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 为了捕捉正常的大脑功能，记录自由行为时的神经活动至关重要 迄今为止，我们对动物神经活动的了解具有高分辨率、高速度和高通量。 清醒的动物主要依靠电极记录，然而，光学成像技术具有侵入性。 已被广泛用于使用荧光膜来可视化小鼠模型中大量神经元的活动 然而，受穿透深度（<1 mm）的限制，技术上具有挑战性。 记录皮质层以外深度的大脑功能，例如海马体中的新大规模。 高分辨率和深度渗透的记录技术对于自由行为的动物将有很大的用处 对于神经科学界来说，光声显微镜（PAM）是这项任务的有希望的候选者。 超声波的穿透力相对较深，但是 PAM 还无法对神经活动进行成像。 自由移动的动物，因为（1）使成像系统小型化具有挑战性，（2）缺乏钙 或可以报告大脑深部神经活动的电压探针，以及（3）光声检测灵敏度 由于来自血液的强背景信号，分子探针传统上较低。在本提案中，我们计划。 克服上述所有技术障碍，开发头戴式深脑光声成像 为了实现这一目标，我们将遵循三个目标策略 (1) 在目标 1 中， 我们将开发一种小型化头戴式 PAM (HM-PAM) 系统，几项关键创新将减少 HM-PAM 的系统占地面积可达 1 cm3，穿透深度约为 3.0 mm，分辨率约为 10−15 µm。 比纯光学显微镜更深 (2) 在目标 2 中，我们将开发新型近红外光开关。 作为 PA 探针的基因编码钙指示剂 (NIR-PS-GECI) 我们将设计和优化一个新类别。 基于光声福斯特共振能量转移 (FRET-PA) 的 NIR-PS-GECI 我们已经证明了这一点。 光开关能够实现差分 PA 成像，是目前最有效的方法之一 因此，我们将应用 NIR-PS-GEIC 的快速光开关来增强 PA 检测灵敏度。 (3) 在目标 3 中，优化的 HM-PAM 和先进的 NIR-PS-GECI 将是 我们将在分离的神经元和体内进行彻底的表征和验证。 总之，我们的建议将建立在自由行为动物的深部大脑神经活动的基础上。 第一个头戴式 PAM 系统、第一个近红外光电开关 GECI 和差分 FRET- 的创新 PA 成像可抑制强烈的背景血液信号，这一技术将提供强大的功能。 用于研究健康、疾病和行为状态下的神经活动的工具包。