The dynamic balance between neuronal volume and chloride handling in network excitability after traumatic brain injury

创伤性脑损伤后网络兴奋性中神经元体积和氯处理之间的动态平衡

• 批准号: 10522950
• 负责人: PUNAM MADHUKAR SAWANT-POKAM
• 金额: $ 38.38万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-15 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10522950
• 关键词: Affect; Alcian Blue; Anions; Astrocytes; Biophysics; Blood Vessels; Brain Injuries; Cations; Cause of Death; Cells; Cerebral Edema; Chlorides; Clinical; Complex; Data; Development; Down-Regulation; Drug Targeting; Edema; Elements; Equilibrium; Event; Extracellular Matrix; Failure; Female; GABA Receptor; Gel; Gelatinase B; Genetic; Glutamates; Goals; Histological Techniques; Homeostasis; Hour; Image; Impairment; Injury; Intervention; Label; Link; Matrix Metalloproteinases; Mediating; Mediator of activation protein; Membrane; Membrane Potentials; Monitor; Morbidity - disease rate; Movement; Mus; Neurons; Outcome; Pattern; Population; Potassium Glutamate; Predisposition; Process; Publishing; Receptor Activation; Recovery; Regulation; Resistance; Resolution; Role; Seizures; Sex Differences; Shapes; Stains; Swelling; Testing; Therapeutic Intervention; Time; Traumatic Brain Injury; Up-Regulation; Water; Water Movements; Whole-Cell Recordings; Work; base; cell type; chloride-cotransporter potassium; controlled cortical impact; extracellular; genetic manipulation; in vivo; in vivo two-photon imaging; insight; male; mild traumatic brain injury; mortality; novel; protective effect; response; reuptake; severe injury; success; synaptic inhibition; therapeutic target; tool; transmission process; two photon microscopy; two-photon

Project Summary/Abstract Cerebral edema is the most significant cause of morbidity and mortality following traumatic brain injury (TBI). Though a major target for intervention, current treatments have limited success, perhaps due to the incomplete understanding of edema mechanisms. The task of this proposal is to dissect potential mechanisms underlying TBI-associated neuronal edema, with a particular focus on how edema affects neuronal and network excitability after mild and severe injury. We recently found that neuronal swelling is associated with reduced expression of the cation chloride transporter KCC2 48 hours after controlled cortical impact (CCI) TBI. To our initial surprise, we discovered that neuronal swelling coincided with reduced cellular and network excitability 48 hours post- CCI. And furthermore, when we treated neuronal swelling we observed increased excitability - suggesting that neuronal volume and excitability changes are intimately linked. These results were unexpected, as we predicted that lower levels of KCC2 expression would increase intraneuronal chloride and reduce the efficacy of inhibition – an effect that would increase excitability. However, the excitability of neurons is also affected by their shape: an edematous neuron is less excitable, which may overpower effects of KCC2 on chloride homeostasis. Moreover, despite continued reductions in KCC2 expression, neuronal volume is normalized by one-week post-CCI. This implies that mechanisms beyond KCC2 expression are involved in neuronal volume regulation. This proposal focuses on mechanisms of neuronal edema and its resolution after TBI. In addition, these studies will examine the contributions of neuronal and astrocytic mechanisms on excitability following injury and during recovery. We will test our hypotheses using genetic manipulations, simultaneous in vivo whole cell recording and two- photon microscopy, and histological techniques. In Aim 1, we examine the mechanisms of neuronal edema at 48 hours and at 1 week after CCI and mild TBI, time points when edema is maximal and resolved, respectively. We hypothesize that extracellular impermeant anions are the missing element in the edema response. In Aims 2 & 3, we examine how neuronal volume (and associated chloride shifts) affect excitability at 48 hours and one week after TBI. In Aim 2, we examine to what extent: 1. GABAergic activity is rendered excitatory after TBI, and 2. neuronal volume impacts mechanisms of chloride and water movement across neuronal membranes to influence inhibition. In Aim 3, we assess the effects of neurons (i.e., neuronal volume and chloride transport) and astrocytes (potassium and glutamate clearance) on excitability. In particular, we will examine susceptibility to seizures and spreading depolarizations, which are two key consequences of the hyperexcitable network after TBI. We hypothesize that failure to reuptake potassium and glutamate by astrocytes may contribute - in conjunction with neuronal effects - to enhanced excitability. Our findings will result in a mechanistic understanding of neuronal edema and excitability, including potential drug targets, which in turn could prompt a reassessment of how and why we treat edema clinically.

项目总结/摘要 脑水肿是创伤性脑损伤（TBI）后发病率和死亡率最重要的原因。 虽然是干预的主要目标，但目前的治疗方法成功有限，可能是由于不完整的 了解水肿机制。这项建议的任务是剖析潜在的机制， TBI相关的神经元水肿，特别关注水肿如何影响神经元和网络兴奋性 在轻微和严重的伤害之后。我们最近发现，神经元肿胀与以下表达减少有关： 在控制性皮质撞击（CCI）TBI后48小时，阳离子氯转运蛋白KCC 2。令我们惊讶的是， 我们发现，神经元肿胀与治疗后48小时细胞和网络兴奋性降低一致， CCI。此外，当我们治疗神经元肿胀时，我们观察到兴奋性增加-这表明， 神经元体积和兴奋性变化密切相关。这些结果出乎意料，正如我们预测的那样 低水平的KCC 2表达会增加神经元内氯化物，降低抑制效果， - 这种效应会增加兴奋性。然而，神经元的兴奋性也受到其形状的影响： 水肿的神经元较不容易兴奋，这可能压倒KCC 2对氯稳态的影响。此外，委员会认为， 尽管KCC 2表达持续降低，但CCI后一周神经元体积正常化。这 这意味着KCC 2表达以外的机制参与神经元体积调节。这项建议 着重于TBI后神经元水肿及其消退的机制。此外，这些研究将审查 神经元和星形胶质细胞机制对损伤后和恢复期间兴奋性的贡献。 我们将测试我们的假设，使用遗传操作，同时在体内全细胞记录和两个- 光子显微镜和组织学技术。在目标1中，我们研究了48 ℃时神经元水肿的机制。 在CCI和轻度TBI后24小时和1周，分别是水肿最大和消退的时间点。我们 假设细胞外不渗透阴离子是水肿反应中缺失的元素。在目标2 & 3，我们研究了神经元体积（和相关的氯变化）如何影响48小时和一周的兴奋性 脑外伤后。在目标2中，我们研究到什么程度：1。GABA能活性在TBI后呈现兴奋性，以及2. 神经元体积影响氯和水穿过神经元膜的运动机制， 抑制作用在目标3中，我们评估神经元的影响（即，神经元体积和氯离子转运）， 星形胶质细胞（钾和谷氨酸清除）对兴奋性的影响。特别是，我们将检查易感性， 癫痫发作和扩散去极化，这是TBI后超兴奋网络的两个关键后果。 我们推测星形胶质细胞不能再摄取钾和谷氨酸可能与 对神经元的影响--增强兴奋性。我们的研究结果将导致一个机械的理解， 神经元水肿和兴奋性，包括潜在的药物靶点，这反过来又可能促使重新评估 我们临床上如何以及为什么治疗水肿。