Mechanisms regulating KCC2 hypofunction during refractory seizures in a mouse model of ischemic neonatal seizures

项目摘要/摘要： 这项研究项目的长期目标是了解更广泛的 苯巴比妥抵抗的新生儿癫痫发作，KCC2功能低下在难治性发作出现中的作用 癫痫发作和新型KCC2功能增强剂对新生儿缺血小鼠模型的疗效 癫痫发作。难治性新生儿癫痫与儿童期癫痫综合征和认知功能高度相关 残疾人士。更有效的治疗方法的开发将受益于对 动物模型中潜在的顽固性和癫痫发生的病理生理学和机制。这个 KCC2氯共转运体是中枢神经系统神经元中主要的氯离子输出器。严重损害 神经元排出氯离子的能力逆转了由GABA介导的跨膜氯离子梯度 去极化而不是超极化。新生儿大脑中的刺激性侮辱通常与 严重的癫痫负担，通常难以通过GABA激动剂进行一线治疗干预 比如苯巴比妥。我们以前的工作表明，缺血显著下调了氯-共转运体的表达 KCC2的表达，但NKCC1的表达，作为氯-进口仍然没有受到影响的趋势 脑缺血后的上调。抢救KCC2术后病理生理功能减退 在新生儿的大脑中，缺血侮辱是一种未经测试的策略。假说：抢救KCC2功能减退 新生儿缺血可恢复生理水平的突触抑制和神经元网络活动。这 抢救将防止难治性癫痫的出现，并成功减轻GABA的癫痫负担 激动剂，从长远来看，将是疾病的改良剂。目标：1.早期和急性的动态布局 缺血后KCC2降解及内源性KCC2功能低下的调节 缺血性癫痫。2.记录与KCC2降解相关的皮层神经元去极化 体外缺血及KCC2激动剂对这种去极化的影响3.抢救难治性缺血 一种新型KCC2激动剂的体内癫痫发作和对长期并存疾病的量化影响。交付成果： 在这个项目成功完成后，我们将更接近于了解动态 新生儿癫痫发作至难治性发作期间KCC2表达的变化影响和 创新：了解未成熟的大脑通过重复转化的机制 新生儿期的癫痫发作将有助于引导循证策略用于顽固性疾病的治疗 癫痫发作通常与儿童严重的长期并存有关。

Project summary/abstract: The long-term goal of this research project is to gain an understanding of the broader relationship between phenobarbital-resistant seizures in neonates, the role of KCC2 hypofunction in the emergence of refractory seizures, and the efficacy of novel KCC2 functional enhancers in a mouse model of neonatal ischemic seizures. Refractory neonatal seizures are highly correlated with childhood seizure syndromes and cognitive disabilities. The development of more effective therapies will benefit from a deeper understanding of the pathophysiology and mechanisms of underlying refractoriness and epileptogenesis in animal models. The KCC2 chloride co-transporter is the chief Cl- extruder in central nervous system neurons. Severe impairment in a neurons ability to extrude Cl- reverses the transmembrane Cl- gradient resulting in GABA mediated depolarization instead of hyperpolarization. Excitotoxic insults in neonatal brains are often associated with severe seizure burdens that are commonly refractory to first-line therapeutic interventions with GABA agonists like phenobarbital. Our previous work has shown that ischemia significantly downregulates Cl- co-transporter KCC2 expression but NKCC1 expression which is the Cl- importer remains unaffected with trends of upregulation in post-ischemic brains. Rescuing the pathophysiological hypofunction of KCC2 following ischemic insults is an untested strategy in neonatal brains. Hypothesis: Rescuing KCC2 hypofunction in neonatal ischemia will restore the physiological levels of synaptic inhibition and neuronal network activity. This rescue will prevent the emergence of refractory seizures and successfully reduce seizure burdens with GABA agonists which in turn will be disease modifying in the long-term. Aims: 1.Plot the dynamics of early and acute KCC2 degradation following ischemia and investigate the regulation of intrinsic KCC2 hypofunction during ischemic seizures. 2. Document the KCC2 degradation related depolarization of cortical neurons following ischemia and the effects of a KCC2 agonist on such depolarization in-vitro 3. Rescue refractory ischemic - seizures in-vivo with a novel KCC2 agonist and quantitate effect on long-term co-morbidities. Deliverables: Upon successful completion of this project, we will move closer to understanding the link between the dynamic changes of KCC2 expression during neonatal seizures to the emergence of refractoriness. Impact and Innovation: Understanding the mechanisms by which the immature brain is transformed with repeated seizures in the neonatal period will help guide evidence-based strategies into treatments for intractable seizures that are often associated with severe long-term co-morbidities in children.