Sleep Spindles: Role of Thalamic Reticular Nucleus and Parvalbumin GABA Neurons

 描述（由申请人提供）： 本研究计划的广泛目标是使用小鼠模型来了解精神分裂症（Sz）睡眠纺锤波改变的细胞机制，精神分裂症是一种在VA中高度流行的疾病，消耗了VA精神保健预算的约40%。最近对Sz患者的研究提供了一致的证据，证明睡眠纺锤波的数量和固有频率异常，睡眠纺锤波是在轻度非快速眼动（NREM）睡眠期间观察到的一种渐强和渐弱的脑电图（EEG）模式。然而，人类研究在破译潜在细胞机制的能力方面受到限制。因此，在这里，我们建议在小鼠中使用最先进的光遗传学技术来研究潜在的神经生物学。 以前的基础科学工作表明，睡眠纺锤波需要丘脑网状核（TRN）中GABA能神经元的活动，其中大多数含有钙结合蛋白，小清蛋白（PV）。Sz的尸检结果表明含有PV的皮质GABA能神经元的活性降低。由于TRN PV神经元来源于相同的发育途径，因此一个似乎合理但迄今为止尚未验证的假设是，睡眠纺锤体异常是由于TRN PV神经元活性下调所致。 最近在Sz患者中使用催眠药右佐匹克隆（靶向TRN神经元表达的含α 3亚基的GABAA受体）的研究改善了睡眠和睡眠梭形异常并改善了记忆巩固。因此，如果成功的话，我们在小鼠中的实验可能会导致新的疗法，以纠正Sz纺锤体异常的基础上操纵TRN PV神经元的活动，从而改善认知，在Sz的核心障碍。 我们的实验有具体的预测，揭示了TRN PV神经元在纺锤体细胞机制中的作用。我们使用视紫红质2（ChR 2）的光遗传学刺激（纺锤波和NREM睡眠的功能获得）和使用质子泵ArchT的抑制（纺锤波和NREM睡眠的功能丧失）来测试对TRN PV神经元和皮质EEG的影响。 特定目标（SA）1中的实验将测试双侧ChR 2刺激是否会引发睡眠纺锤波并增加NREM睡眠，从而减少NREM发作期间的听觉感觉传递。单元记录将显示识别的TRN PV神经元将以与纺锤体相关的突发放电。 SA 2中的实验将测试TRN PV神经元的双侧光学抑制是否会减少纺锤体和NREM睡眠，模拟Sz中的纺锤体异常。 在SA 3的实验将调查一个重要的和迄今为止研究不足的输入途径TRN，在基底前脑中产生。我们将测试，如果光学刺激的基底前脑GABA/PV输入TRN将减少睡眠纺锤体在体内和抑制TRN PV神经元在体外，从而测试功能的作用，这解剖学记录的途径，与预测的抑制TRN PV神经元产生纺锤体减少。体外实验将检验我们的假设，即通过含有α 3-亚基的GABAA受体介导对TRN PV神经元的作用，模拟右佐匹克隆的α 3-亚基作用。 最后，在SA 4中，我们将测试是否双侧 TRN PV神经元的光遗传学操纵将改变新物体识别任务中的记忆回忆，从而模仿在Sz.我们预测Chr 2将改善，Arch T将损害性能。

 DESCRIPTION (provided by applicant): The broad objective of this research program is to use a mouse model to understand the cellular mechanisms underlying alterations in sleep spindles in schizophrenia (Sz), a disease which is highly prevalent in the VA and which consumes ~40 % of the VA mental healthcare budget. Recent studies in Sz patients have provided consistent evidence for abnormalities in the number and intrinsic frequency of sleep spindles, a waxing and waning electroencephalographic (EEG) pattern observed during light non-REM (NREM) sleep. However, human studies are limited in their ability to decipher the underlying cellular mechanisms. Thus, here we propose to use state-of-the-art optogenetic techniques in mice to investigate the underlying neurobiology. Previous basic science work suggested that sleep spindles require the activity of GABAergic neurons in the thalamic reticular nucleus (TRN), most of which contain the calcium binding protein, parvalbumin (PV). Postmortem findings in Sz indicate reductions in the activity of cortical GABAergic neurons containing PV. Since TRN PV neurons are derived from the same developmental pathway, a plausible and so far untested hypothesis is that sleep spindle abnormalities are due to downregulation of the activity of TRN PV neurons. Recent studies in Sz patients using the hypnotic, eszopiclone, which targets the a3-subunit-containing GABAA receptors expressed by TRN neurons, ameliorates sleep & sleep spindle abnormalities and improves memory consolidation. Thus, if successful, our experiments in mice could lead to novel therapies to correct Sz spindle abnormalities based on the manipulation of the activity of TRN PV neurons and thereby improve cognition, a core impairment in Sz. Our experiments have specific predictions that shed light on the role of TRN PV neurons in the cellular mechanisms of spindles. We test the effect on TRN PV neurons and cortical EEG using optogenetic stimulation with Channelrhodopsin2 (ChR2) (gain of function for spindles and NREM sleep) and inhibition using the proton pump ArchT (loss of function for spindles and NREM sleep). Experiments in Specific Aim (SA) 1 will test if bilateral ChR2 stimulation will elici sleep spindles and increase NREM sleep, thereby also reducing auditory sensory transmission during NREM episodes. Unit recordings will show identified TRN PV neurons will fire in bursts associated with spindles. Experiments in SA 2 will test if bilateral optical inhibition of TRN PV neurons will reduce spindles and NREM sleep, modeling spindle abnormalities in Sz. Experiments in SA 3 will investigate an important and so far poorly investigated input pathway to TRN, arising in the basal forebrain. We will test if optical excitation of the basal forebrain GABA/PV input to TRN will reduce sleep spindles in vivo and inhibit TRN PV neurons in vitro, thereby testing the functional role of this anatomically documented pathway, with predicted inhibition of TRN PV neurons producing spindle reduction. In vitro experiments will test our hypothesis of mediation of effects on TRN PV neurons by GABAA receptors containing an a3-subunit, mimicking the a3-subunit effect of eszopiclone. Finally in SA 4 we will test if bilateral optogenetic manipulation of TRN PV neurons will alter memory recall in the Novel Object Recognition Task, thus mimicking the cognitive deficits seen in Sz. We predict Chr2 will improve and Arch T will impair performance.