Loss of Grin2a Promotes Hippocampal Hyperexcitability and Epileptiform Activity

Grin2a 的缺失会促进海马过度兴奋和癫痫样活动

• 批准号: 10357918
• 负责人: Chad R. Camp
• 金额: $ 4.32万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2023-02-11
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10357918
• 关键词: Adolescent; Adult; Antiepileptic Agents; Apical; Automobile Driving; Behavioral; Biological Assay; Brain; CNR1 gene; Calcium Spikes; Caring; Cell surface; Child; Data; Dendrites; Dendritic Spines; Development; Disease; Early Intervention; Epilepsy; Epileptogenesis; Equilibrium; Evaluation; Exhibits; Fever; Financial Hardship; Frequencies; Genes; Glutamate Receptor; Hippocampus (Brain); Human; Interneurons; Ligands; Link; Long-Term Depression; Long-Term Potentiation; Measures; Mediating; Modeling; Morphology; Mus; Mutation; N-Methyl-D-Aspartate Receptors; NMDA receptor A1; Parvalbumins; Patients; Permeability; Pharmacology; Phenotype; Prevalence; Prognosis; Property; Pyramidal Cells; ST5 gene; Seizures; Slice; Stains; Surface; Symptoms; Synapses; Synaptic plasticity; Therapeutic; Therapeutic Intervention; Time; Variant; Wild Type Mouse; brain abnormalities; de novo mutation; density; epileptic encephalopathies; exome sequencing; experimental study; genetic variant; gephyrin; insight; loss of function; loss of function mutation; neural circuit; neuronal cell body; neuronal circuitry; neuronal excitability; neurotransmission; patch clamp; piriform cortex; postsynaptic; prevent; primary caregiver; receptor; receptor expression; receptor function; social

PROJECT SUMMARY/ABSTRACT: De novo mutations in the GRIN2A gene, which encodes the GluN2A subunit of the N-methyl-D-aspartate receptor (NMDAR), are linked to several forms of epileptic encephalopathy (EE). EE is a group of devastating epilepsies, which often involve intractable seizures and different brain abnormalities. The prognosis for EE patients is poor, partly due to inadequate pharmacological options that dampen symptoms, but have little ability to rectify underlying aberrant neuronal circuitry. In addition, children with EE often require around-the-clock care, placing a heavy social and financial burden on primary caregivers, making the need for early intervention with circuitry-correcting therapeutics urgent and necessary. Here, I propose experiments that explore the cellular mechanisms driving epileptiform activity in patients with loss-of-function (LoF) GRIN2A mutations, a group of genetic variants that reduce the function or cell surface expression of GluN2A-containing NMDARs. Whole exome sequencing revealed that multiple patients with EE have de novo mutations in GRIN2A, with a prevalence of roughly 1 in 80,000. Evaluation for functional activity has revealed that 56% of epilepsy-related GRIN2A mutations are LoF variants, displaying diminished receptor function and/or decreased surface expression. These results are intriguing as one might hypothesize that the loss of excitatory synaptic GluN2A subunit would decrease excitability, rather than promote epileptiform activity. To understand how diminished GluN2A function/expression can promote compensatory alterations that increase excitability, I will use Grin2a +/- and -/- mice as a model for loss-of-function GRIN2A variants. Preliminary behavioral data indicate that Grin2a -/- (2AKO) mice have a lower threshold for febrile-induced seizures compared to wildtype (WT) mice. In adult brain slices, 2AKO mice exhibit an increased frequency of epileptiform burst-firing activity in the CA1 region of the hippocampus compared to WT mice. These data indicate that the loss of GluN2A activity is sufficient to drive an epileptic phenotype, as observed in human patients with heterozygous LoF GRIN2A mutations. Taken together, these data suggest that 2AKO mice have an altered excitatory-to-inhibitory balance. Therefore, I hypothesize that epileptiform activity in 2AKO mice is: 1) dependent on a critical developmental window, 2) may be due to changes in postsynaptic receptor expression and hippocampal connectivity, and 3) could be modulated by aberrant forms of synaptic plasticity. Information on how and when the imbalance in excitation/inhibition is generated will help advance a cellular and temporal understanding on the mechanisms involved in epileptogenesis in LoF GRIN2A patients. Thus, these experiments could help guide specific anti-epileptic treatment options for LoF GRIN2A patients suffering from EE. Moreover, if aspects of epileptogenesis are shared across the spectrum of epileptic disorders, data obtained from this proposal may uncover overarching disease mechanisms without being restricted to just LoF GRIN2A patients.

项目摘要/摘要： 编码N-甲基-D-天冬氨酸GluN2A亚单位的GRIN2A基因的从头突变 受体(NMDAR)与几种形式的癫痫脑病(EE)有关。EE是一群毁灭性的 癫痫，通常涉及顽固性癫痫发作和不同的大脑异常。脑血栓形成的预后 病人很穷，部分原因是没有足够的药物选择来抑制症状，但几乎没有能力 来矫正潜在的异常神经回路。此外，患有EE的儿童通常需要全天候护理， 给主要照顾者带来沉重的社会和经济负担，使早期干预的必要性 电路矫正疗法迫在眉睫且必要。在这里，我提议进行一些实验，探索细胞 功能丧失(LoF)GRIN2A突变患者癫痫样活动的驱动机制，一组 降低含有GluN2A的NMDARs的功能或细胞表面表达的基因变体。整体 外显子组测序显示，多名EE患者存在GRIN2A新基因突变， 大约80000人中就有1人。功能活动评估显示，56%的癫痫相关GRIN2a 突变是LOF变异体，表现为受体功能减弱和/或表面表达减少。这些 结果很耐人寻味，因为人们可能会假设，兴奋性突触GluN2A亚单位的丢失将 降低兴奋性，而不是促进癫痫样活动。 了解GluN2A功能/表达减弱如何促进代偿性改变 为了增加兴奋性，我将使用GRIN2A+/-和-/-小鼠作为功能丧失的GRIN2A变体的模型。 初步的行为学数据表明，GRIN2A-/-(2AKO)小鼠有较低的发热诱导阈值 与野生型(WT)小鼠比较。在成年脑片中，2AKO小鼠表现出增加的频率 与WT小鼠相比，海马区CA1区的癫痫样爆发放电活动。这些数据表明 GluN2A活性的丧失足以导致癫痫表型，正如在人类 杂合性LOF GRIN2A突变。综上所述，这些数据表明，2AKO小鼠有一种改变 兴奋与抑制的平衡。因此，我假设2AKO小鼠的癫痫样活动是：1) 取决于一个关键的发育窗口，2)可能是由于突触后受体表达的变化 和海马区的连接，以及3)突触可塑性的异常形式可以调节。信息 关于兴奋/抑制失衡是如何以及何时产生的，将有助于推进细胞和时间 对LoF GRIN2A患者癫痫发病机制的认识因此，这些实验 可以帮助指导患有EE的LoF GRIN2A型患者的特定抗癫痫治疗选择。此外， 如果癫痫发生的各个方面都在癫痫障碍的谱系中共享，那么从这个角度获得的数据 该提案可能会揭示主要的疾病机制，而不仅仅限于LOF GRIN2A患者。