Imaging of Hippocampal Activity Across Sleep/Wake and Disease States

描述（由申请人提供）：在θ频率范围（6-9 Hz）内的EEG功率，特别是在清醒和REM睡眠期间，被认为主要是由于海马锥体细胞的同步放电。迄今为止，支持这一假设的研究依赖于细胞外单位记录，其中监测相对少量的细胞，并且只能在短时间内跟踪单个细胞的活性。在这里，我们将通过监测数百个可单独识别的细胞在数周到数月内的活动，提供海马体在唤醒状态下神经活动的前所未有的网络水平视图。我们将把脑电图和肌电图活动的联合收割机遥测记录与一种令人兴奋的新技术结合起来，当与遗传编码的荧光钙指标（例如，GCaMP 5和6），能够同时从未麻醉的自由移动动物的局部脑区成像数百个神经元的活动。Insopix nVista HD成像系统由微型（<2 g）荧光显微镜（可安装在小鼠颅骨上）、软件（可在高达~0.5 mm 2的视野内进行高速成像（20-100 Hz））和显微内窥镜（可在皮质下大脑结构中进行微米级分辨率成像）组成。由于在海马体及其层状组织中进行的行为研究的丰富历史，该区域是理想的神经结构，可以利用该技术获得对唤醒状态下神经活动的新见解。首先，我们将确定CA 1内的局部网络在觉醒状态下的活动，以及在稳态睡眠压力可能不同的情况下，例如在“开灯”早期与黑暗期早期以及对睡眠剥夺和随后的恢复睡眠的反应。与EEG一起同时测量数百个细胞的活动将使我们不仅能够在睡眠和清醒时跟踪单个细胞的活动，而且还能够评估局部网络中关于EEG频率和特定EEG事件的同步性。在定义了野生型小鼠海马网络的参数后，我们将确定亨廷顿病（HD）小鼠模型中CA 1内局部网络的活性。我们最近的特点是在睡眠和觉醒的变化，并在脑电图的R6/2小鼠模型的HD和发现都极大地增加了功率的θ范围和θ峰频率（TPF）的疾病进展放缓。使用theta功率和TPF作为生物标志物，我们将利用Insopix技术的力量，对已识别的神经元进行长期记录，以确定R6/2小鼠中海马网络如何随着疾病进展而重组。观察数百个细胞的钙动力学的能力，同时记录脑电图在自由行为的小鼠可能会导致新的见解海马在行为状态中的作用和网络的变化，发生在海马在HD。

DESCRIPTION (provided by applicant): EEG power in the theta frequency range (6-9 Hz), particularly during waking and REM sleep, is believed to primarily be due to synchronous firing of hippocampal pyramidal cells. To date, studies supporting this hypothesis have relied on extracellular unit recordings where a relatively small number of cells are monitored and the activity of individual cells can only be followed for short periods of time. Here, we will provide n unprecedented network level view of neural activity in the hippocampus across arousal states by monitoring the activity of hundreds of individually identifiable cells over weeks to months. We will combine telemetric recording of EEG and EMG activity with an exciting new technology that, when used in conjunction with genetically-encoded fluorescent calcium indicators (e.g., GCaMP5 and 6), enables imaging the activity of hundreds of neurons simultaneously from a local brain region in unanesthetized, freely-moving animals. The Inscopix nVista HD imaging system is comprised of a miniature (<2 g) fluorescence microscope that can be borne on the skull of a mouse, software that enables high-speed imaging (20-100 Hz) over a field of view up to ~0.5 mm2, and microendoscopes that allow imaging with micron-scale resolution in subcortical brain structures. Because of the rich history of behavioral studies conducted in the hippocampus and its laminar organization, this region is the ideal neural structure in which to utilize this technology to obtain novel insights into neural activity across arousal states. First,we will determine the activity of localized networks within CA1 across arousal states and during conditions when homeostatic sleep pressure can be expected to differ, such as early in the "lights on" period vs. early in the dark period and in response to sleep deprivation and subsequent recovery sleep. Concurrent measurement of the activity of hundreds of cells in conjunction with the EEG will enable us to not only follow the activity of individual cells across sleep and wakefulness, but also to evaluate synchrony within the local network with respect to EEG frequencies and specific EEG events. Having defined the parameters of the hippocampal network in wildtype mice, we will then determine the activity of local networks within CA1 in a mouse model of Huntington's disease (HD). We have recently characterized changes in sleep and wakefulness and in the EEG of the R6/2 mouse model of HD and found both tremendously increased power in the theta range and slowing of the theta peak frequency (TPF) as disease progresses. Using theta power and TPF as biomarkers, we will exploit the power of the Inscopix technology to enable long-term recordings of identified neurons to determine how the hippocampal network is reorganized as disease progresses in R6/2 mice. The ability to observe calcium dynamics of hundreds of cells while simultaneously recording EEG in freely-behaving mice may lead to new insights into the role of the hippocampus in behavioral states and the network changes that occur in the hippocampus in HD.