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。一些 最近的研究已经证明了使用超声波刺激周围神经，脊髓， 和大脑。然而，很难确定超声刺激是否是通过直接调制超声信号来实现的。 膜电压或通过间接突触或网络途径。为了揭示超声波的作用机理 调制，我们形成了一个互补的专业知识团队（薛汉：神经科学与技术;纪欣 Cheng：成像和光声技术; Edward Boyden：神经技术）。具体来说，我们将部署 并将合作伙伴实验室最近建立的三项新技术整合在一起。首先，我们将使用 微型光纤光声转换器（FOC）（直径0.4mm）它可以被放置在大脑内部 具有前所未有的亚毫米空间分辨率的局部超声。第二，我们将使用尖端的 基因编码的电压传感器，用于量化超声刺激对细胞中单个细胞的影响 大脑的时间分辨率为1毫秒，这超出了常用的Ca 2+成像。第三，我们将部署 亚微米空间分辨率受激拉曼散射显微镜用于绘制阈值处的膜电压 和亚阈值水平来监测单个神经元的不同区域处的膜对超声的响应。 将这些新技术与大规模成像平台相结合， 局部给药，最近在汉实验室开发，我们将进行细胞和 在培养的原代神经元中，在亚细胞水平上超声刺激的生物物理机制， 不同的大脑区域。具体而言，我们将（1）检查个体的空间反应轮廓 通过基于FOC的神经刺激和体内大规模Ca 2+成像，观察清醒脑中的神经元;（2）检查 基于FOC神经刺激清醒脑中单个神经元的时间响应曲线 用基因编码的电压传感器进行电压成像;（3）检查膜的参与 变形和机械敏感通道激活的超声神经调节。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理解，以及 确定膜变形和机械敏感通道的因果作用。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了新的基础 以及神经系统疾病的治疗。

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.