文献光遗传学激活胆碱能神经元在和LDT课件

1、Optogenetic activation of cholinergic neurons in thePPT or LDT induces REM sleep Christa J. Van DortProc Natl Acad Sci U S A.2014 Dec 29. pii: 201423136. Epub ahead of print Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Harvard Medical School, Boston一般认为

2、广义的基底前脑包括以下结构:(1)下丘脑视前区和前区，(2)隔核群，(3)终纹床核，(4)斜角带核群, (5)Meynert基底核在内的无名质，(6)伏核(nucleus accumbens),(7)嗅结节，(8)嗅皮质和(9)杏仁核群。狭义的基底前脑主要指半球前内侧面和基底面的一些靠近脑表面的灰质，它们是(1)腹侧纹状体苍白球(ventral striatopallidal system)，又称腹侧基底节;(2)杏仁核的延伸部(bed nucleus-amygdalacontinuum, extended amygdala); (3)Meynert基底核(basalnucleus of Me

3、ynert):(4)隔核及Broca氏斜角带核。近年来文献所指的基底前脑多指这些核群。而灵长类的无名质(substantia innominata)也包括在这些灰质群中。basal forebrainBackgroundEarly pharmacological and unit recording studies Suggested that ACh was important for REM sleep regulationinjection of cholinergic drugs into the dorsal mesopontine tegmentum reliably induce

4、d a state very similar tonatural REM sleep in catsElectrical stimulationof the laterodorsal tegmentum (LDT) 被盖背外侧in cats increased the percentage of time spent in REM sleep (14), and activation of the pedunculopontine tegmentum (PPT)脚间脑桥被盖 in rats induced wakefulness and REM sleepIf cholinergic PPT

5、and LDT neurons are necessary for REM sleep to occur, as the early studies suggest, then lesioning the PPT or LDT should decrease REM sleep. In cats, lesions of the PPT and LDT do disrupt REM sleep, but lesions in rodents have had little effect on REM sleep or increased REM sleepThe PPT and LDT are

6、made up of heterogeneous populations of cells, including distinct populations of cholinergic, GABAergic, and glutamatergic neurons Rapid eye movement (REM) sleep is a critical component of restful sleep, yet the mechanisms that control REM sleep are incompletely understood. Brainstem cholinergic neu

7、rons have been implicated in REM sleep regulation, but heterogeneous cell types in the area have made it difficult to determine the specific role of each population, leading to a debate about the importance of cholinergic neurons. Therefore, we selectively activated brainstem cholinergic neurons to

8、determine their role in REM sleep regulation. 光遗传学技术的运用包括四个步骤：第一、找寻合适的光敏蛋白。蛋白质可以是具有天然的光敏性，也可以是经过化学修饰而具有光敏性。第二、遗传信息传递。通过转染、病毒转导、转基因动物系的建立等方式将光敏蛋白的遗传信息传递给目标细胞。第三、可控性演示。通过从时间和空间上控制演示光线的特定性，实现对细胞活动的精确演示。第四、读取研究结果。可采用电极通过检测细胞膜内外电压来测量光敏蛋白的荧光效果变化，并可用荧光性生物传感器来检测不同细胞的读出值。3 REM 快速动眼睡眠又称异相睡眠或快波睡眠。此睡眠时相的脑电图特征是呈

9、现去同步化的快波。各种感觉和躯体运动功能进一步减退。此外，还可有间断性的阵发性表现：如出现眼球快速运动、部分肢体抽动、心率变快、血压升高、呼吸加快等表现。此时易导致心绞痛、哮喘、阻塞性肺气肿缺氧的发作。快波睡眠期间，脑内蛋白质合成增加，新的突触联系建立，这有利于幼儿神经系统的成熟、促进学习记忆活动和精力的恢复。在快波睡眠时，将受试者唤醒，80%的人报告说正在做梦，所以做梦也是快波睡眠的一个特征。脑桥的内部结构以斜方体为界，分为腹侧的基底部和背侧的被盖部。Materials and MethodsAdult male mice (n = 23) expressing ChR2 under the

10、 ChAT and their WT littermates were implanted with EEG and EMG electrodes and bilateral fiber optics in the PPT or LDT. Blue light from a laser was used to stimulate the cholinergic PPT or LDT neurons optogenetically during NREM sleep. Stimulations were 60 s, 80 s, or 180 s long, separated by at lea

11、st 1 min, Fig. 2A demonstrates that LDT neuronshad reliable rapid-onset action potentials following 5-ms light pulses. Fig. 2B shows the ability of a cell to follow the light pulses at 5 Hz over 2 s.图2Activation of Cholinergic Neurons in the PPT and LDT Increases the Probability of REM Sleep.EEG and

12、 EMG traces demonstrate that optogenetic activation of cholinergic neurons in the PPT during non-REM (NREM) sleep induced REM sleep (Fig. 3).1 optogenetic activation of the PPT or LDT increased REM sleep (Fig. 5 A and D).2 NREM sleep decreased between ChR2+ mice and ChR2 micefor all PPT stimulations

13、 and the 60-s LDT stimulation (Fig. 5 B and E3 Wakefulness did not change for either PPT or LDT stimulation, except for a small increase in wakefulness for the 60-s PPT stimulation Fig. 5 C and FIncrease in REM Sleep Is Due to More REM Sleep EpisodesThe increase in REM sleep occurred by increasing t

14、he number of REM sleep episodes (percentage of stimulations that induced REM sleep; Fig. 6 A and D) but not the duration of REM sleep episodes(Fig. 6 B and E). The induced REM sleep was electrophysiologicallysimilar to natural REM sleep. Power spectralanalysis of the EEG during induced REM sleep was

15、 not significantly different from the power spectra during natural REM sleepin the same ChR2+ mice (Fig. 6 C and F),Fiber Optics Were Localized to the PPT and LDTFig. 7 summarizesthe results of the histological analyses demonstrating that the tipsof the fiber optics were localized just above the PPT

16、 or LDTbilaterally. Stained sections were compared with the mouse brainatlas (33) to identify their final location (Fig. 7A).Conclusions1 The present findings demonstrate that activation of cholinergicneurons in the PPT or LDT during NREM sleep is sufficient toincrease REM sleep in mice. 2 The induced REM sleep state closely resembles natural