Effects of dietary alpha-linolenic acid on SUDEP, seizures, and neural structure and function in mouse models of epilepsy

项目摘要 目前可用的药物无法控制约30%的癫痫患者的癫痫发作。这是一个严重的医疗 这是一个问题，因为患有严重进行性耐药性癫痫的患者有很高的突然发作风险。 癫痫意外死亡（SUDEP）因此，需要新的和有效的战略， 癫痫的治疗迫在眉睫在这些难治性形式的潜在的预防性治疗中， 癫痫，基于饮食的方法有很大的希望，因为它们往往是经济的，并与一些广告， 副作用。我们的长期目标是为开发新的 采取有效干预措施，防治耐药性癫痫。为此，有必要阐明 通过饮食调整减少癫痫发作和SUDEP的分子和细胞机制。的 这个探索性项目的总体目标是确定1）饮食的有益影响， 补充必需β-3多不饱和脂肪酸（PUFA）β-亚麻酸（ALA）对小鼠 癫痫模型，以及2）“治疗性饮食”如何影响大脑的形态和生理功能 与癫痫有关的神经元。我们的中心假设是口服ALA和乳清 通过诱导形态学和生理学变化降低癫痫小鼠模型的表型严重程度 在大脑GABA能神经元中。这一假设部分是基于我们在果蝇中的初步发现， 小鼠：1）用乳清补充饮食显著抑制神经元和行为 果蝇的过度兴奋，以及癫痫小鼠的自发性癫痫发作和SUDEP，和2）ALA在 乳清介导果蝇中这种饮食诱导的癫痫发作抑制。当前项目将使用鼠标 电压门控钠通道突变体，人类癫痫的成熟模型。目标#1是确定 ALA对SUDEP和癫痫发作的影响。建立Dravet综合征（Scn 1aR 1407 X/+）和早期婴儿期模型的小鼠 癫痫性脑病（Scn 8aN 1768 D/+）将在断奶后每天喂食ALA，自发性癫痫发作和 SUDEP将在连续视频监控下进行监测。高血压引起的癫痫发作也会 在存在或不存在ALA喂养的情况下对频率和严重程度进行评分。目标#2是描述饮食- 引起脑神经元的形态和生理变化。乳清蛋白和ALA对乳细胞增殖的影响 新皮层GABA能中间神经元的形态学将通过检查它们的数量、树突状细胞、神经元和神经元来评估。 Scn 1aR 1407 X/+突变体的复杂性、轴突投射和末端密度。食疗的效果 对GABA能神经元功能的影响也将通过来自新皮层的膜片钳记录来确定， 切片这项拟议中的研究意义重大，因为它有望确定饮食的有益影响。 在良好控制条件下对遗传性癫痫小鼠模型的修改，以及 大脑神经元由饮食诱导。这些结果为今后的研究奠定了坚实的基础。 分子和细胞的机制，通过修改饮食防止癫痫。

PROJECT SUMMARY Currently available medications fail to control seizures in ~30% of epilepsy patients. This is a serious medical problem because patients with severe progressive forms of drug-resistant epilepsy have a high risk of sudden unexpected death in epilepsy (SUDEP). Thus, the need for novel and effective strategies for the prevention and treatment of epilepsy is pressing. Among the potential adjunctive therapies for such refractory forms of epilepsy, diet-based approaches hold great promise as they are often economical and associated with few ad- verse side effects. Our long-term goal is to provide the scientific foundation necessary for developing new effective interventions for combating drug-resistant epilepsy. To this end, it will be necessary to elucidate the molecular and cellular mechanisms through which dietary modifications reduce seizures and SUDEP. The overall objectives of this exploratory project are to determine 1) the beneficial effects of dietary supplementation with -linolenic acid (ALA), an essential -3 polyunsaturated fatty acid (PUFA), on mouse models of epilepsy, and 2) how “therapeutic diets” affect the morphology and physiological function of brain neurons that are relevant to epilepsy. Our central hypothesis is that orally administered ALA and milk whey reduce phenotypic severity in mouse models of epilepsy by inducing morphological and physiological changes in brain GABAergic neurons. This hypothesis is based, in part, on our preliminary findings in fruit flies and mice: 1) supplementation of diets with milk whey significantly suppresses neuronal and behavioral hyperexcitability in Drosophila, as well as spontaneous seizures and SUDEP in epileptic mice, and 2) ALA in milk whey mediates this diet-induced suppression of seizures in Drosophila. The current project will use mouse voltage-gated sodium channel mutants, well-established models of human epilepsy. Aim #1 is to determine the effects of ALA on SUDEP and seizures. Mice that model Dravet syndrome (Scn1aR1407X/+) and early infantile epileptic encephalopathy (Scn8aN1768D/+) will be fed ALA daily after weaning, and spontaneous seizures and SUDEP will be monitored under continuous video surveillance. Hyperthermia-induced seizures will also be scored for frequency and severity in the presence or absence of ALA feeding. Aim #2 is to delineate diet- induced morphological and physiological alterations in brain neurons. The effects of milk whey and ALA on the morphology of neocortical GABAergic interneurons will be assessed by examining their numbers, dendritic complexity, axonal projections, and terminal density in Scn1aR1407X/+ mutants. The effects of therapeutic diets on the function of GABAergic neurons will also be determined, by patch clamp recordings from neocortical slices. The proposed research is significant because it is expected to identify the beneficial effects of dietary modifications on mouse models of genetic epilepsy under well-controlled conditions, as well as the changes in brain neurons that are induced by diet. These outcomes represent a solid foundation for future studies of the molecular and cellular mechanisms by which modified diets protect against epilepsy.