Unveiling the mechanisms of ultrasound neuromodulation via spatially confined stimulation and temporally resolved recording

项目摘要 早在20世纪20年代，超声波就被探索为调节神经和肌肉的一种方式。一批 最近的研究证明了使用超声波刺激周围神经、脊髓、 还有大脑。然而，很难确定超声波刺激是否是通过直接调节 膜电压或通过间接突触或网络通路。为了揭示超声波的作用机理 调制，我们组成了一个专业互补的团队(薛涵：神经科学与技术；纪欣 程：成像和光声技术；爱德华·博伊登：神经科技)。具体地说，我们将部署 并整合了最近在联合PI的实验室中建立的三项新技术。首先，我们将使用 微型光纤光声转换器(FOC)(直径0.4 mm)它可以定位在大脑内部以传递 具有前所未有的亚毫米空间分辨率的局部化超声。第二，我们将使用尖端技术 基因编码的电压传感器，用于量化超声刺激对脑内单个细胞的影响 大脑的时间分辨率为1毫秒，这超出了常用的钙成像。三是部署 用亚微米空间分辨率受激拉曼散射显微镜测绘阈值电压 和亚阈值水平，以监测单个神经元不同区域的膜对超声波的反应。 将这些新技术与大规模成像平台集成在一起，允许同时进行颅内成像 最近在HAN实验室开发的局部药物传递，我们将对细胞和 超声刺激在原代培养神经元亚细胞水平的生物物理机制 清醒小鼠不同的大脑区域。具体地说，我们将(1)考察个体的空间反应轮廓 以FOC为基础的神经刺激和大范围的体内钙成像；(2)检测 基于FOC的神经刺激和在体清醒脑内单个神经元的时间反应谱 用基因编码的电压传感器进行电压成像；以及(3)检查膜的参与 超声神经调节中的变形和机械敏感通道激活。我们建议的研究将 提供对大脑中超声神经调节的时空分布的系统了解，以及 确定膜变形和机械敏感通道的原因作用。这些新知识将 为超声神经刺激器的合理设计和基础神经科学研究奠定新的基础 以及神经疾病的治疗。

Project Summary Ultrasound has been explored as a modality to modulate nerves and muscles back in the 1920s. A number of recent studies have demonstrated the feasibility of using ultrasound to stimulate peripheral nerves, spinal cord, and brain. Yet, it has been difficult to determine whether ultrasound stimulation is via direct modulation of the membrane voltage or via indirect synaptic or network pathways. In order to unveil the mechanisms of ultrasound modulation, we formed a team of complementary expertise (Xue Han: neuroscience and technology; Ji-Xin Cheng: imaging and opto-acoustic technology; Edward Boyden: neurotechnology). Specifically, we will deploy and integrate three novel technologies that have been established in the co-PI's labs recently. First, we will use a miniature fiber optoacoustic converter (FOC) (0.4 mm in dia.) that can be positioned inside the brain to deliver localized ultrasound with an unprecedented sub-millimeter spatial resolution. Second, we will use cutting-edge genetically encoded voltage sensors to quantify the effects of ultrasound stimulation on individual cells in the brain at a temporal resolution of 1 millisecond that is beyond commonly used Ca2+ imaging. Third, we will deploy submicron spatial resolution stimulated Raman scattering microscopy to map membrane voltage at threshold and sub-threshold level to monitor membrane response to ultrasound at different regions of a single neuron. Integrating these novel technologies with a large-scale imaging platform that allows simultaneous intracranial local drug delivery, recently developed in the Han lab, we will perform a systematic analysis of the cellular and the biophysical mechanisms of ultrasound stimulation at sub-cellular level in cultured primary neurons, and in different brain regions of awake mice. Specifically, we will (1) examine the spatial response profile of individual neurons in awake brains by FOC-based neurostimulation and large-scale Ca2+ imaging in vivo; (2) examine the temporal response profile of individual neurons in awake brains by FOC-based neurostimulation and in vivo voltage imaging with genetically encoded voltage sensors; and (3) examine the involvement of membrane deformation and mechanosensitive channel activation in ultrasound neuromodulation. Our proposed studies will deliver a systematic understanding of the spatiotemporal profiles of ultrasound neuromodulation in the brain, and identify the causal role of membrane deformation and mechanosensitive channels. These new knowledge will build a new foundation for rational design of ultrasound neuro-stimulators and for basic neuroscience research as well as treatment of neurological disorders.