Head-mounted Photoacoustic Imaging of Deep-brain Neural Activities in Freely Behaving Animals

自由行为动物深脑神经活动的头戴式光声成像

• 批准号: 9924909
• 负责人: Vladislav Verkhusha
• 金额: $ 200.72万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9924909
• 关键词: Animal Model; Animals; Area; Behavioral; Blood; Brain; Brain imaging; Brain region; Calcium; Collaborations; Communities; Detection; Disease; Electrodes; Engineering; Fluorescence Resonance Energy Transfer; Focused Ultrasound; Functional Imaging; Goals; Head; Health; Hemoglobin; Hippocampus (Brain); Image; Imaging Techniques; Imaging technology; Knowledge; Measures; Medicine; Membrane; Microscopy; Midbrain structure; Molecular Probes; Monitor; Mus; Neurons; Neurosciences; Optical Methods; Optics; Pathologic Processes; Penetration; Performance; Phytochrome; Protein Engineering; Publications; Reporting; Research Personnel; Resolution; Scanning; Series; Signal Transduction; Speed; System; Techniques; Technology; Texas; Time; Ultrasonic Transducer; Ultrasonic wave; Universities; Variant; Water; Yang; awake; base; calcium indicator; college; design; experience; experimental study; fluorescence imaging; head mounted display; hemodynamics; imaging modality; imaging system; improved; in vivo; innovation; miniaturize; mouse model; novel; optical imaging; photoacoustic imaging; relating to nervous system; scaffold; sensor; two photon microscopy; voltage

Abstract To capture the normal brain functions, it is critically important to record the neural activities in freely-behaving animals, with high resolution, high speed, and high throughput. So far, our knowledge about neuronal activity of awake animals mainly relies on electrode recording, which, however, is invasive. Optical imaging techniques have been widely used to visualize activity of a large number of neurons in mouse models using fluorescent membrane voltage or calcium indicators. However, limited by the penetration depth (<1 mm), it is technically challenging to record the brain functions at depths beyond the cortex layer, such as in the hippocampus. A new large-scale recording technology with high resolution and deep penetration in freely-behaving animals would be of great utility for the neuroscience community. Photoacoustic microscopy (PAM) is a promising candidate for this task due to the relatively deep penetration of ultrasound waves. However, PAM has not been able to image neural activities of freely-moving animals, because (1) it is challenging to miniaturize the imaging system, (2) there lacks calcium or voltage probes that can report neural activities in deep brain, and (3) photoacoustic detection sensitivity of molecular probes is traditionally low due to the strong background signals from blood. In this proposal, we plan to overcome all of the above technical obstacles and develop head-mounted photoacoustic imaging of deep-brain neural activities in freely-behaving animals. To achieve this goal, we will follow a three-aim strategy. (1) In Aim 1, we will develop a miniaturized head-mounted PAM (HM-PAM) system. Several key innovations will reduce the system footprint to 1 cm3. HM-PAM will achieve a penetration depth of ~3.0 mm with ~10−15 µm resolution, which is deeper than that with pure optical microscopy. (2) In Aim 2, we will develop novel near-infrared photoswitchable genetically-encoded calcium indicators (NIR-PS-GECIs) as PA probes. We will engineer and optimize a new class of NIR-PS-GECIs based on photoacoustic Förster resonance energy transfer (FRET-PA). We have proven that the photoswitching, which enables differential PA imaging, is currently one of the most effective approaches to enhance the PA detection sensitivity. We will thus apply fast photoswitching of the NIR-PS-GEICs to enhance the detection sensitivity of HM-PAM. (3) In Aim 3, the optimized HM-PAM and advanced NIR-PS-GECIs will be thoroughly characterized and validated in dissociated neurons and in vivo. We will perform proof-of-concept experiments of deep-brain neural activity in freely-behaving animals. In summary, our proposal will build on the innovations of the first head-mounted PAM system, the first NIR photoswitching GECIs, and the differential FRET- PA imaging that rejects the strong background blood signals. This enabling technology will provide a powerful toolkit for studying neural activities in health, disease, and behavioral states.

抽象的 为了捕获正常的大脑功能，记录自由行为的神经活动至关重要 具有高分辨率，高速和高通量的动物。到目前为止，我们对神经元活动的了解 清醒的动物主要依赖电极记录，但是，这是侵入性的。光学成像技术具有 我们被广泛用于使用荧光膜在小鼠模型中可视化大量神经元的活性 电压或钙指标。但是，受渗透深度（<1毫米）的限制，它在技术上挑战 记录大脑在皮层层以外的深度（例如海马中）的功能。一个新的大型 录制技术具有高分辨率和对自由动物的深度渗透的技术将是极大的实用性 对于神经科学社区。光声显微镜（PAM）是由于 超声波的相对深度穿透。但是，PAM无法成像神经活动 自由移动的动物，因为（1）将小型成像系统微型化是一项挑战，（2）缺乏钙 或电压问题，可以报告深脑中的神经元活动，以及（3）光声检测灵敏度 由于血液中的背景信号很强，传统上，分子问题在传统上很低。在此提案中，我们计划 要克服上述所有技术障碍，并发展出深脑的头部光声成像 自由行为动物的神经活动。为了实现这一目标，我们将遵循三aim策略。 （1）在AIM 1中， 我们将开发一个小型的头部安装PAM（HM-PAM）系统。几项关键创新将减少 系统足迹至1 cm3。 HM-PAM将通过〜10-15 µm的分辨率实现〜3.0毫米的穿透深度，该深度 与纯光学显微镜相比，要深。 （2）在AIM 2中，我们将开发新颖的近红外照片开关 作为PA探针，遗传编码的钙指标（NIR-PS-GECIS）。我们将设计并优化一个新课程 基于光声förster共振能量转移（FRET-PA）的NIR-PS-GECIS的摄入。我们已经证明了 可以实现差异成像的照片处理是目前最有效的方法之一 增强PA检测灵敏度。因此，我们将应用NIR-PS-GEICS快速拍照来增强 HM-PAM的检测灵敏度。 （3）在AIM 3中，优化的HM-PAM和Advanced NIR-PS-Gecis将是 在解离神经元和体内得到了彻底的特征和验证。我们将执行概念验证 自由举止动物中深脑神经活动的实验。总而言之，我们的建议将以 第一个头戴式PAM系统的创新，第一个NIR Photoswitching Gecis和Disialial Fret- PA成像拒绝强烈的背景血液信号。这种促成技术将提供强大的 用于研究健康，疾病和行为状态的神经元活动的工具包。