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

项目摘要 Dravet综合征是一种严重的神经发育障碍，每16，000名儿童中就有1名受影响，其定义为： 难治性癫痫，发育迟缓，智力残疾，自闭症谱系障碍，以及高 猝死率。Dravet综合征是由SCN 1A基因突变引起的，SCN 1A基因编码钠， (Na+）通道Nav1.1 SCN 1A突变如何导致Dravet综合征的临床实体 目前尚不清楚;这种知识上的差距极大地限制了这种诊断的实际影响。 治疗、生活质量和长期预后。 先前在Dravet综合征实验动物模型（包括Scn 1a +/-小鼠）中的研究表明， Nav1.1通过大脑皮层中GABA能抑制性中间神经元的功能障碍导致癫痫， 最突出的确定的异常是受损的动作电位产生的关键亚型， 这种中间神经元称为小清蛋白阳性快速尖峰中间神经元（PV-IN）。然而，这里提供的数据 这表明，令人惊讶的是，PV-IN功能障碍是短暂的，仅限于早期的短暂时间窗。 发展，随后恢复高频率射击。初步数据显示， Dravet综合征的病理部位实际上是PV-IN轴突，具有异常动作电位传播 导致传导延迟和突触失败，即使PV-IN已经恢复了产生突触的能力， 高频动作电位这一发现对小说的发展具有重要意义 Dravet综合征的治疗方法，如细胞移植，基因治疗或精准医学。 这个新的5年应用程序从实验室的早期阶段的研究人员使用创新的神经科学 方法来测试这一新的假设的病理机制Dravet综合征。提出 实验将建立PV-IN轴突中Na+通道的分子特性和生理特性 在Scn 1a +/-小鼠中，与野生型对照相比，使用来自中间神经元轴突的靶向记录， 轴突Na+通道的详细免疫组织化学（目的1）;确定PV-IN轴突的影响 功能障碍的前馈抑制的时间在大脑皮层电路（目的2）;并评估活动 使用体内成像和电生理学，在清醒的行为Scn 1a +/-小鼠中确定神经元子集 证实体外结果（目标3）。 拟议的实验的总体结果将提出一个统一的假设，以病理生理学 Dravet综合征这些知识对于开发新的靶向治疗至关重要。 目前无法治愈的疾病。这项研究的长期目标是 从实验模型系统的临床前数据到新的，以机制为导向的 治疗人类患者。

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.