TRPM2 channels and synaptic dysfunction following ischemic injury in the developing brain.

发育中大脑缺血损伤后的 TRPM2 通道和突触功能障碍。

基本信息

批准号：
9386009
负责人：
Robert M Dietz
金额：
$ 19.33万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-07-15 至 2022-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9386009
关键词：
Acute Adolescent Adult Behavior Biological Preservation Brain Brain Injuries Calcineurin Cell Death Cell model Cerebral Ischemia Child Clinical Research Cognitive deficits Communication Data Development Electrophysiology (science)Exhibits Face Functional disorder Genetic Heart Arrest Hippocampus (Brain)Impairment Infant Injury Innovative Therapy Intervention Ion Channel Ischemia Lead Learning Long-Term Potentiation Measures Memory Memory impairment Mentors Modeling Molecular Mus Neurologic Neuronal Dysfunction Neurons Oxidative Stress Pathologic Peptides Pharmacology Phenotype Physiological Physiology Play Puberty Recovery Reperfusion Injury Research Research Proposals Role Signal Transduction Stimulus Stress Subfamily lentivirinae Survivors Synapses Synaptic plasticity Techniques Testing Therapeutic Time Training Transfection Translating Work behavior test career design developmental plasticity experience experimental study fitness functional disability functional outcomes improved in vivo inhibitor/antagonist innovation juvenile animal knock-down mouse model neuron loss neuroprotection neurotoxicity new therapeutic target novel novel therapeutics preclinical study prepuberty prevent protein expression protein function receptor skills small hairpin RNA synaptic function young adult

项目摘要

Project Summary Global cerebral ischemia caused by cardiac arrest results in many neurological sequelae, including deficits in learning and memory. These deficits are as evident in children as they are in adults. The resulting neurological sequelae from cardiac arrest (CA) in children likely arise from both neuronal death and altered physiology in surviving neurons. TRPM2 channels are non-selective ion channels that are activated by hyperoxidative stress and are compelling targets in preventing neurotoxicity and cellular dysfunction. Our lab has recently designed a novel inhibitor of TRPM2 channels, known as tatM2NX, to better understand the role of TRPM2 in neuronal death and dysfunction. A useful measure to assess neuronal dysfunction is to investigate the level of synaptic function. The ability for neurons to undergo synaptic plasticity (long-term potentiation; LTP) in the hippocampus is recognized as an innate measure of function and is a widely accepted cellular model for learning and memory. This proposal makes use of a novel cardiac arrest model in juvenile mice (p21-25) to investigate the hypothesis that activation of TRPM2 channels contributes to impairment of synaptic function and cognitive deficits following global cerebral ischemia. We have found that inhibiting TRPM2 soon after juvenile CA leads to preservation of synaptic function, despite no significant change in neuronal death. We will further test the hypothesis by inhibiting TRPM2 at delayed time points (7-14 days after PCA). Inhibition of TRPM2 will be done with pharmacology (in vivo tatM2NX) or genetic modulation (TRPM2-/-, lentiviral shRNA TRPM2 knockdown) and function will be assessed by electrophysiology and behavior. Preliminary data suggest that delayed inhibition of TRPM2 reverses synaptic impairments after CA. We will use signal transduction techniques to identify the mechanism for synaptic impairment after ischemia-induced TRPM2 activation. We will focus on the hypothesis that activation of TRPM2 signals calcineurin, leading to decreased synaptic function. Finally, we have found that while ischemia in young animals results in impairment of synaptic function up to 14 days after CA, there is remarkable endogenous recovery to control levels. The final aim of the project will investigate the role of developmental changes in TRPM2 expression in neurons through puberty into adulthood that may account for endogenous recovery of impaired synaptic function. Overall, this project has high translational potential through the opportunity of redefining therapeutic windows after global cerebral ischemia in children. Experiments outlined in this proposal will provide important training for an independent research career. In this proposal, I will learn hippocampal lentivirus shRNA transfection, behavior testing, and signal transduction techniques to assess protein expression and function. The mentor team assembled has experience and expertise to assure completion of this project. Upon completion of this proposal, I will combine these skills with electrophysiology techniques that I am already familiar with and apply for an R01 to further characterize TRPM2 activity in the maturing juvenile mouse after cardiac arrest. The impact of this work will be in establishing a novel therapeutic strategy to improve neurological consequences of ischemia in the young brain.

项目概要心脏骤停引起的全脑缺血会导致许多神经系统后遗症，包括功能缺陷学习和记忆。这些缺陷在儿童中和在成人中一样明显。由此产生的神经学儿童心脏骤停（CA）的后遗症可能是由神经元死亡和生理改变引起的幸存的神经元。 TRPM2 通道是由过度氧化应激激活的非选择性离子通道并且是预防神经毒性和细胞功能障碍的引人注目的目标。我们实验室最近设计了一个 TRPM2 通道的新型抑制剂，称为 tatM2NX，以更好地了解 TRPM2 在神经元中的作用死亡和功能障碍。评估神经元功能障碍的一个有用措施是研究突触水平功能。海马神经元进行突触可塑性（长时程增强；LTP）的能力被认为是功能的先天测量，并且是广泛接受的学习和学习细胞模型记忆。该提案利用幼年小鼠的新型心脏骤停模型 (p21-25) 来研究 TRPM2 通道的激活会导致突触功能和认知功能受损的假设全脑缺血后的缺陷。我们发现，在幼年 CA 后不久抑制 TRPM2 会导致尽管神经元死亡没有显着变化，但突触功能得以保留。我们将进一步测试通过在延迟时间点（PCA 后 7-14 天）抑制 TRPM2 的假设。 TRPM2的抑制将会完成药理学（体内 tatM2NX）或基因调节（TRPM2-/-，慢病毒 shRNA TRPM2 敲低）功能将通过电生理学和行为进行评估。初步数据显示，延迟 TRPM2 的抑制可逆转 CA 后的突触损伤。我们将使用信号转导技术确定缺血诱导的 TRPM2 激活后突触损伤的机制。我们将重点关注假设 TRPM2 的激活向钙调磷酸酶发出信号，导致突触功能下降。最后，我们发现虽然幼年动物的缺血会导致突触功能受损长达 14 天 CA，控制水平有显着的内源性恢复。该项目的最终目标将调查从青春期到成年期神经元中 TRPM2 表达的发育变化的作用可能解释受损突触功能的内源性恢复。总体而言，该项目转化率较高通过重新定义儿童全脑缺血后治疗窗口的机会来发挥潜力。该提案中概述的实验将为独立研究生涯提供重要的培训。在这个建议，我学习海马慢病毒shRNA转染、行为测试、信号转导评估蛋白质表达和功能的技术。组建的导师团队经验丰富，的专业知识，以确保该项目的完成。完成本提案后，我会将这些技能与我已经熟悉并应用 R01 来进一步表征的电生理学技术心脏骤停后成熟幼年小鼠的 TRPM2 活性。这项工作的影响将在建立一种新的治疗策略来改善年轻大脑缺血的神经系统后果。