GABA-BETA MECHANISMS OF ABSENCE SEIZURES

失神发作的 GABA-β 机制

• 批准号: 3417938
• 负责人: DAVID A HOSFORD
• 金额: $ 15.46万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-05-01 至 1996-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3417938
• 关键词: GABA receptor; adenylate cyclase; autoradiography; behavior test; brain subcortex; calcium flux; chemical binding; chemical kinetics; cyclic AMP; electroencephalography; enzyme activity; epilepsy; forskolin; gamma aminobutyrate; high performance liquid chromatography; laboratory mouse; ligands; microinjections; molecular site; neocortex; neural information processing; neurons; neuropsychological tests; radioimmunoassay; radiotracer; receptor binding; second messengers; synaptosomes; thalamus; voltage /patch clamp

Absence seizures afflict a significant percentage of children and up to 20% of all patients with epilepsy. Attempts to understand the molecular mechanisms of absence seizures may be facilitated by the use of lethargic (1h/1h) mutant mice, which harbor a single-locus defect and express absence seizures. GABAB receptor activation appears to be required for absence seizures in this model. The goal of this proposal is threefold. First, we will establish the role of GABAB receptors by examining the pharmacologic specificity with which they regulate absence seizures in 1h/1h mice. Second, we will identify the neural network in which GABAB receptors regulate these seizures. Third, we will use tissues from this neural network to characterize the mechanisms triggered by GABAB receptors in 1h/1h mice. In the short-term, the results forthcoming may establish GABAB antagonists as new antiabsence anticonvulsants in humans. In the long- term, these studies will help to characterize the aberrant genetic regulation of the mechanisms underlying absence seizures. This type of information may ultimately enable the control of absence seizures at the regulatory level. Specific aim 1 is to use EEG recordings to firmly establish the role of specific GABAB receptors and the role of specific neuronal structures in these seizures. The effects on seizure frequency of diverse compounds acting at GABAB and other receptors will be recorded by electrodes implanted in neocortex and subcortical sites. Specific aim 2 is to use radioligand binding techniques within slid- mounted sections and membranes of selected neuronal structures to characterize alterations intrinsic to the GABAB receptor in 1h/1h mice. These studies will begin to characterize mechanisms underlying the role of GABAB receptors in absence seizures. Specific aim 3 is to use microinjection techniques to test hypotheses that neuronal structures which express seizures (aim 1) or have enriched GABAB receptor binding (aim 2) regulate absence seizures in 1h/1h mice. This will be accomplished by measuring seizure frequency after microinjections of GABAB agonists or antagonists into candidate neuronal structures. These experiments will be used to choose appropriate tissues for the next aims. Specific aim 4 is to use biochemical techniques to test hypotheses that: i) GABAB receptor-mediated inhibition of net 45Ca+2 uptake into synaptosomes is greater in 1h/1h than nonepileptic control (+/+) mice; ii) GABAB inhibition of adenylate cyclase activity is greater in 1h/1h than +/+ mice. These experiments will help identify second messenger systems underlying the effect of GABAB receptors in absence seizures. Specific aim 5 is to use current- and voltage-clamp recording techniques in thalamic neurons of brain slices from 1h/1h and control mice to test hypotheses that: i) GABAB receptor-mediated IPSPs and low-threshold calcium spikes (LTCSs) are of greater magnitude in 1h/1h mice; ii) GABAB receptors are required for LTCSs. These experiments will further pinpoint candidate mechanisms that link GABAB receptor activation to absence seizures.

失神发作折磨着相当大比例的儿童， 20%的癫痫患者。 试图理解分子 失神发作的机制可以通过使用昏睡剂来促进 (1h/1h）突变小鼠，其具有单基因座缺陷并表达 失神发作 GABAB受体激活似乎是需要的， 在这个模型中， 这项建议有三个目标。 首先，我们将建立 GABAB受体的作用，通过检查药理学特异性， 它们调节1h/1h小鼠的失神发作。 二是 确定GABAB受体调节这些的神经网络 癫痫发作 第三，我们将使用这个神经网络的组织， 描述GABAB受体在1h/1h小鼠中触发的机制。 在短期内，即将到来的结果可能会建立GABAB 拮抗剂作为新的抗失神抗惊厥药在人类。 在漫长的- 长期而言，这些研究将有助于描述异常的遗传特征， 调节失神发作的潜在机制。 这种类型的 这些信息最终可以使控制失神发作成为可能， 监管层面。 具体目标1是使用EEG记录来牢固地确立 特异性GABAB受体和特异性神经元结构在 这些癫痫 不同化合物对癫痫发作频率的影响 作用于GABAB和其他受体的信号将被电极记录 植入新皮层和皮层下部位。 具体目标2是在载玻片内使用放射性配体结合技术， 所选神经元结构的固定切片和膜， 表征1h/1h小鼠中GABAB受体固有的改变。 这些研究将开始描述这种作用的潜在机制 GABAB受体在失神发作中的作用。 具体目标3是使用显微注射技术来检验假设 表达癫痫发作（aim 1）或具有丰富的 GABAB受体结合（目的2）调节1h/1h小鼠的失神发作。 这将通过测量癫痫发作频率来实现 将GABAB激动剂或拮抗剂微量注射到候选神经元中 结构. 这些实验将用于选择合适的组织 为了下一个目标。 具体目标4是使用生物化学技术来检验以下假设： i）GABAB受体介导的对45Ca +2净摄取的抑制 1h/1h突触体大于非癫痫对照（+/+）小鼠; ii）GABAB对腺苷酸环化酶活性的抑制在1h/1h中更大 +/+小鼠。 这些实验将有助于识别第二信使 GABAB受体在失神发作中的作用的潜在系统。 具体目标5是使用电流钳和电压钳记录技术 在1h/1h和对照小鼠脑片的丘脑神经元中， 假设：i）GABAB受体介导的IPSP和低阈值 钙峰（LTCS）在1h/1h小鼠中具有更大的幅度; ii）GABAB 受体是LTCS所必需的。 这些实验将进一步 确定GABAB受体激活与 失神发作