Mechanisms of Seizure Genesis in Human Hypothalamic Hamartomas

描述(申请人提供)：下丘脑错构瘤(HH)是一种罕见的先天性人类大脑畸形，与痉挛(或大笑)发作有关，在生命的早期很难诊断，而且出了名的难以接受药物治疗。尽管临床上越来越多地认识到这种情况，但癫痫发生的机制在很大程度上是未知的。重要的是，癫痫发作以前被证明是由错构瘤本身引起的。在巴罗神经研究所，在过去的几年里，我们治疗了100多名患有这种疾病的患者，构成了有史以来最大的单中心系列；我们目前是美国唯一的HH管理多学科项目，也是世界上最活跃的HH项目。因此，我们在研究HH癫痫发生机制方面具有得天独厚的优势。本研究的主要目的是结合细胞电生理、荧光成像、免疫细胞化学和基因表达技术，确定手术切除的HH组织中癫痫发作的发生机制。我们的初步数据表明，HH组织包含两个不同的神经元群体；小的HH神经元表现出内在的起搏样活动，而大的HH神经元是静止的。我们已经证明，当GABAA受体激动剂Muscimol灌流时，大HH神经元的亚群去极化。基于这些有趣的发现，我们推测这种GABAA受体介导的去极化可能是阳离子-氯共转运体KCC2(相对于其表亲NKCC1)表达减少的结果，这种反应可能参与癫痫的发生。具体地说，我们假设麝香酚会增加细胞内钙水平，这一作用可以被NKCC1拮抗剂布美他尼阻断。此外，我们假设HH组织将使用免疫细胞化学和单细胞RT-PCR表达技术显示KCC2相对于NKCC1的表达减少。尽管HH本身并不常见，估计患病率为50,000-100,000人中有1人，但它可能是皮质下癫痫的最佳人类模型，也是理解儿童灾难性癫痫后果的最有趣的模型之一。因此，对这种疾病的详细科学理解对于加快以发育性癫痫为重点的翻译研究具有重要意义。公共卫生相关性：癫痫是一种常见的神经疾病，仅在美国就有300多万人受到影响，许多新诊断的患者是婴儿和儿童。为了开发新的、更有效的治疗方法，有必要了解癫痫在儿科人群中是如何发展的。这项研究建议，虽然专注于一种导致失控癫痫的罕见脑瘤，但可能为癫痫的传播和进展机制提供新的见解。

DESCRIPTION (provided by applicant): The hypothalamic hamartoma (HH) is a rare congenital human brain malformation associated with gelastic (or laughing) seizures that are difficult to diagnosis early in life and are notoriously refractory to medical therapy. Despite increasing clinical recognition of this condition, the mechanisms of epileptogenesis are largely unknown. Importantly, seizures have previously been shown to originate from the hamartoma itself. At the Barrow Neurological Institute, we have treated over 100 patients with this condition over the past several years, constituting the largest single-center series ever; we are currently the only multidisciplinary program for HH management in the United States, and the most active HH program in the world. Thus, we are uniquely positioned to study the mechanisms of epileptogenesis in HH. The major goal of the proposed studies is to determine the mechanisms of seizure genesis in surgically resected HH tissue using a combination of cellular electrophysiological, fluorescence imaging, immunocytochemical and gene expression techniques. Our preliminary data demonstrate that HH tissue contains two distinct populations of neurons; small HH neurons exhibit intrinsic pacemaker-like activity, whereas large HH neurons are quiescent. We have demonstrated that a subpopulation of large HH neurons depolarize when perfused with the GABAA-receptor agonist muscimol. Based on these intriguing findings, we hypothesize that such GABAA-receptor mediated depolarization may be a consequence of decreased expression of the cation-chloride co-transporter KCC2 (relative to its cousin, NKCC1), and that such responses may contribute to seizure genesis. Specifically, we hypothesize that muscimol will increase intracellular calcium levels, an effect that can be blocked by the NKCC1 antagonist bumetanide. Further, we hypothesize that HH tissue will demonstrate decreased expression of KCC2 relative to NKCC1 using immunocytochemical and single-cell RT-PCR expression techniques. Although HH itself is uncommon with an estimated prevalence of 1 in 50,000-100,000 it is perhaps the best human model for subcortical epilepsy, and one of the most intriguing models for understanding the consequences of catastrophic epilepsy in childhood. As such, a detailed scientific understanding of this disorder has major ramifications for expediting translational research focused on the developmental epilepsies. PUBLIC HEALTH RELEVANCE: Epilepsy is a common neurological disorder that afflicts more than 3 million people in the United States alone, and many newly diagnosed patients are infants and children. There is a substantial need to understand how epilepsy develops in the pediatric population in order to develop newer, more effective treatments. This research proposal, while focused on a rare brain tumor that causes uncontrolled seizures, may provide novel insights into the mechanisms of seizure propagation and progression.