Dravet Syndrome Anti-Epileptic Control by Targeting GIRK Channels

通过针对 GIRK 通道进行 Dravet 综合征抗癫痫控制

• 批准号: 10638439
• 负责人: Diomedes E. Logothetis
• 金额: $ 59.6万
• 依托单位: NORTHEASTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-01 至 2027-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10638439
• 关键词: 2 year old; Address; Affect; Age Months; Agonist; Animal Model; Animals; Antiepileptic Agents; Ataxia; Brain; CNR1 gene; Cannabidiol; Cardiac; Cause of Death; Cells; Child; Classification; Compensation; Computer Models; Coupled; Defect; Developmental Delay Disorders; Disease; Electroencephalography; Ensure; Epidiolex; Epilepsy; Excision; Experimental Designs; FDA approved; Febrile Convulsions; Fenfluramine; Frequencies; Future; G-Protein-Coupled Receptors; GIRK1 subunit, G protein-coupled inwardly-rectifying potassium channel; GTP-Binding Proteins; Genes; Genetic; HTR2A gene; Health; Heterozygote; Hippocampus; Hyperthermia; Impairment; Infant; Interneurons; Ion Channel; Ion Channel Protein; Knock-out; Knowledge; Life; Mediating; Membrane; Membrane Potentials; Modeling; Molecular; Monitor; Mus; Muscle hypotonia; Mutation; Nerve; Neurons; Neurotransmitters; Patients; Pattern; Pharmaceutical Preparations; Phenotype; Phosphatidylinositols; Plants; Population; Potassium; Potassium Channel; Predisposition; Prefrontal Cortex; Proteins; Pyramidal Cells; Recurrence; Resort; Rest; Rodent Model; SCN1A protein; Seizures; Signal Transduction; Sleep; Sleep disturbances; Slice; Speech; Sudden Death; Synapses; Temperature; Testing; Work; density; dimer; dosage; dravet syndrome; drug discovery; epileptiform; experimental study; gamma-Aminobutyric Acid; inward rectifier potassium channel; ketogenic diet; loss of function; loss of function mutation; mouse model; nervous system disorder; neuronal circuitry; pharmacologic; prevent; receptor; side effect; small molecule; sudden unexpected death in epilepsy; synergism; two photon microscopy; two-photon; voltage; voltage gated channel; 2 year old; Address; Affect; Age Months; Agonist; Animal Model; Animals; Antiepileptic Agents; Ataxia; Brain; CNR1 gene; Cannabidiol; Cardiac; Cause of Death; Cells; Child; Classification; Compensation; Computer Models; Coupled; Defect; Developmental Delay Disorders; Disease; Electroencephalography; Ensure; Epidiolex; Epilepsy; Excision; Experimental Designs; FDA approved; Febrile Convulsions; Fenfluramine; Frequencies; Future; G-Protein-Coupled Receptors; GIRK1 subunit, G protein-coupled inwardly-rectifying potassium channel; GTP-Binding Proteins; Genes; Genetic; HTR2A gene; Health; Heterozygote; Hippocampus; Hyperthermia; Impairment; Infant; Interneurons; Ion Channel; Ion Channel Protein; Knock-out; Knowledge; Life; Mediating; Membrane; Membrane Potentials; Modeling; Molecular; Monitor; Mus; Muscle hypotonia; Mutation; Nerve; Neurons; Neurotransmitters; Patients; Pattern; Pharmaceutical Preparations; Phenotype; Phosphatidylinositols; Plants; Population; Potassium; Potassium Channel; Predisposition; Prefrontal Cortex; Proteins; Pyramidal Cells; Recurrence; Resort; Rest; Rodent Model; SCN1A protein; Seizures; Signal Transduction; Sleep; Sleep disturbances; Slice; Speech; Sudden Death; Synapses; Temperature; Testing; Work; density; dimer; dosage; dravet syndrome; drug discovery; epileptiform; experimental study; gamma-Aminobutyric Acid; inward rectifier potassium channel; ketogenic diet; loss of function; loss of function mutation; mouse model; nervous system disorder; neuronal circuitry; pharmacologic; prevent; receptor; side effect; small molecule; sudden unexpected death in epilepsy; synergism; two photon microscopy; two-photon; voltage; voltage gated channel

PROJECT SUMMARY Dravet Syndrome (DS) is a devastating form of epilepsy caused by loss of function of NaV1.1 (80-90% of cases), the predominant voltage-gated Na+ channel expressed in inhibitory (GABAergic) interneurons in the hippocampus and prefrontal cortex. This causes a decrease in the release of inhibitory neurotransmitter (GABA), resulting in hyperexcitability. The disease manifests itself within the first year of life and is usually triggered by hyperthermia causing frequent and prolonged seizures that result in a host of health problems including developmental delay, speech impairment, ataxia, hypotonia and sleep disturbances. Two small molecule monotherapies have been approved recently by the FDA: Epidiolex (Cannabidiol or CBD) in 2018 and Fintepla (fenfluramine or FA) in 2020 for patients two years of age and older. Even though they reduce the frequency of seizures, these drugs at their effective dosages cause multiple side effects. Their mechanism(s) of action to reduce epileptiform activity remain(s) unknown. G protein-gated inwardly rectifying K+ (GIRK) channels have been strongly implicated in epilepsy. They are activated by the Gβγ dimer of G proteins and by [Na+]i in a synergistic manner. The basis of synergism lies in that they each work allosterically to control predominantly the two channel gates: Gβγ, the membrane gate and Na+, the cytosolic gate. In the case of the GABAergic interneurons, we hypothesize that NaV1.1 and GIRK1/2 are coupled, such that the Na+ entering through NaV1.1 promotes GIRK1/2 activity to hyperpolarize the cell and ensure removal of NaV1.1 inactivation for fast spiking. In DS this mechanism becomes compromised causing cell depolarization and inactivation of voltage-gated channels at large present in GABAergic neurons failing to compensate for the loss of NaV1.1. We use GAT1508, a specific activator of GIRK1/2 to compensate for the compromised Na+ entry. Since GAT1508 opens the cytosolic gate, we ask whether it synergizes with CBD (via CB1R) and FA (via 5-HT1DR) to open more fully the membrane gates. In Aim 1, experiments designed to test the hypothesis are aimed at the cellular level in both heterologous expression and in native GABAergic neurons. In Aims 2 and 3, we utilize a DS mouse model, heterozygous for the Scn1a gene that encodes NaV1.1, and in Aim 2, we test the hypothesis at the brain slice level, where synapses and transmitter release remain intact, and compare the DS model to a wild-type animal model. In Aim 3, we pursue experiments at the whole animal level (DS model), using simultaneous EEG and 2-photon microscopy to monitor the neuronal circuits involved.

项目摘要 Dravet综合征（DS）是由NAV 1.1（80-90％的病例）的功能丧失引起的癫痫的毁灭性形式 在抑制性（GABA能）中间神经元中表达的主要电压门控通道 海马和额叶皮层。这会导致抑制性神经递质（GABA）的释放减少， 导致过度兴奋。该疾病在生命的第一年内表现出来，通常是由 热疗经常引起并长时间癫痫发作，导致许多健康问题 发育延迟，言语障碍，共济失调，低骨和睡眠障碍。 FDA最近已批准了两个小分子单等层：Epidiolex（大麻二酚或 CBD）在2018年和2020年的Fintepla（Fenfluramine或FA），两岁及以上的患者。虽然 它们降低了癫痫发作的频率，这些药物处于有效剂量上会导致多种副作用。他们的 降低癫痫病活性的作用机制仍然未知。 G蛋白门控的内部矫正K+（GIRK）通道已与癫痫密切相关。他们是 由G蛋白的Gβγ二聚体和[Na+] I以协同的方式激活。协同作用的基础在于 它们每个人都作变构的作用以控制两个通道门：Gβγ，膜门和 Na+，胞质门。对于GABA能中间神经元，我们假设NAV1.1和GIRK1/2 耦合，使Na+通过NAV1.1进入的Na+会促进GIRK1/2活性以使细胞超极化和 确保删除NAV1.1灭活以进行快速峰值。在DS中，这种机制被损害 在GABA能神经元中，电压门控通道的细胞沉积和灭活，无法 补偿NAV1.1的损失。我们使用GAT1508（GIRK1/2的特定激活剂）来补偿 NA+进入。由于GAT1508打开了胞质门，我们询问它是否与CBD合成（通过 CB1R）和FA（通过5-HT1DR）更全面地打开膜大门。在AIM 1中，旨在测试的实验 该假设针对异源表达和天然GABA能神经元的细胞水平。 在AIMS 2和3中，我们利用DS鼠标模型，用于编码NAV1.1的SCN1A基因的杂合子，并在 AIM 2，我们在大脑切片水平上测试假设，突触和发射器释放保持完整，并且 将DS模型与野生型动物模型进行比较。在AIM 3中，我们在整个动物一级进行实验 （DS模型），使用简单的脑电图和2光子显微镜监测所涉及的神经元电路。