Mechanisms of Brain Dysfunction in Tuberous Sclerosis

描述(由申请人提供)：神经系统并发症，如癫痫、智力低下和自闭症，是结节性硬化症(TSC)儿童显著发病率和死亡率的常见来源。尤其是癫痫，可能特别具有破坏性，而且往往对现有的治疗方法难以奏效。了解TSC致痫的分子、生化和生理机制对于开发更有效的治疗TSC患者癫痫发作的方法至关重要。癫痫发作和TSC的其他神经症状反映了典型的发育脑异常的存在，如皮质结节。皮质结节含有分化异常的神经胶质细胞和过度的胶质细胞，提示胶质细胞可能在癫痫和脑功能障碍的病理生理学中起重要作用。为了了解胶质细胞在TSC脑功能障碍中的作用，我们建立了一种新的TSC小鼠模型(TSC1[GFAP]CKO小鼠)，在该模型中，TSD在胶质细胞中的失活会导致胶质细胞生长、增殖和分化的异常，神经元的解体，以及严重的癫痫。这项建议的主要目的是阐明TSD中神经胶质细胞缺陷导致TSC1[GFAP]CKO小鼠癫痫发作的分子、生化和电生理机制。我们推测，胶质细胞TSD失活导致星形胶质细胞对谷氨酸和钾的摄取受损，这可能通过导致细胞外谷氨酸和钾稳态异常、兴奋性神经元死亡和过度兴奋的神经元回路而促进癫痫的发生。在这个项目中，我们建议确定TSD失活对TSC1[GFAP]CKO小鼠的1)特定星形胶质细胞谷氨酸转运体和钾通道的表达和功能，2)细胞外谷氨酸和钾水平，3)诱导兴奋性毒性神经元死亡，以及4)神经元兴奋性的影响。此外，先前在TSC中涉及的特定细胞信号通路在调节这些效应中的参与将被调查。在这些研究中，我们将在两个独立研究实验室之间成功、协同合作的背景下，采用分子遗传学、生化和电生理方法的独特组合来解决这些关键问题。除了明确在TSC中导致癫痫发作的细胞和分子机制外，这些研究还应该提供对神经胶质细胞在癫痫发生中的一般作用的更广泛的见解，并确定针对星形胶质细胞的癫痫发作的新疗法，可能使所有癫痫患者受益。由于癫痫影响了大约1%的人，这项研究有可能对公共健康产生重大影响。

DESCRIPTION (provided by applicant): Neurological complications, such as epilepsy, mental retardation, and autism, represent common sources of significant morbidity and mortality in children with tuberous sclerosis complex (TSC). Epilepsy, in particular, can be especially devastating and is often refractory to available treatments. Understanding the molecular, biochemical, and physiological mechanisms underlying epileptogenesis in TSC is critical to the development of more effective therapies for seizures in patients with TSC. Seizures and other neurological symptoms of TSC reflect the presence of characteristic developmental brain abnormalities, such as cortical tubers. Cortical tubers contain abnormally differentiated neuroglial cells and excessive gliosis, suggesting that glial cells may be important in the pathophysiology of epilepsy and brain dysfunction in TSC. To understand the contribution of glial cells to TSC brain dysfunction, we developed a novel mouse model of TSC (Tsc1[GFAP]CKO mice), in which Tsd inactivation in glia results in abnormalities in glial growth, proliferation, and differentation, neuronal disorganization, and severe epilepsy. The primary goal of this proposal is to elucidate the molecular, biochemical, and electrophysiological mechanisms by which glial defects in Tsd lead to seizures in Tsc1[GFAP]CKO mice. We hypothesize that glial Tsd inactivation results in impaired uptake of glutamate and potassium by astrocytes, which may promote epileptogenesis by causing abnormal extracellular glutamate and potassium homeostasis, excitotoxic neuronal death, and hyperexcitable neuronal circuits. In this project, we propose to determine the effects of glial Tsd inactivation on 1) the expression and function of specific astrocyte glutamate transporters and potassium channels, 2) extracellular glutamate and potassium levels, 3) induction of excitotoxic neuronal death, and 4) neuronal excitability on both the single cell and circuit levels in Tsc1[GFAP]CKO mice. Furthermore, the involvement of specific cellular signaling pathways, previously implicated in TSC, in mediating these effects will be investigated. In these studies, we will employ a unique combination of molecular genetic, biochemical, and electrophysiological approaches in the context of a successful, synergistic collaboration between two independent research laboratories to address these critical issues. In addition to defining the cellular and molecular mechanisms causing seizures specifically in TSC, these studies should provide broader insights into the general role of glial cells in epileptogenesis and identify novel therapies for seizures directed at astrocytes that may benefit all patients with epilepsy. Since epilepsy affects approximately 1% of all people, this research has the potential to have significant impact on public health.