Interneuron axonopathy underlies circuit dysfunction in a mouse model of Dravet syndrome

项目总结 德拉韦综合征是一种严重的神经发育障碍，每16,000名儿童中就有一人受到影响，其定义为 难治性癫痫、发育迟缓、智力残疾、自闭症谱系障碍和高 猝死率。德拉维综合征是由编码钠的基因SCN1A突变引起的 (NA+)通道Nav1.1 SCN1A突变如何导致被称为Dravet综合征的临床实体 仍然不清楚；这种知识的差距已经深刻地限制了这种诊断的实际影响。 关于这种疾病患者的治疗、生活质量和长期结果。 先前在包括Scn1a+/-小鼠在内的Dravet综合征实验动物模型中的工作表明， Nav1.1通过大脑皮层GABA能抑制中间神经元功能障碍导致癫痫， 最显著的异常是严重亚型的动作电位产生受损。 中间神经元被称为小白蛋白阳性快速放电中间神经元(PV-IN)。然而，这里提供的数据 这表明，令人惊讶的是，PV-IN功能障碍是暂时的，在早期被限制在一个短暂的时间窗口 发展，随后恢复高频发射。初步数据显示，特定的 Drave氏综合征的病理部位实际上是PV-IN轴突，动作电位传播异常 导致传导延迟和突触衰竭，即使PV-INS已经恢复了产生 高频动作电位。这一发现对小说的发展具有重要的意义 德拉韦综合征的治疗方法，如细胞移植、基因治疗或精确医学。 这项来自早期研究人员实验室的为期5年的新申请使用了创新的神经科学 测试这一新假说的方法，以了解Drave氏综合征的病理机制。建议 实验将确定PV-IN轴突中Na+通道的分子同一性和生理特性 在Scn1a+/-小鼠中，与野生型对照相比，使用来自神经元间轴突的靶向记录和 轴突Na+通道的详细免疫组织化学研究(Aim 1)；确定PV-in对轴突的影响 功能障碍对大脑皮层环路前馈抑制时间的影响(目标2)；并评估其活动 利用活体成像和电生理学对清醒的、表现为Scn1a+/-的小鼠确定的神经元亚群的研究 证实体外研究结果(目标3)。 拟议实验的总体结果将提出关于病理生理学的统一假说。 德拉维综合征的症状。这些知识对于开发新的、有针对性的治疗方法是至关重要的。 目前是不治之症。这一系列研究的长期目标是应用 从实验模型系统到新的、以机械为导向的开发的临床前数据 人类患者的治疗方法。

PROJECT SUMMARY Dravet syndrome is a severe neurodevelopmental disorder that affects 1 in 16,000 children and is defined by treatment-resistant epilepsy, developmental delay, intellectual disability, autism spectrum disorder, and a high rate of sudden death. Dravet syndrome is caused by mutation in the gene SCN1A, which encodes the sodium (Na+) channel Nav1.1 How SCN1A mutation leads to the clinical entity known as Dravet syndrome remains unclear; this gap in knowledge has profoundly limited the practical impact that such a diagnosis has on treatment, quality of life, and long-term outcome for patients with this disorder. Prior work in experimental animal models of Dravet syndrome including Scn1a+/- mice suggests that loss of Nav1.1 leads to epilepsy via dysfunction of GABAergic inhibitory interneurons in the cerebral cortex, with the most prominent identified abnormalities being impaired action potential generation in a critical subtype of interneuron known as the parvalbumin-positive fast-spiking interneuron (PV-IN). However, data presented here indicates that, surprisingly, PV-IN dysfunction is transient, being restricted to a brief time window in early development, with subsequent recovery of high frequency firing. Preliminary data suggests that the specific locus of pathology in Dravet syndrome is actually PV-IN axons, with abnormal action potential propagation leading to conduction delay and synaptic failure, even though PV-INs have recovered the ability to generate action potentials at high frequency. This finding has important implications for the development of novel treatment approaches for Dravet syndrome, such as cell transplantation, gene therapy, or precision medicine. This new 5-year application from the lab of an early stage investigator uses innovative neuroscience approaches to test this new hypothesis as to the mechanism of pathology in Dravet syndrome. Proposed experiments will establish the molecular identity and physiological properties of Na+ channels in PV-IN axons in Scn1a+/- mice as compared to wild-type controls using targeted recordings from interneuron axons and detailed immunohistochemistry of axonal Na+ channels (Aim 1); determine the impact of PV-IN axonal dysfunction on the timing of feedforward inhibition in cerebral cortical circuits (Aim 2); and assess the activity of defined subsets of neurons in awake, behaving Scn1a+/- mice using in vivo imaging and electrophysiology to corroborate in vitro findings (Aim 3). The overall outcome of the proposed experiments will set forth a unifying hypothesis as to the pathophysiology of Dravet syndrome. Such knowledge is critical to the development of novel, targeted therapies for this currently incurable and untreatable disease. The long-term objective of this line of research is to apply preclinical data from experimental model systems to the development of new, mechanistically oriented therapies in human patients.