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

摘要_项目2 许多工作表明，在睡眠期间，淋巴流动在清除大脑垃圾方面发挥着关键作用。然而， 驱动淋巴流动的机制以及大脑活动状态对其调节的机制还知之甚少。喜欢 U19应用程序中的其他项目，项目2将专注于了解是什么驱动动脉周围脑脊液流入。 我们的动脉周围脑脊液泵送模型建立在已有文献记载的功能性充血机制基础上， 即对神经活动作出反应的一过性血流量和血容量的增加。我们假设脑脊液平行 沿动脉周围间隙泵入，以补偿局部或整体血管容量的变化。因此， NREM睡眠期间的淋巴液体转运高峰是因为协调的神经活动驱动了更大的变化 在血容量上比清醒状态下更大。目标1将具有高时空分辨率的图像的效果 晶须刺激对神经血管单位单个细胞群中钙/cAMP信号的影响 穿透小动脉的直径和脑脊液在动脉周围间隙的速度(以颗粒测量) 跟踪)。我们将通过对神经元的光遗传刺激来确定动脉周围脑脊液泵送的驱动因素， 通过双光子或宏观成像同时观察小鼠的星形胶质细胞或平滑肌细胞。 目标2将检验这样的假设，即自然睡眠状态不仅决定神经回路的活动模式， 还包括血管周围空间的物理尺寸和功能特性，从而 动脉周围脑脊液抽吸的疗效。在业务上，我们将使用目标1中提出的相同模式，包括 觉醒和睡眠(NREM)小鼠的感觉胡须刺激和细胞特异性光遗传激活。目标 3将确定动脉周围脑脊液泵是否可以预测淋巴清除。我们将利用一个 基于皮质内注射小分子荧光示踪剂(DB53，BBB不透性)的新清除法 从大脑清除后积聚在血液中，所以血液中的信号反映了DB53脑流出总量， 与清关路线无关。我们将系统地操作动脉周围脑脊液泵送并确定 对皮质内给药后DB53清除的影响。 总体而言，项目2将通过测试神经回路活动的假设来为U19提案做出贡献 通过控制神经血管的结构和物理性质来确定淋巴溶质的清除 从而使脑脊液泵入脑周围的疗效在脑状态依赖机制中发挥作用。项目2 主要依赖于项目1中的流体动力学建模，而项目3将提供关于 在睡眠过程中，哪些神经元群体控制着神经血管耦合。项目2和项目3将共同充当 为项目4中的神经影像研究提供重要的细胞信息来源，桥接细胞 在人脑中观察到的多尺度动力学下溶质清除的机制和后果。

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.