Imaging of Hippocampal Activity Across Sleep/Wake and Disease States

描述(申请人提供)：脑电功率在theta频率范围(6-9赫兹)，特别是在清醒和快速眼动睡眠期间，被认为主要是由于海马体锥体细胞的同步放电。到目前为止，支持这一假说的研究依赖于细胞外单元记录，其中相对较少的细胞被监测，单个细胞的活动只能被跟踪很短的时间。在这里，我们将通过监测数百个可单独识别的细胞的活动在几周到几个月的时间里，提供前所未有的海马区神经活动在唤醒状态下的网络水平视图。我们将把EEG和EMG活动的遥测记录与一项令人兴奋的新技术结合起来，当与基因编码的荧光钙指示器(例如GCaMP5和GCaMP6)结合使用时，能够从未麻醉、自由移动的动物的局部大脑区域同时成像数百个神经元的活动。Inscope ix nVista HD成像系统由可安装在小鼠头骨上的微型(2 G)荧光显微镜、可在~0.5 mm2的视场内进行高速成像(20-100赫兹)的软件以及可在皮质下大脑结构中以微米级分辨率成像的显微内窥镜组成。由于在海马体及其层状组织中进行的行为研究有丰富的历史，该区域是利用这项技术对唤醒状态下的神经活动进行新见解的理想神经结构。首先，我们将确定CA1内局部网络在觉醒状态和在稳态睡眠压力预期不同的情况下的活动，例如在“亮灯”时期的早期与黑暗时期的早期，以及对睡眠剥夺和随后的恢复睡眠的反应。结合脑电同时测量数百个细胞的活动将使我们不仅能够跟踪单个细胞在睡眠和清醒期间的活动，而且还可以评估局部网络中关于EEG频率和特定EEG事件的同步性。在确定了野生型小鼠的海马区网络参数后，我们将确定亨廷顿病(HD)小鼠模型中CA1区局部网络的活动。我们最近描述了HD的R6/2小鼠模型在睡眠和觉醒以及脑电方面的变化，发现随着疾病的进展，theta范围内的功率极大增加，theta峰值频率(TPF)减慢。使用theta power和tpf作为生物标记物，我们将利用Inscope ix技术的力量，对已识别的神经元进行长期记录，以确定随着R6/2小鼠疾病的进展，海马体网络是如何重组的。在自由行为的小鼠身上同时记录脑电的同时观察数百个细胞的钙动力学的能力，可能会让我们对海马体在行为状态中的作用以及HD海马体中发生的网络变化有新的见解。

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.