GABA B MECHANISMS OF ABSENCE SEIZURES IN LETHARGIC MICE

GABA B 昏睡小鼠失神发作的机制

• 批准号: 2379674
• 负责人: DAVID A HOSFORD
• 金额: $ 17.14万
• 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 1999-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2379674
• 关键词: GABA receptor; electroencephalography; epilepsy; laboratory mouse; microelectrodes; muscle relaxants; neocortex; neurons; neuropharmacology; pyramidal cells; synaptosomes; thalamus; voltage /patch clamp

Absence seizures afflict a significant percentage of children with epilepsy. Attempts to understand the molecular mechanisms of absence seizures may be facilitated by studying animal models. Our earlier studies using the lethargic (lh/lh) mutant mouse model of absence seizures showed: i) an apparent requirement for GABAB receptors in absence seizures; ii) an increased number of GABAB receptors in neocortex and thalamic nuclei compared to control (+/+) mice; iii) no change in electrophysiologic responses mediated by postsynaptic GABAB receptors in thalamic neurons in lh/lh compared to +/+ mice; and iv) alterations in biochemical functions mediated by presynaptic GABAB receptors in neocortex and thalamus in lh/lh compared to +/+ mice. The short-term goals of this proposal are to seek physiologic correlates to our findings regarding GABAB responses in lh/lh and +/+ mice, and to determine if these responses cause absence seizures. We will perform 6 specific aims to answer the following questions. First (aims 1-3)), what is the physiologic correlate of our evidence that presynaptic GABAB receptor-mediated function is altered in thalamic and neocortical neurons from lh/lh mice? Second (aims 4 and 5), is the altered function mediated by presynaptic GABAB receptors in lh/lh mice caused by the increased numbers of GABAB receptors in these mice? Third (aim 6), are absence seizures in lh/lh mice caused by the increased numbers of GABAB receptors in these mice? In the long-term, the answers forthcoming will increase our understanding of the physiologic and pathophysiologic roles of GABAB receptor-mediated function. More importantly, this information may lead to better therapies for patients with absence seizures. In specific aims 1 and 2 we will use Cs+-filled microelectrodes and whole- cell voltage-clamp techniques in ventrobasal (VB) thalamic neurons (aim 1) or neocortical pyramidal neurons (aim 2) of lh/lh and +/_ slices to measure the effect of presynaptic GABAB receptor activation on IPSCs (autoreceptor function) and EPSCs (heteroreceptor function) evoked by local stimulation. We will activate presynaptic GABAB receptors by: i) applying the GABAB receptor agonist (-) baclofen to the bath; and ii) measuring paired-pulse and homosynaptic depression. In specific aim 3 we will use Cs+ filled microelectrodes and whole-cell voltage clamp techniques in VB thalamic neurons to measure the effect of presynaptic GABAB receptor activation on spindles evoked by stimulation of nucleus reticularis thalami (NRT). In specific aim 4 we will down-regulate GABAB receptors in lh/lh mice by chronic administration of GABAB receptor agonists. We will then measure the effect of presynaptic GABAB receptors on [3H]-GABA release in thalamic and neocortical synaptosomes from these mice and from "normal" lh/lh mice. In specific aim 5 we will use whole-cell voltage-clamp techniques in VB thalamic neurons of down-regulated lh/lh mice to measure the effect of presynaptic GABAB receptors on spindles evoked by stimulation of NRT. In specific aim 6 we will implant bipolar recording electrodes into neocortex of +/+ and lh/lh mice. After up-regulating GABAB receptors by chronic administration of GABAB antagonists to half of the +/+ mice, we will use EEG recordings to compare absence seizure frequency in the groups.

失神发作折磨着很大一部分患有 癫痫 试图理解缺席的分子机制 通过研究动物模型可以促进癫痫发作。 我们早期的研究 使用失神发作的昏睡（LH/LH）突变小鼠模型显示： i）失神发作时对GABAB受体的明显需求; 新皮质和丘脑核团中GABAB受体数量增加 与对照（+/+）小鼠相比; iii）电生理学无变化， 丘脑神经元突触后GABAB受体介导的反应 lh/lh与+/+小鼠相比;和iv）生化功能的改变 在lh/lh时由新皮层和丘脑中的突触前GABAB受体介导 与+/+小鼠相比。 这项建议的短期目标是寻找生理相关性， 我们关于lh/lh和+/+小鼠中GABAB反应的发现， 确定这些反应是否会导致失神发作 我们将执行6 具体目的是回答以下问题。 第一（目标1-3）， 我们的证据表明突触前GABA 受体介导的功能在丘脑和新皮层神经元中改变 从lh/lh小鼠？ 第二（目标4和5），改变的功能介导 突触前GABAB受体在lh/lh小鼠引起的增加 GABAB受体的数量 第三（目标6）是缺席 由GABAB受体数量增加引起的lh/lh小鼠癫痫发作 在这些老鼠？ 从长远来看，即将到来的答案将增加我们的 了解GABAB的生理和病理生理作用 受体介导的功能。 更重要的是，这些信息可能会导致 更好的治疗失神发作的患者。 在具体的目标1和2中，我们将使用Cs+填充的微电极和全- 丘脑腹侧基底核神经元的细胞电压钳技术（aim 1） 或lh/lh和+/_切片的新皮质锥体神经元（aim 2）来测量 突触前GABAB受体激活对IPSC（自身受体）的影响 功能）和由局部刺激诱发的EPSC（异源受体功能）。 我们将通过以下方式激活突触前GABAB受体：i）将GABAB 受体激动剂（-）巴氯芬到浴缸中;和ii）测量配对脉冲 和同突触抑制 在具体目标3中，我们将使用Cs+填充的微电极和全细胞 电压钳技术在VB丘脑神经元，以测量的影响， 突触前GABAB受体激活 丘脑网状核（NRT）。 在具体目标4中，我们将通过以下方法下调lh/lh小鼠中的GABAB受体： 长期给予GABAB受体激动剂。 然后我们将测量 突触前GABAB受体对丘脑[3 H]-GABA释放的影响 以及来自这些小鼠和来自“正常”LH/LH小鼠的新皮质突触体。 在具体目标5中，我们将在VB中使用全细胞电压钳技术， 下调LH/LH小鼠的丘脑神经元，以测量 刺激NRT诱发的突触前GABAB受体。 在具体目标6中，我们将双极记录电极植入到 +/+和lh/lh小鼠的新皮质。 在上调GABAB受体后， 对一半的+/+小鼠长期给予GABAB拮抗剂，我们 将使用EEG记录来比较各组中的失神发作频率。