Seizure-induced Neuronal Loss and Epileptogenesis in the Immature Brain

描述（由申请人提供）：关于未成熟大脑脑损伤后癫痫的发展是否涉及海马形成中的神经元死亡存在争议。一些研究报道，与在成年动物模型中广泛观察到的不同，未成熟的动物可以发展为癫痫，而海马形成中没有可观察到的神经元损失。与此问题直接相关的两个关键方法学问题是：1)确定神经元损失的解剖学方法是否可以检测到可能是细微的或解剖学上限制在海马体亚区域的损伤；2)不同的癫痫检测（癫痫阈值的变化或癫痫监测）是否准确地检测到哪些动物发生了癫痫。本项目利用匹罗卡品锂（LiPC）癫痫动物模型（一种成熟的化学惊厥性癫痫持续状态（SE）模型）来重新检查长时间癫痫发作是否会导致未成熟动物癫痫的发展，特别是神经元死亡是否与这一过程有关。我们建议使用flouo -jade B（一种急性神经元损伤的组织化学标记物）、无偏体视技术的尼氏细胞计数和海马体积磁共振成像作为检测潜在神经元损伤和死亡的三种独立方法。研究人员将使用无线脑电图遥测系统对动物进行持续监测，并对动物的行为进行视频分析，以确保准确评估哪些动物会患上癫痫。我们的方法的优势在于神经元损伤、细胞死亡和癫痫发展之间更有意义的相关性，因为我们的电图监测和解剖分析将比以前的研究更完整。确定神经元丢失或其他因素是否有助于未成熟大脑中的癫痫发生，对于理解儿童在脑损伤后如何发展为癫痫至关重要，并将有助于临床医生开发新的干预措施来治疗和预防这种情况。该提案概述了一个结构化的监督研究经验，这对于帮助申请人发展长期的研究儿童癫痫的职业生涯至关重要。该计划的制定得到了强有力的机构支持，并在癫痫研究人员的指导下提供了良好的环境，这些研究人员在培训成功的科学家方面有着长期的记录。它是专门设计让申请人获得新的研究技能和过渡到一个独立的临床医师科学家。

DESCRIPTION (provided by applicant): There is a controversy over whether neuronal death in the hippocampal formation is involved in the development of epilepsy after a brain insult in the immature brain. Several studies have reported that, unlike what is widely observed in adult animal models, immature animals can develop epilepsy without observable neuronal loss in the hippocampal formation. Two critical methodological issues directly related to this problem are: 1) whether anatomic methods for determining neuronal loss can detect injury that may be subtle or anatomically restricted to sub-regions of the hippocampus and 2) whether the different assays for epilepsy (changes in seizure threshold or seizure monitoring) accurately detect which animals have developed epilepsy. This project utilizes the lithium pilocarpine (LiPC) animal model of epilepsy, a well established model of chemoconvulsant status epilepticus (SE), to reexamine whether prolonged seizures result in the development of epilepsy in immature animals, and, in particular, whether neuronal death is associated with this process. We propose to use Flouro-jade B (a histochemical marker for acute neuronal injury), Nissl cell counting with unbiased stereology techniques, and magnetic resonance imaging of hippocampal volume as three independent methods for detecting potential neuronal injury and death. Animals will be monitored continuously with a wireless EEG telemetry system with video analysis of behavior to ensure an accurate assessment of which animals develop epilepsy. The strength of our approach is that a more meaningful correlation between neuronal injury, cell death, and the development of epilepsy can be made because our electrographic monitoring and anatomical analysis will be more complete than previous studies. Determining whether neuronal loss or other factors contribute to epileptogenesis in the immature brain is critical for understanding how children develop epilepsy after brain insult and will aid clinicians in developing novel interventions for treating and preventing this condition. This proposal outlines a structured supervised research experience that is critical for helping the applicant develop a long-term research career studying pediatric epilepsies. The plan has been developed with strong institutional support, and provides an excellent environment with mentored guidance from epilepsy researchers who have a long track record of training successful scientists. It is specifically designed to allow the applicant to acquire new research skills and transition into an independent clinical physician scientist. PUBLIC HEALTH RELEVANCE: Some researchers have claimed that, unlike what is commonly thought to occur in adults, children can develop the disorder of epilepsy after an insult to the brain without significant brain injury with loss of brain cells. We propose to use an animal model of epilepsy in young rats to induce them to have epilepsy, and then assess for evidence of neuronal injury and loss of neurons using three separate methods (Flouro-jade B-a chemical marker for injured neurons, actual counting of remaining neurons to see if they have been lost, and neuroimaging with magnetic resonance imaging). These experiments will help us better understand the relationship between neuronal injury/death and epilepsy and will provide information about what changes may occur when children get epilepsy.