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

 描述(由申请人提供)： 这项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)睡眠纺锤体变化背后的细胞机制，Sz是一种在退伍军人中非常普遍的疾病，它消耗了退伍军人管理局精神卫生保健预算的约40%。最近对Sz患者的研究为睡眠纺锤体的数量和固有频率的异常提供了一致的证据，这是在轻度非快速眼动(NREM)睡眠中观察到的一种起伏的脑电(EEG)模式。然而，人类研究在破译潜在细胞机制方面的能力有限。因此，在这里，我们建议使用最先进的光遗传学技术在小鼠身上研究潜在的神经生物学。以往的基础科学工作表明，睡眠纺锤体需要丘脑网状核(TRN)中GABA能神经元的活动，其中大部分含有钙结合蛋白-小蛋白(PV)。在Sz的尸检结果表明，含有PV的皮质GABA能神经元的活动减少。由于TRN PV神经元来自相同的发育途径，一个可信且迄今未被验证的假设是，睡眠纺锤体异常是由于TRN PV神经元活动下调所致。最近在Sz患者中使用催眠药eszopiclone的研究，它针对TRN神经元表达的包含A3亚单位的GABAA受体，改善睡眠和睡眠纺锤体异常，并改善记忆巩固。因此，如果成功，我们在小鼠身上的实验可能会导致基于对TRN PV神经元活动的操纵来纠正Sz纺锤体异常的新疗法，从而改善认知，这是Sz的一种核心损伤。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。我们测试了通道视紫红质2(Channelopoursin2，CHR2)的光遗传刺激(纺锤波和NREM睡眠的功能获得)和质子泵ARCHT(纺锤波和NREM睡眠的功能丧失)抑制对TRN PV神经元和皮质脑电的影响。特定目标(SA)1的实验将测试双侧ChR2刺激是否会诱发睡眠纺锤波并增加NREM睡眠，从而也减少NREM发作期间的听觉感觉传递。单位记录将显示已识别的TRN PV神经元将以与纺锤波相关的爆发形式放电。在SA 2中的实验将测试双侧TRN PV神经元的光学抑制是否会减少纺锤体和NREM睡眠，模拟Sz的纺锤体异常。SA 3中的实验将调查一条重要的、迄今研究较少的TRN输入途径，它发生在基底前脑。我们将测试光刺激基础前脑GABA/PV输入TRN是否会在体内减少睡眠纺锤波，并在体外抑制TRN PV神经元，从而测试这一解剖学证明的途径的功能作用，并预测抑制TRN PV神经元产生纺锤波减少。体外实验将验证我们的假说，即含有A3亚单位的GABAA受体，模拟埃索匹克隆的A3亚单位效应，介导对TRN PV神经元的影响。最后，在SA 4中，我们将测试双边 对TRN PV神经元的光遗传操作将改变新对象识别任务中的记忆回忆，从而模仿Sz中看到的认知缺陷。我们预测ChR2将有所改善，而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.