Project 2: Periarterial CSF pumping: Dependence on state of brain activity

摘要_项目2 许多研究表明，胶质淋巴流在睡眠期间的大脑废物清除中起着关键作用。然而 驱动胶质淋巴流动的机制及其通过脑活动状态的调节知之甚少。像 U19申请中的其他项目，项目2将侧重于了解驱动动脉周围CSF流入的因素。 我们将动脉周围CSF泵送模型建立在功能性充血的充分记录机制的基础上， 也就是说，响应神经活动的血流量和血容量的短暂增加。我们假设CSF与 沿动脉周围空间沿着泵送以补偿血管容积的局部或整体变化。因此，在本发明中， 胶质淋巴液运输在NREM睡眠期间达到峰值，因为协调的神经活动驱动更大的变化 比清醒状态下的血容量要多目标1将具有高时空分辨率的图像效果 触须刺激对神经血管单位单个细胞群中Ca 2 +/cAMP信号传导的影响 穿透性小动脉的直径，以及动脉周围空间中CSF的速度（通过颗粒测量） 跟踪）。我们将使用神经元的光遗传学刺激来确定动脉周围CSF泵送的驱动因素， 星形胶质细胞或平滑肌细胞，通过双光子或宏观成像同时观察小鼠。 目标2将检验自然睡眠状态不仅决定神经回路的活动模式， 而且血管周围空间的物理尺寸和功能特性， 动脉周围CSF泵送的有效性。在操作上，我们将使用目标1中提出的相同范例，包括 感觉须刺激和细胞特异性光遗传学激活，在清醒和睡眠（NREM）小鼠。目的 3将确定动脉周围CSF泵送是否预测胶质淋巴清除。我们将利用一个 基于皮质内注射小荧光示踪剂（DB 53，BBB-不可渗透）的新型清除试验 其在从脑清除后在血液中积累，使得血液中的信号反映总的DB 53脑流出， 独立于清除路线。我们将系统地操作动脉周围脑脊液泵送， 其对皮质内递送后DB 53清除的影响。 总的来说，项目2将通过测试神经回路活动 通过控制神经血管的结构和物理性质来决定胶质淋巴溶质的清除率 单位，从而在脑状态依赖性机制中动脉周围CSF泵送的功效。计划2 关键取决于项目1中的流体动力学模型，而项目3将提供以下方面的关键信息 哪些神经元群体在睡眠期间控制神经血管耦合。项目2和项目3将共同作为 这是项目4中神经影像学研究的重要细胞信息来源， 机制和后果的溶质清除与多尺度动力学观察在人脑中。

Abstract_Project 2 Many lines of work show that glymphatic flow plays a key role in brain waste clearance during sleep. Yet, the mechanisms driving glymphatic flow and its regulation by the state of brain activity are poorly understood. Like the other projects in the U19 application, Project 2 will focus on understanding what drives periarterial CSF influx. We are basing the periarterial CSF pumping model on the well-documented mechanism of functional hyperemia, i.e., transient increases in blood flow and volume in response to neural activity. We postulate that CSF in parallel is pumped along the periarterial spaces to compensate for local or global changes in vascular volume. Thus, glymphatic fluid transport peaks during NREM sleep because coordinated neural activity drives larger changes in blood volume than in the awake state. Aim 1 will with high spatio-temporal resolution image the effect of whisker stimulation on Ca2+/cAMP signaling in the individual cell populations of the neurovascular unit on the diameters of penetrating arterioles, and on the velocity of CSF in periarterial spaces (as measured by particle tracking). We will determine the drivers of periarterial CSF pumping using optogenetic stimulation of neurons, astrocytes, or smooth muscle cells with simultaneous observations via 2-photon or macroscopic imaging in mice. Aim 2 will test the hypothesis that natural sleep states determine not only the activity pattern of neural circuits, but also the physical dimensions and the functional properties of the perivascular spaces, and thereby the efficacy of periarterial CSF pumping. Operationally, we will use the same paradigms proposed in Aim 1, including sensory whisker stimulations and cell-specific optogenetic activation, in awake and sleeping (NREM) mice. Aim 3 will establish whether periarterial CSF pumping predicts glymphatic clearance. We will take advantage of a novel clearance assay based on intracortical injection of a small fluorescent tracer (DB53, BBB-impermeable) that accumulates in blood after clearance from brain, so that the signal in blood reflects total DB53 brain efflux, independent of the clearance routes. We will systematically manipulate periarterial CSF pumping and determine its effect on DB53 clearance after intracortical delivery. Overall, Project 2 will contribute to the U19 proposal by testing the hypothesis that neural circuit activity determines glymphatic solute clearance by controlling the structural and physical properties of the neurovascular unit and thereby the efficacy of periarterial CSF pumping in a brain state dependent mechanism. Project 2 depends critically on the fluid dynamic modelling in Project 1, while Project 3 will provide key information on which neuronal populations control neurovascular coupling during sleep. Projects 2 and 3 will together serve as an important source of cellular information for the neuroimaging studies in Project 4, bridging cellular mechanisms and consequences for solute clearance with the multiscale dynamics observed in the human brain.