元电活动的通道功能，导致电路级功能障碍和 由此产生的ASD内表型。 我们建议探索以下目的的这些新颖的SCN2A突变模型：1）我们将获得详细的 有关其神经元电生理特征和突触特性的信息，从而定义 SCN2A突变如何扰动神经元功能。 2）我们将采用纤维光度法和化学遗传学工具 （Dreadds）测试SCN2A突变是否会降低兴奋性驱动器到基础杏仁核和 从而产生社会功能障碍并在我们的SCN2A小鼠模型中观察到的危险检测受损。 3）我们将利用最初的电生理发现来测试我们针对的潜在理论策略 为了抵消与与ASD相关的SCN2A功能丧失突变相关的降低的Na+电流。我们的 总体目标是定义由SCN2A突变引起的细胞功能障碍的范围，并追踪这些功能 通过电路水平到行为表现的异常。

项目成果

L-type calcium channels and neuropsychiatric diseases: Insights into genetic risk variant-associated genomic regulation and impact on brain development.

• DOI: 10.1080/19336950.2023.2176984
• 发表时间: 2023-12
• 期刊: CHANNELS
• 影响因子: 3.3
• 作者: Baker, Madelyn R.;Lee, Andrew D. S.;Rajadhyaksha, Anjali M.
• 通讯作者: Rajadhyaksha, Anjali M.

Direct Observation of Compartment-Specific Localization and Dynamics of Voltage-Gated Sodium Channels.

• DOI: 10.1523/jneurosci.0086-22.2022
• 发表时间: 2022-07-13
• 期刊: JOURNAL OF NEUROSCIENCE
• 影响因子: 5.3
• 作者: Liu, Hui;Wang, Hong-Gang;Pitt, Geoffrey;Liu, Zhe
• 通讯作者: Liu, Zhe

Scn2a severe hypomorphic mutation decreases excitatory synaptic input and causes autism-associated behaviors.

• DOI: 10.1172/jci.insight.150698
• 发表时间: 2021-08-09
• 期刊: JCI insight
• 影响因子: 8
• 作者: Wang HG;Bavley CC;Li A;Jones RM;Hackett J;Bayleyen Y;Lee FS;Rajadhyaksha AM;Pitt GS
• 通讯作者: Pitt GS