Neuronal Seizure Burden versus Cell Death after Neonatal Brain Injury

项目摘要/摘要 每1000名足月新生儿中有2-6人并发缺氧缺血性脑损伤(HI)，并与显著的 死亡率和不良的神经学结果。HI也是新生儿癫痫(NS)的主要原因。当前 治疗既无效，又具有潜在的危害性。这一提议的中心假设是新生儿 癫痫独立加重新生儿缺氧缺血性脑病后的脑损伤。新生儿癫痫发作是否是关键的第二阶段 HI后受伤还是仅仅是损伤严重程度的生物标志物一直是一个长期争论的问题，我们现在有了 可用于以必要的时间和蜂窝分辨率解决问题的技术。具体来说， 对癫痫发作和神经元死亡的活体实时监测克服了过去研究的许多局限性 以便能够更严格地理解这些动态过程。这项提案的近期目标是 是为了验证双光子成像方法在活体内实时监测新生儿癫痫发作和细胞死亡 并确定神经元癫痫负荷与细胞死亡概率之间的关系。 新生儿缺氧缺血性脑病模型。表达神经元荧光蛋白和钙指示剂的转基因小鼠 将接受新生儿HI，随后进行慢性延时双光子成像。这些实验 建议将在癫痫负担和癫痫发作概率之间产生高分辨率的相关数据 神经元死亡。长期目标是获得明确的数据，表明NS是否独立有害 或者不是在HI之后，这对于确定如何积极治疗患者以及如何优先处理研究是至关重要的 努力制定改进的NS治疗策略。在本建议书中获得的技术和数据将是 应用于美国国立卫生研究院资助的未来研究，由申请者检查这些过程并测试治疗方法 临床相关的病理生理状况。 这项提议将创新和严谨的方法与直接的翻译影响结合在一起， 在实验室的科学和职业发展方面以及通过申请者的 科学咨询委员会，以及马萨诸塞州综合医院和 哈佛医学院。与申请人的职业发展相结合地完成拟议的目标 计划将导致申请人在获奖期结束时作为临床医生-科学家的独立性。这个 建议书密切反映申请者的研究和临床兴趣，并提供关键的培训以完成 申请人的长期职业目标是成为新生儿和婴儿临床护理方面的专家 在进行基于实验室的脑机制研究和治疗策略的同时发现癫痫 大脑发育中的损伤、癫痫发作和癫痫的发生。

PROJECT SUMMARY/ABSTRACT Hypoxic-ischemic brain injury (HI) complicates 2-6 per 1,000 term births and is associated with significant mortality and poor neurologic outcomes. HI is also the leading cause of neonatal seizures (NS). Current treatments are both ineffective and potentially harmful. The central hypothesis of this proposal is that neonatal seizures independently worsen brain injury after neonatal HI. Whether neonatal seizures are a critical second hit after HI or just a biomarker of injury severity has been a long-debated question and we now have the technologies available to address it at the temporal and cellular resolution necessary to answer it. Specifically, in vivo real-time monitoring of seizures and neuronal death overcomes many of the limitations of past studies to enable a more rigorous understanding of these dynamic processes. The immediate goals of this proposal are to validate two-photon imaging methods for in vivo real-time seizure and cell death monitoring in a neonatal pup and to define the relationship between neuronal seizure burden and the probability of cell death in a neonatal HI model. Transgenic mouse lines expressing neuronal fluorescent proteins and calcium indicators will undergo neonatal HI and be followed with chronic time-lapse two-photon imaging. The experiments proposed will generate high-resolution correlational data between seizure burden and the probability of neuronal death. The long-term goal is to obtain definitive data indicating whether NS are independently harmful or not after HI, which is essential to determine how aggressively to treat patients and how to prioritize research efforts to develop improved NS treatment strategies. The techniques and data acquired in this proposal will be applied to future NIH-funded studies by the applicant to examine these processes and test treatments in this clinically relevant pathophysiologic condition. This proposal combines innovative and rigorous methodologies with directly translational implications, excellent mentorship in science and career development in the laboratory as well as through the applicant's scientific advisory committee, and extensive institutional resources at Massachusetts General Hospital and Harvard Medical School. Completing the proposed aims in conjunction with the applicant's career development plan will lead to the independence of the applicant as a clinician-scientist by the end of the award period. The proposal closely reflects the applicant's research and clinical interests and provides key training to accomplish the applicant's long-term career goal to become an expert in the clinical care of neonates and infants with epilepsy while conducting laboratory-based research into mechanisms of and therapeutic strategies for brain injury, seizures, and epileptogenesis in the developing brain.