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Integrated research for the mechanism of sleep/wakefulness regulation

睡眠/觉醒调节机制的综合研究

基本信息

批准号：
20680021
负责人：
YAMANAKA Akihiro
金额：
$ 13.23万
依托单位：
National Institute for Physiological Sciences
依托单位国家：
日本
项目类别：
Grant-in-Aid for Young Scientists (A)
财政年份：
2008
资助国家：
日本
起止时间：
2008 至 2010
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-20680021/
关键词：
睡眠覚醒インビボトランスジェニックマウスチャネルロドプシン2 細胞外記録オレキシン視床下部スライスパッチクランプ脳波筋電図パッチクランプ遺伝子改変マウス緑色蛍光タンパク質光感受性タンパク質ナルコレプシー

项目摘要

Sleep/wakefulness is a physiological phenomenon which observed only in the whole body. Thus, it is essential to study using whole animal. Optogenetics is a recently developed technique to control the activity of neurons by expressing light activated proteins, such as channelrhodopsin2(ChR2) or halorhodopsin(Halo). This technique allows us to control the activity of orexin-producing neurons(orexin neurons) using light in vivo and in vitro. First, we confirmed the function of CHR2 and Halo in the orexin neurons using slice patch clamp and decided a condition of light illumination. Next, we proceeded in vivo experiment using conscious mouse. Optical fibers were bilaterally implanted into the hypothalamus. The activity of orexin neurons was inhibited by illuminating orange light into the hypothalamus through the optical fibers. At the same time, the activity of serotonergic neurons in the raphe nucleus is recorded by the extracellular recordings. Acute inhibition of orexin neurons induced slow wave sleep and the activity of serotonergic neurons was decreased in accordance with initiation of slow wave sleep. This result showed that inhibition of orexin neurons induced slow wave sleep in mice.

睡眠/清醒是一种只在全身观察到的生理现象。因此，利用整个动物进行研究是非常必要的。光遗传学是最近发展起来的一种通过表达光激活蛋白来控制神经元活动的技术，如通道视紫红质2(ChR2)或卤视紫质(Halo)。这项技术允许我们在体内和体外利用光来控制产生食欲素的神经元(食欲素神经元)的活动。首先，我们利用薄片膜片钳证实了ChR2和Halo在增食欲素神经元中的功能，并确定了光照条件。接下来，我们用清醒的小鼠进行了体内实验。双侧将光纤植入下丘脑。通过光纤向下丘脑照射橙色光可抑制增食欲素神经元的活动。同时，通过细胞外记录记录中缝核内5-羟色胺能神经元的活动。急性抑制增食欲素神经元可引起慢波睡眠，5-羟色胺能神经元的活性随慢波睡眠的启动而降低。结果表明，抑制增食欲素神经元可诱导小鼠慢波睡眠。