Probing epileptic circuits with novel Cre- and drug-regulated genetic approaches

用新型 Cre 和药物调节的遗传方法探索癫痫回路

• 批准号: 8913446
• 负责人: EDWARD PEREZ-REYES
• 金额: $ 23.7万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-02-01 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8913446
• 关键词: Affect; Aftercare; American; Animal Model; Animals; Appearance; Back; Behavior; Birth; Cells; Cessation of life; Cholecystokinin; Clinical; Complex; Data; Dendrites; Dependovirus; Development; Disease; Doxycycline; Economics; Electroencephalography; Epilepsy; Epileptogenesis; Food; Frequencies; Functional disorder; Future; Generations; Grant; Hilar; Hippocampus (Brain); Human; Implant; Injection of therapeutic agent; Interneurons; Left; Measures; Medical; Medicine; Methods; Monitor; Motor Seizures; Mus; Neurons; Neurosciences; Output; Parvalbumins; Pathology; Patients; Pharmaceutical Preparations; Potassium Channel; Publishing; Pyramidal Cells; Rattus; Recurrence; Replacement Therapy; Research; Rodent; Sampling; Science; Sclerosis; Seizures; Serotyping; Site; Somatostatin; Status Epilepticus; Synapses; Temporal Lobe Epilepsy; Testing; Time; Transcriptional Activation; Transplantation; Tropism; United States National Institutes of Health; Viral; Withdrawal; base; cell type; critical period; density; dentate gyrus; entorhinal cortex; excitotoxicity; genetic approach; granule cell; human FRAP1 protein; innovation; insight; migration; mimetics; mossy fiber; nervous system disorder; neurogenesis; neuronal circuitry; neuropeptide Y; neurotropic; novel; novel strategies; optogenetics; public health relevance; recombinase; research study; response; small molecule; vesicular GABA transporter; voltage

 DESCRIPTION (provided by applicant): Epilepsy is a major neurological disorder with significant economic and human burdens. One of the most common forms is mesial temporal lobe epilepsy (TLE). Unfortunately, medical treatment of TLE fails in many cases, leaving a large unmet clinical need. Therefore, a thorough understanding of the neuronal circuits involved in seizure initiation and propagation will drive the development of novel therapies. A hallmark of mesial temporal lobe epilepsy for many patients is hippocampal sclerosis. The hippocampus is particularly susceptible to excitotoxicity. Key sites of neuronal death are the hilus of the dentat gyrus, where many inhibitory GABAergic interneurons are lost. Recurrent seizures trigger an increase in both neurogenesis and the development of remaining neurons. This is particularly true for the dentate gyrus, whose responses include: birth and migration of new granule cells, appearance of novel basal dendrites, and an increase in their axonal projections including excitatory collaterals back onto neighboring granule cells (mossy fiber sprouting). Which of all these responses is most responsible for the development of spontaneous seizures? This proposal will test the hypothesis that reduced activity of inhibitory interneurons is a key driver f epileptogenesis. This hypothesis was developed to explain the serendipitous finding that expression of a modified leak K+ channel (TREK-M) in rat dentate hilar neurons triggered limbic seizures similar to those that occur in TLE patients. The research will use a novel chemogenetic approach based on adeno-associated viral delivery of TREK-M whose expression is dependent on the action of Cre recombinase and regulated by doxycycline. This allows us to exploit Cre-driver mice that have been developed as a result of the NIH Neuroscience Blueprint. The ability of chemogenetic silencing of GABAergic neurons will be first tested in mice where Cre is expressed in all GABAergic interneurons. The first aim will validate delivery, record seizure activity, and examine the pathology that results, focusing on mossy fiber sprouting. The second aim will begin to dissect which of the many GABAergic subtypes are critical for seizure generation. This innovative chemogenetic approach will likely have a major impact on the field of neuroscience, as it provides a longer time scale to probe complex behaviors than is possible with optogenetics. The studies may also provide the first animal model of spontaneous recurrent seizures without neuronal death, which would mimic human TLE patients who do not show hippocampal sclerosis.

 描述（由申请人提供）：癫痫是一种严重的神经系统疾病，具有显著的经济和人类负担。最常见的形式之一是内侧颞叶癫痫（TLE）。不幸的是，TLE的医学治疗在许多情况下失败，留下大量未满足的临床需求。因此，彻底了解癫痫发作的启动和传播所涉及的神经元回路将推动新疗法的发展。许多患者的内侧颞叶癫痫的标志是海马硬化。海马体对兴奋性毒性特别敏感。神经元死亡的关键部位是齿状回的门，在那里许多抑制性GABA能中间神经元丢失。复发性癫痫发作触发神经发生和剩余神经元发育的增加。这对于齿状回尤其如此，其反应包括：新颗粒细胞的出生和迁移，新基底树突的出现，以及它们的轴突投射的增加，包括兴奋性侧枝回到邻近的颗粒细胞上（苔藓纤维发芽）。在所有这些反应中，哪一种对自发性癫痫发作的发生最负责任？该提议将检验抑制性中间神经元的活性降低是癫痫发生的关键驱动力的假设。这一假说的发展是为了解释一个偶然的发现，即大鼠齿状门神经元中修饰的泄漏K+通道（TREK-M）的表达触发了类似于TLE患者中发生的边缘系统癫痫发作。该研究将使用一种新的化学遗传学方法，该方法基于TREK-M的腺相关病毒递送，TREK-M的表达依赖于Cre重组酶的作用并受多西环素调节。这使我们能够利用作为NIH神经科学蓝图的结果而开发的Cre-driver小鼠。首先在Cre在所有GABA能中间神经元中表达的小鼠中测试GABA能神经元的化学发生沉默的能力。第一个目标将验证交付，记录癫痫发作活动，并检查病理结果，重点是苔藓纤维发芽。第二个目标将开始剖析许多GABA能亚型中哪些对癫痫发作至关重要。这种创新的化学遗传学方法可能会对神经科学领域产生重大影响，因为它提供了比光遗传学更长的时间尺度来探测复杂的行为。这些研究还可能提供第一个自发复发性癫痫发作而没有神经元死亡的动物模型，这将模拟没有显示海马硬化的人类TLE患者。