Genetic Modifiers of Childhood Epilepsy

癫痫是一种常见的神经系统疾病，每26名美国人中就有一人会在一生中受到影响。SCN1A的突变， 编码神经元电压门控钠通道Nav1.1是癫痫最常见的遗传原因。 据报道，在患有不同严重程度的癫痫的个体中，已有超过1600个SCN1A突变，范围从 轻度热性惊厥至德拉韦综合征，一种严重的婴儿发作的癫痫脑病，由 杂合性功能丧失突变。德拉韦综合征的特征是各种类型的癫痫发作， 发育迟缓和死亡风险增加。单基因癫痫的一个共同特征是多变的。 携带相同突变的个体的表达能力，表明临床严重性受遗传因素的影响 修饰符。Scn1a杂合性缺失(Scn1a+/-)小鼠概括了Dravet综合征的核心特征 表型，包括自发性癫痫发作和死亡风险增加。Scn1a的丢失导致减少 海马GABA能中间神经元钠电流导致抑制和兴奋/抑制失败 大脑的不平衡。Scn1a+/-小鼠的表型严重程度强烈依赖于菌株背景。SCN1A+/- 耐药129品系(129，Scn1a+/-)的小鼠没有明显的表型，活着正常。相比之下， [129xB6]F1品系(F1.Scn1a+/-)的SCN1A+/-小鼠表现出自发性癫痫发作和过早死亡， 50%在1个月龄前死亡。在神经元亚型水平上，依赖于应变的差异也很明显。 易感F1Scn1a+/-小鼠GABA能中间神经元钠电流密度降低 与野生型相比，虽然钠电流密度在分离的中间神经元中保持不变 129.SCN1A+/-与野生型窝种有关。这表明，129菌株的中间神经元补偿了 NAV1.1丢失，而F1中间神经元不丢失。基于菌株在表型上的差异 整个动物和细胞水平，我们假设遗传修饰物影响Scn1a+/-表型严重程度 由于Scn1a杂合子背景下不同神经元亚型代偿能力的差异 删除。我们先前定位了几个影响早产儿死亡率的Dravet生存修饰物(DSM)基因座 SCN1A+/-小鼠。在目前的提案中，我们将从三个方面阐述我们的假设。首先，我们会表现得很好 小鼠7号和8号染色体上两个DSM基因座的定位和候选基因分析 单细胞rna-seq分析以表征特定细胞的细胞组成和基因表达的差异 癫痫易感基因F1Scn1a+/-和抗性基因F1Scn1a+/-在表型发病关键期的亚群 129.SCN1A+/-小鼠。第三，我们将通过转录来评估体内候选基因的修饰潜力。 调节对候选基因表达的上调和下调。这些研究的结果将确定修饰语 影响Scn1a+/-小鼠表型严重性的基因和途径。修饰基因的鉴定 Dravet综合征的影响程度将为癫痫的病理生理学提供洞察，并将提示 改善人类患者治疗的新治疗方法。

Epilepsy is a common neurological that will affect 1 in 26 Americans during their lifetime. Mutations in SCN1A, encoding the neuronal voltage-gated sodium channel Nav1.1, are the most common genetic cause of epilepsy. Over 1600 SCN1A mutations have been reported in individuals with epilepsy of varying severity, ranging from mild febrile seizures to Dravet syndrome, a severe infant-onset epileptic encephalopathy caused by heterozygous loss-of-function mutations. Dravet syndrome is characterized by a variety of seizure types, developmental delay and elevated mortality risk. A common feature of monogenic epilepsies is variable expressivity in individuals carrying the same mutation, suggesting that clinical severity is influenced by genetic modifiers. Mice with heterozygous deletion of Scn1a (Scn1a+/-) recapitulate core features of Dravet syndrome phenotypes, including spontaneous seizures and increased mortality risk. Loss of Scn1a results in reduced sodium current in hippocampal GABAergic interneurons, resulting in failure of inhibition and excitatory/inhibitory imbalance in the brain. Phenotype severity in Scn1a+/- mice is strongly dependent on strain background. Scn1a+/- mice on the resistant 129 strain (129.Scn1a+/-) have no overt phenotype and live a normal lifespan. In contrast, Scn1a+/- mice on a [129xB6]F1 strain (F1.Scn1a+/-) exhibit spontaneous seizures and premature lethality, with 50% dying by 1 month of age. Strain-dependent differences are also evident at the level of neuron subtypes. GABAergic interneurons isolated from the susceptible F1.Scn1a+/- mice exhibit decreased sodium current density compared to wildtype littermates, while sodium current density is preserved in interneurons isolated from 129.Scn1a+/- relative to wildtype littermates. This suggests that interneurons from strain 129 compensate for the loss of Nav1.1, while F1 interneurons do not. Based on the strain-dependent difference in phenotypes at the whole animal and cellular levels, we hypothesize that genetic modifiers influence Scn1a+/- phenotype severity due to differences in compensatory capacity among neuronal subtypes in the context of Scn1a heterozygous deletion. We previously mapped several Dravet survival modifier (Dsm) loci that influence premature lethality of Scn1a+/- mice. In the current proposal, we will address our hypothesis in three aims. First, we will perform fine mapping and candidate gene analysis at two Dsm loci on mouse chromosomes 7 and 8. Second, we will perform single cell RNA-seq analysis to characterize differences in cell composition and gene expression in specific cell subpopulations during the critical phase of phenotype onset in epilepsy susceptible F1.Scn1a+/- and resistant 129.Scn1a+/- mice. Third, we will evaluate the modifier potential of candidate genes in vivo using transcriptional modulation to up- and down-regulate candidate gene expression. Results from these studies will identify modifier genes and pathways that influence phenotype severity in Scn1a+/- mice. Identification of modifier genes that influence severity of Dravet syndrome will provide insight into the pathophysiology of epilepsy and will suggest novel therapeutic approaches for improved treatment of human patients.