PATHOPHYSIOLOGY OF DEVELOPING DYSPLASTIC HUMAN CORTEX

基于神经影像学（MRI和PET）和切除组织的检查，皮质发育不良（CD）已被认为是癫痫的主要病理基础。 加州大学洛杉矶分校的儿科癫痫手术计划治疗患有顽固性癫痫发作的儿童人群，在我们的经验中，一半的病例患有CD，而其余的病例有中风和脑炎（非CD）等病因。 关于发育不良皮质中细胞的电生理特性、CD和非CD区域致癫痫的潜在机制以及正常的出生后皮质发育如何影响致癫痫新皮质，我们知之甚少。 该研究项目旨在通过对这些儿童的手术组织进行协调的形态学和电生理学研究来解决这些基本的病理生理学问题。 我们的工作假设是，异常的新皮层是癫痫，因为兴奋和抑制过程之间的不平衡，皮层轴突回路异常组织的病理过程的结果。 拟议的研究将有两个目标：1）检查CD和非CD组织中兴奋性和抑制性过程发生改变的假设; 2）评估人类出生后发育期间新皮层神经元中兴奋性和抑制性过程的发育。 每个特定目标将利用类似的实验设计，结合术前临床和术中ECoG，以确定将研究哪些区域进行细胞内电生理学和形态学评估。 使用最先进的形态学和电生理学技术，实验将比较CD和非CD新皮层中“异常”出现的神经元，以确定：1）N-甲基-D-天冬氨酸（NMDA）和非NMDA的[α-氨基-3-羟基-5-甲基-4-异恶唑丙酸（AMPA）和红藻氨酸（KA）]离子型谷氨酸受体是否存在差异和/或改变; 2）是否存在抑制性神经元数量的差异和/或抑制性GABAA受体的增加;以及3）组织中癫痫发作的一种可能病因是否来自异常轴突回路。 这些目标将通过在视觉上识别的发育异常和正常外观的细胞中检查来实现：1）由NMDA、AMPA、KA和GABAA受体激活诱导的电生理膜电流的改变; 2）使用原位杂交和免疫组织化学技术表达NMDA、非NMDA和GABAA亚单位的神经元的数量和位置;和3）树突和轴突连接，如通过用生物胞素或荧光黄填充记录的发育不良神经元和正常外观细胞所鉴定的。 这些发现将提供重要的基础信息，为了解发育不良的新皮质的病理生理学，提出了顽固性儿童癫痫的病理机制，并提供了深入了解可能的方法来控制儿童癫痫发作导致的CD和非CD。

Based on neuroimaging (MRI and PET) and examination of surgically-resected tissue, cortical dysplasia (CD) has become recognized as a major pathological substrate in epilepsy. UCLA's Pediatric Epilepsy Surgery Program treats populations of children with intractable seizures, and in our experience one-half of the cases have CD while the remaining have etiologies such as strokes and encephalitis (non-CD). Very little is known about the electrophysiological properties of cells in dysplastic cortex, the underlying mechanism(s) that make CD and non-CD areas epileptogenic, and how normal postnatal cortical development affects epileptogenic neocortex. This research project is designed to address these fundamental pathophysiologic questions by performing coordinated morphologic and electrophysiologic studies on surgical tissue from these children. Our working hypothesis is that abnormal neocortex is epileptogenic because of an imbalance between excitatory and inhibitory processes and that cortical axon circuits are abnormally organized as a consequence of the pathologic process. The proposed studies will have two goals: 1) To examine the hypothesis that excitatory and inhibitory processes are altered in CD and non-CD tissue; and 2) to assess development of excitatory and inhibitory processes in neocortical neurons during human postnatal development. Each specific aim will utilize a similar experimental design incorporating pre-surgical clinical and intraoperative ECoG to determine which regions are to be studied for intracellular electrophysiology and morphologic assessments. Using state-of-the-art morphologic and electrophysiologic techniques, experiments will compare "abnormal"-appearing neurons in CD and non-CD neocortex to determine: 1) If there are differences and/or alterations in N-methyl-D-aspartate (NMDA) and non-NMDA alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and kainate (KA)] ionotropic glutamate receptors; 2) if there are differences in the number of inhibitory neurons and/or an increase in inhibitory GABAA receptors; and 3) if one possible etiology of seizures in the tissue is from abnormal axon circuitry. These goals will be accomplished by examining in visually-identified dysplastic and normal-appearing cells: 1) The alterations in electrophysiologic membrane currents induced by activation of NMDA, AMPA, KA, and GABAA receptors; 2) the number and location of neurons expressing NMDA, non-NMDA, and GABAA subunits using in situ hybridization and immunohistochemical techniques; and 3) the dendritic and axonal connections as identified by filling recorded dysplastic neurons and normal-appearing cells with biocytin or Lucifer Yellow. The findings will provide important fundamental information necessary for the understanding of the pathophysiology of dysplastic neocortex, suggest pathologic mechanisms of intractable childhood epilepsy, and provide insights into possible ways of controlling childhood seizures resulting from CD and non-CD.