A Delta GABA Receptor as a Target for Essential Tremor Therapy

Delta GABA 受体作为特发性震颤治疗的靶点

• 批准号: 9495274
• 负责人: Charles Adrian Handforth
• 金额: --
• 依托单位: VA GREATER LOS ANGELES HEALTHCARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2018-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9495274
• 关键词: Affect; Age; Alcohols; American; Animal Model; Behavior; Brain; Brain Mapping; Breeding; Calcium; Cells; Cerebellar Nuclei; Cerebellar cortex structure; Cerebellum; Clinical Trials; Data; Development; Disabled Persons; Dose; Essential Tremor; Ethanol; Exons; Formulation; GABA Receptor; General Population; Genes; Goals; Guidelines; Harmaline; Human; Impairment; In Vitro; Inferior; Intention Tremor; Knock-out; Knockout Mice; Laboratory Research; Lead; Measures; Mediating; Modeling; Molecular Target; Motion; Mus; Muscimol; Occupational; Occupations; Olives - dietary; Patients; Persons; Pharmaceutical Preparations; Pharmacotherapy; Prevalence; Receptor Activation; Recommendation; Reporting; Retirement; Role; Sedation procedure; Slice; Specificity; Testing; Tremor; Upper Extremity; Veterans; Wild Type Mouse; Wine; Withdrawal; Work; alcohol effect; base; delta opioid receptor; design; digital; disability; drug development; experimental study; gamma-Aminobutyric Acid; ganaxolone; granule cell; interest; motor impairment; nervous system disorder; neurosteroids; prevent; programs; promoter; public health relevance; receptor; social; therapy development

DESCRIPTION (provided by applicant): Essential tremor affects 0.5-0.9% of the general population and about 4.5% of those above age 65. Medications for tremor are re-purposed drugs found empirically rather than through laboratory research. As such they lack potency and specificity, and are often not well tolerated. In our program we seek to identify molecular targets to which anti-tremor drugs could be designed, offering greater on-target specificity and potency. A clue is that patients often report that as little as 1-2 glasses of wine can suppress tremor. Brain mapping studies in essential tremor indicate that hyper-oscillatory activity within circuits of the cerebellum underlies tremor. When patients are given low doses of alcohol, not only does the tremor diminish, but excessive activity in the cerebellar cortex also diminishes. We postulate that low-dose alcohol works on tremor by activating GABAA receptors that contain alpha6, beta3 and delta subunits, found only in the cerebellar cortex. If we can show that this receptor can mediate tremor suppression, it could represent an important target for anti-tremor therapy. The GABAA receptor is a pentamer made of alpha, beta, and either gamma or delta subunits. Delta GABAA receptors exert tonic inhibition, and are much more sensitive to GABA. Studies with receptors expressed in cells, and with brain slices, have shown that THIP, muscimol and neurosteroids such as ganaxolone selectively activate delta GABAA receptors rather than gamma GABAA receptors. In addition, ethanol in the range of 1-2 glasses of wine has been found to activate those delta GABAA receptors that use beta3 subunits. We hypothesize that activation of delta GABAA receptors by THIP, muscimol, ganaxolone, and low-dose alcohol will suppress tremor in the harmaline model of essential tremor in wild-type but not littermate delta knockout mice. In addition, the tremor-suppressing effect of alcohol in wild-type mice should be reversed by the alcohol antagonist Ro15-4513, providing further evidence that alcohol is suppressing tremor via a GABAA receptor. We use the harmaline tremor model, which has many similarities to tremor of essential tremor. Motion power is digitally analyzed to provide measures of motion in the tremor bandwidth as a ratio of overall motion power. Our pilot data indicate that THIP, muscimol, ganaxolone, and low-dose alcohol all suppress harmaline tremor in wild-type but not in delta knockout mice. The effect of low-dose alcohol was blocked by Ro15-4513. We propose to replicate these findings (Aim 1). The delta subunit is associated with either alpha 4 or with alpha 6 in GABAA receptors. The alpha4- delta receptors mediate sedation and motor impairment when activated, whereas no behavior has previously been found associated with alpha6-delta receptor activation. Our pilot experiments showed that THIP and low-dose alcohol suppress harmaline tremor in wild-type but not in alpha6 knockout mice, indicating the alpha6-delta receptor mediates tremor suppression. We propose to replicate this finding and test ganaxolone as well (Aim 2). GABAA receptors containing alpha6-delta are found only in the cerebellar cortex, where they associate with beta3, beta2 or beta1. The next step is to investigate whether cerebellar beta3 is important for tremor suppression by these delta-receptor acting drugs by breeding mice with beta3 missing in the part of the cerebellum where alpha6 is expressed (Aim 3). It is anticipated that beta3 will be critical for low-dose alcohol's anti-tremor action, but itmay not be critical for THIP or ganaxolone. Should appropriate strains of knockout mice become available, we will test whether beta2 or beta1 are important for mediating the anti-tremor action of THIP and ganaxolone. These experiments are expected to lead to the identification of a GABAA receptor subtype, alpha6-beta2/3- delta, found only in the cerebellar cortex that will constitute a molecular target to which anti-tremor therapies can be designed to activate. Such a therapy, through its high selectivity, is anticipated to be potent and well tolerated. As the first drug to be developed specifically for essential tremor from laboratory research, it would be expected to confer greater efficacy than existing drugs, preventing disability and loss of occupation in veterans with essential tremor.

描述（由申请人提供）： 特发性震颤影响普通人群的 0.5-0.9%，约占 65 岁以上人群的 4.5%。治疗震颤的药物是根据经验而不是通过实验室研究发现的药物的重新用途。因此，它们缺乏效力和特异性，并且通常不能很好地耐受。在我们的项目中，我们寻求确定可以设计抗震颤药物的分子靶标，从而提供更大的靶向特异性和效力。一个线索是，患者经常报告说，只要喝 1-2 杯酒就可以抑制震颤。特发性震颤的脑图谱研究表明，小脑回路内的过度振荡活动是震颤的根源。 当患者摄入低剂量的酒精时，不仅震颤会减少，小脑皮质的过度活动也会减少。我们假设低剂量酒精通过激活仅存在于小脑皮质中的含有 α6、β3 和 δ 亚基的 GABAA 受体来缓解震颤。如果我们能够证明这种受体可以介导震颤抑制​​，那么它可能成为抗震颤治疗的重要靶点。 GABAA 受体是由 α、β 以及 γ 或 δ 亚基组成的五聚体。 Delta GABAA 受体发挥强直抑制作用，并且对 GABA 更加敏感。对细胞中表达的受体以及脑切片的研究表明，THIP、蝇蕈醇和神经类固醇（例如加奈索酮）选择性激活 δ GABAA 受体，而不是 γ GABAA 受体。此外，还发现 1-2 杯葡萄酒中的乙醇可以激活那些使用 beta3 亚基的 delta GABAA 受体。我们假设 THIP、蝇蕈醇、加奈索酮和低剂量酒精激活 δ GABAA 受体将抑制野生型而非同窝 δ 基因敲除小鼠的骆驼蓬碱原发性震颤模型中的震颤。此外，酒精对野生型小鼠的震颤抑制作用应该可以被酒精拮抗剂Ro15-4513逆转，这进一步证明酒精通过GABAA受体抑制震颤。我们使用骆驼蓬碱震颤模型，它与特发性震颤的震颤有很多相似之处。对运动功率进行数字分析，以提供颤抖带宽中的运动测量值，作为总体运动功率的比率。我们的试验数据表明，THIP、蝇蕈醇、加奈索酮和低剂量酒精均能抑制野生型骆驼蓬碱震颤，但不能抑制 Delta 基因敲除小鼠。 Ro15-4513 可以阻断低剂量酒精的作用。我们建议复制这些发现（目标 1）。 δ 亚基与 alpha 4 或 alpha 相关 GABAA受体中的6个。 α4-δ受体在激活时介导镇静和运动障碍，而之前尚未发现与α6-δ受体激活相关的行为。我们的初步实验表明，THIP 和低剂量酒精可以抑制野生型骆驼蓬碱震颤，但不能抑制 α6 敲除小鼠，这表明 α6-δ 受体介导震颤抑制​​。我们建议复制这一发现并测试加奈索酮（目标 2）。含有 α6-δ 的 GABAA 受体仅存在于小脑皮质中，在那里它们与 β3、β2 或 β1 相关。下一步是通过培育小脑表达 α6 部分缺失 β3 的小鼠，研究小脑 β3 对这些 δ 受体作用药物抑制震颤是否重要（目标 3）。预计 beta3 对于低剂量酒精的抗震颤作用至关重要，但对于 THIP 或加奈索酮可能并不重要。如果有合适的基因敲除小鼠品系可用，我们将测试β2或β1对于介导THIP和加奈索酮的抗震颤作用是否重要。这些实验预计将鉴定出仅在小脑皮质中发现的 GABAA 受体亚型 α6-β2/3-δ，它将构成抗震颤疗法可激活的分子靶点。这种疗法由于其高选择性，预计将是有效的且耐受性良好。作为第一个 该药物是根据实验室研究专门开发用于特发性震颤的药物，预计将比现有药物具有更好的功效，防止患有特发性震颤的退伍军人残疾和失去职业。