The STAT3 Response of Excitatory Neurons to Epileptogenic Brain Injury

兴奋性神经元对癫痫性脑损伤的 STAT3 反应

• 批准号: 10610469
• 负责人: Amy R. Brooks-Kayal
• 金额: $ 65.69万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10610469
• 关键词: Acute; Adverse effects; Animal Model; Astrocytes; Attenuated; Binding Sites; Brain; Brain Diseases; Brain Injuries; Brain-Derived Neurotrophic Factor; Ca(2+)-Calmodulin Dependent Protein Kinase; Cell Nucleus; Cells; Chromatin; Computer Models; Cyclic AMP; DNA; Data; Development; Disease; Disease Progression; Electrophysiology (science); Encephalitis; Enterobacteria phage P1 Cre recombinase; Epilepsy; Epileptogenesis; Frequencies; GABA Receptor; GABA-A Receptor; GRM5 gene; Gene Expression Regulation; Gene Silencing; Genes; Genetic Transcription; Genome; Genomics; Glutamate Receptor; Glutamates; Hippocampus; Human; Immunity; Impaired cognition; Individual; Inflammation; Inflammatory; Injections; Intervention; JAK2 gene; Janus kinase; Kainic Acid; Knock-out; Knowledge; Laboratories; Long-Term Depression; Malignant neoplasm of brain; Mediating; Mediator; Medical; Memory impairment; Metabotropic Glutamate Receptors; Microglia; Modeling; Molecular; Monitor; Morphology; Mus; N-Methyl-D-Aspartate Receptors; Nerve Degeneration; Neuroglia; Neuronal Plasticity; Neurons; Neuropharmacology; Pathogenesis; Pathologic; Pathway interactions; Patients; Pharmacology; Pilocarpine; Population; Predisposition; Property; Prosencephalon; Protein Tyrosine Kinase; RNA; Receptor Down-Regulation; Receptor Signaling; Recurrence; Reporting; Role; STAT protein; STAT3 gene; Seizures; Signal Transduction; Slice; Sorting; Status Epilepticus; Symptoms; Synaptic plasticity; Tamoxifen; Temporal Lobe Epilepsy; Testing; Therapeutic Intervention; Tissues; Transgenic Organisms; Wild Type Mouse; biocytin; brain cell; calmodulin-dependent protein kinase II; cell type; chromatin remodeling; conditioned fear; dimer; excitatory neuron; gene network; gene repression; glial activation; granule cell; inducible Cre; inflammatory marker; inhibitor; inhibitory neuron; kainate; molecular imaging; mouse model; multiple omics; neural; neural circuit; neuroinflammation; neuronal excitability; neurotransmission; new therapeutic target; prevent; promoter; receptor; receptor downregulation; response; response to injury; synaptogenesis; targeted treatment; transcription factor; transcriptome; transcriptome sequencing; transcriptomics

Abstract Temporal lobe epilepsy (TLE) is a progressive disorder mediated by pathological changes in molecular cascades and neural circuit remodeling in the hippocampus resulting in increased susceptibility to spontaneous seizures and cognitive dysfunction. Targeting these cascades could prevent or reverse symptom progression and has the potential to provide viable disease-modifying treatments that could reduce the portion of TLE patients (>30%) not responsive to current medical therapies. The Janus Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) pathway has recently been implicated in the pathogenesis of TLE. This pathway is known to be involved in inflammation and immunity, and to be critical for neuronal functions such as synaptic plasticity and synaptogenesis. Our laboratories previously showed that a STAT3 inhibitor, WP1066, could greatly reduce the number of spontaneous recurrent seizures (SRS) in an animal model of pilocarpine-induced status epilepticus (SE). While this suggests promise for JAK/STAT inhibitors as disease-modifying therapies, the potential adverse effects of systemic or global CNS pathway inhibition limits their use. Development of more targeted therapeutics will require a detailed understanding of JAK/STAT-induced epileptogenic responses in different cell types. To this end, we have developed a new transgenic line where dimer-dependent STAT3 signaling is functionally knocked out (fKO) by tamoxifen-induced Cre expression specifically in forebrain excitatory neurons (eNs) via the Calcium/Calmodulin Dependent Protein Kinase II alpha (CamK2a) promoter. We now report that STAT3 KO in excitatory neurons (eNSTAT3fKO) markedly reduces the progression of epilepsy (SRS frequency) in the intrahippocampal kainate (IHKA) TLE model and protects mice from kainic acid (KA)-induced memory deficits as assessed by Contextual Fear Conditioning. Using data from bulk hippocampal tissue RNA-sequencing, we further discovered a transcriptomic signature for the IHKA model that contains a substantial number of genes, particularly in synaptic plasticity and inflammatory gene networks, that are down-regulated after KA-induced SE in wild-type but not eNSTAT3fKO mice. In this application, we will test the hypothesis that STAT3 signaling in excitatory neurons is a key driver of epilepsy progression via the selective silencing of genes that regulate synaptic plasticity and neuroinflammation. With an integration of open discovery using multiomics and quantitative molecular imaging (Aims 1 and 3), in combination with electrophysiology and neuropharmacology (Aim 2), we will elucidate the genome’s response to injury (24 h and 4 wks after IHKA) within different cell types and determine why STAT3 KO in eNs inhibits disease progression after KA injection by identifying direct and indirect effects of loss of eNSTAT3 expression on both excitatory and inhibitory neurons. We will also determine the relationship between eNSTAT3 signaling and glial activation by examining effects of eNSTAT3KO on the glial transcriptome and inflammatory markers of microglia and astrocytes. Our results will ascertain if cell-type specific modulation of STAT3 signaling or its downstream targets are promising strategies for therapeutic intervention.

抽象的 颞叶癫痫（TLE）是由分子级联反应的病理变化介导的进行性疾病 海马中的神经回路重塑，导致对发起癫痫发作的敏感性增加 和认知功能障碍。针对这些级联反应可以预防或逆转症状的进展，并具有 提供可以减少TLE患者部分的可行疾病改良治疗的潜力（> 30％） 对当前的医疗疗法没有反应。 Janus激酶/信号换能器和转录激活剂 （JAK/STAT）最近在TLE的发病机理中隐含了途径。该途径已知 参与炎症和免疫力，对于神经元功能（例如合成可塑性和）至关重要 突触发生。我们的实验室先前表明，STAT3抑制剂WP1066可以大大减少 在毛果果诱导的癫痫持续状态的动物模型中的赞助复发性癫痫发作（SRS）数量 （SE）。虽然这表明将JAK/Stat抑制剂作为疾病改良疗法的承诺，但潜在的对面 系统性或全球CNS途径抑制的影响限制了它们的使用。开发更具针对性的疗法 将需要详细了解不同细胞类型中的JAK/Stat诱导的癫痫反应。到 这个目的，我们开发了一条新的转基因线，其中二聚体依赖性STAT3信号在功能上是 通过他莫昔芬诱导的CRE表达（FKO）在前脑兴奋性神经元（ENS）中通过 钙/钙调蛋白依赖性蛋白激酶IIα（CAMK2A）启动子。我们现在报告了STAT3 KO 兴奋性神经元（ENSTAT3FKO）显着降低了癫痫的进展（SRS频率） 海马内海藻酸盐（IHKA）TLE模型，并保护小鼠免受海藻酸（KA）诱导的记忆缺陷 通过上下文恐惧条件评估。使用来自散装海马组织RNA测序的数据，我们 进一步发现了IHKA模型的转录组签名，其中包含大量基因， 特别是在突触可塑性和炎症基因网络中，这些基因网络在Ka诱导的SE之后被下调 在野生型，但不是enstat3fko小鼠中。在此应用中，我们将测试以下假设。 兴奋性神经元是通过调节基因的选择性沉默的癫痫进展的关键驱动力 突触可塑性和神经炎症。通过使用多组学和 定量分子成像（目标1和3），结合电生理学和神经药理学 （AIM 2），我们将在不同的细胞类型中阐明基因组对损伤的反应（IHKA后24小时和4周） 并确定为什么ENS中的STAT3 KO通过识别直接和 ENSTAT3表达丧失对兴奋性和抑制性神经元的间接影响。我们还将确定 ENSTAT3信号传导与神经胶质激活之间的关系通过检查ENSTAT3KO对Glial的影响 小胶质细胞和星形胶质细胞的转录组和炎症标记。我们的结果将确定特定于细胞类型的 STAT3信号或其下游目标的调节是治疗干预的承诺策略。