Mechanisms of seizure resistance in a mouse genetic model with altered metabolism

代谢改变的小鼠遗传模型的癫痫抵抗机制

• 批准号: 10057397
• 负责人: GARY I YELLEN
• 金额: $ 38.46万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-03-01 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10057397
• 关键词: ATP sensitive potassium channel complex; Acute; Address; Affect; Alleles; Animals; Anticonvulsants; Apoptosis Regulation Gene; Apoptotic; Attention; Bad protein; Brain; Brain region; Carbohydrates; Caregivers; Cells; Cellular Metabolic Process; Cytoplasmic Granules; Diet; Diet therapy; Disease; Effectiveness; Engineering; Enterobacteria phage P1 Cre recombinase; Epilepsy; Event; Exhibits; Gene Mutation; Genes; Genetic; Genetic Models; Glucose; Hippocampus (Brain); Hydrocarbons; Individual; Intractable Epilepsy; Ion Channel; Kainic Acid; Ketone Bodies; Knock-out; Learning; Mediating; Metabolic; Metabolism; Modeling; Modification; Mus; Mutation; Neurons; Pentylenetetrazole; Peptides; Permeability; Pharmacology; Point Mutation; Resistance; Seizures; Series; Signal Transduction; Site; Slice; Status Epilepticus; Testing; base; brain cell; brain metabolism; cell type; cellular targeting; conditional knockout; dentate gyrus; effective therapy; entorhinal cortex; experience; genetic manipulation; in vivo; ketogenic diet; knockout animal; mimetics; mouse genetics; mouse model; prevent; public health relevance; response; restoration

PROJECT SUMMARY/ABSTRACT Drug-resistant epilepsy is seriously debilitating and very common, affecting about one-third of the 1-2% of people who experience epilepsy during their lifetime. One of the most effective treatments for drug-resistant epilepsy is dietary therapy, in the form of a very-low-carbohydrate, ketogenic diet. Despite its effectiveness, this diet is not very widely used because of the stringency of the diet and the high commitment required of clinicians and other caregivers. It would be very valuable to understand the mechanism by which altered metabolism produces resistance to epileptic seizures, to “reverse-engineer” it, and to discover alternative pharmacologic ways of tapping into this potent and apparently unique anti-seizure mechanism. We have identified a mouse model that recapitulates the seizure resistance seen in ketogenic diet, but that involves a mutation in a single gene, Bad. The seizure resistance in this genetic model is due to alteration in brain cell metabolism, with less glucose utilization and better utilization of alternative fuels such as ketone bodies, similar to the metabolic changes on a ketogenic diet. We have also discovered a downstream mechanism that is altered both by Bad alteration and by ketogenic diet: a metabolically-sensitive class of ion channels, the ATP-sensitive potassium channels (KATP channels), become more activated in response to metabolic changes. These channels are critical for seizure resistance of the Bad-altered mice, and we have also found that they are responsible for anti-seizure effects of BAD knockout in a brain slice model of seizure. We have also recently learned that KATP channel activation depends on the expression of the BAD protein in individual neurons, which means that the effects of BAD can be genetically targeted to individual cell types or to specific brain regions. This ability to target the genetic manipulation of the BAD protein – which cannot be done for a global manipulation like diet – creates the opportunity to learn the cellular sites of action where BAD modification is required to produce seizure resistance. We now have a conditional knockout allele of the Bad gene (Bad flox/flox) that can be used in combination with various “driver lines” that express Cre recombinase in specific cells. We will determine whether BAD knockout is effective in slice seizure models or against seizures in mice, when the knockout is restricted to certain targets, for instance, to neurons in specific brain regions like the dentate gyrus that are hypothesized to function as “seizure gates”. We will also test a pharmacological approach to producing the anti-seizure effects of BAD, by asking whether a specific class of BAD-mimetic compounds is capable of reversing or mimicking the effect of BAD knockout on seizure-like events in slices. These studies will advance our mechanistic understanding of metabolic seizure resistance and more generally of endogenous “seizure gates”, and will explore new pharmacologic approaches to drug-resistant epilepsy.

项目摘要/摘要 耐药性癫痫病很严重，非常普遍，影响了约1-2％的三分之一 一生中经历癫痫的人。防毒的最有效治疗方法之一 癫痫是饮食疗法，其形式是非常低碳水化合物的生酮饮食。尽管有效 由于饮食的严格程度和所需的高承诺 临床医生和其他护理人员。了解改变的机制将非常有价值 代谢产生对癫痫发作的抵抗力，以“反向工程”，并发现替代方案 利用这种潜力且显然是独特的抗塞氏菌机制的药理方法。 我们已经确定了一种小鼠模型，该模型概括了生酮饮食中的癫痫发作耐药性，但 涉及一个基因中的突变，不好。该遗传模型中的癫痫发作性是由于改变 脑细胞代谢，葡萄糖的利用率较低和更好地利用替代燃料（例如酮） 身体，类似于生酮饮食的代谢变化。我们还发现了下游 通过不良改变和生酮饮食改变的机制：一种代谢敏感的离子类 通道，ATP敏感的钾通道（KATP通道），响应于 代谢变化。这些渠道对于改变不良的小鼠的癫痫发作至关重要，我们有 还发现，它们在脑部癫痫发作模型中负责不良基因敲除的抗癫痫发作。 我们最近还了解到，KATP通道激活取决于不良蛋白在 单个神经元，这意味着不良的影响通常针对单个细胞类型或 到特定的大脑区域。 这种靶向不良蛋白质遗传操作的能力 - 对于全球无法做到 像饮食这样的操纵 - 创造机会学习细胞的作用部位，在不良修改中 需要产生癫痫发作性。我们现在有一个有条件的淘汰等位基因 （不良的flox/flox）可以与特定表达CRE重组酶的各种“驱动线”结合使用 细胞。我们将确定不良敲除在切片癫痫发作模型中是否有效或反对小鼠癫痫发作， 当敲除仅限于某些目标时，例如，在特定大脑区域中的神经元（如 假设作为“癫痫门”起作用的齿状回。我们还将测试药品 通过询问特定类别的不良模拟，产生不良效果的方法 化合物能够逆转或模仿坏敲除对切片中类似癫痫发作的事件的影响。 这些研究将提高我们对代谢癫痫抗性的机械理解，更普遍地 内源性的“癫痫大门”，并将探索耐药性癫痫的新药物方法。