Seizure-induced Neuronal Loss and Epileptogenesis in the Immature Brain

描述（由申请人提供）：关于海马结构中的神经元死亡是否参与未成熟脑中脑损伤后癫痫的发展存在争议。几项研究报道，与成年动物模型中广泛观察到的不同，未成年动物可以发展癫痫，而海马结构中没有可观察到的神经元损失。与该问题直接相关的两个关键方法学问题是：1）用于确定神经元损失的解剖学方法是否可以检测可能是细微的或解剖学上限制于海马子区域的损伤，以及2）癫痫的不同测定（癫痫发作阈值或癫痫发作监测的变化）是否准确地检测哪些动物已经发展成癫痫。本项目利用匹鲁卡品锂（LiPC）癫痫动物模型，一个完善的模型，化学惊厥癫痫持续状态（SE），重新检查是否长时间的癫痫发作导致癫痫的发展在未成熟的动物，特别是，是否神经元死亡与此过程相关。我们建议使用Flouro-jade B（急性神经元损伤的组织化学标记物），无偏体视学技术的Nissl细胞计数和海马体积的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。将使用无线EEG遥测系统对动物进行连续监测，并对行为进行视频分析，以确保准确评估哪些动物发生癫痫。我们的方法的优势在于，神经元损伤，细胞死亡和癫痫的发展之间的更有意义的相关性，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了解儿童如何在脑损伤后发生癫痫至关重要，并将有助于临床医生开发治疗和预防这种疾病的新干预措施。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研究儿科癫痫的研究生涯至关重要。该计划是在强大的机构支持下制定的，并提供了一个良好的环境，由癫痫研究人员提供指导，他们在培训成功的科学家方面有着长期的记录。它是专门设计的，让申请人获得新的研究技能和过渡到一个独立的临床医生科学家。 公共卫生相关性：一些研究人员声称，与通常认为发生在成年人身上的情况不同，儿童在大脑受到损伤后可能会患上癫痫症，而不会造成脑细胞的严重损伤。我们建议使用癫痫的动物模型在年轻的大鼠诱导他们有癫痫，然后评估神经元损伤和损失的神经元的证据，使用三种不同的方法（Flouro-jade B-一种化学标记受损的神经元，剩余的神经元的实际计数，看看他们是否已经失去了，神经成像与磁共振成像）。这些实验将帮助我们更好地了解神经元损伤/死亡和癫痫之间的关系，并将提供有关儿童癫痫时可能发生的变化的信息。

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.