Development of compressed ultrafast microscopy for real-time multi-scale neuroimaging

神经科学的一个普遍目标是实时记录在不同空间和时间尺度上发生的快速、自发的神经活动。传统的电生理学主要依靠微电极记录神经元膜电位。然而，总的来说，这种侵入性方法在记录地点的数量上是有限的，容易受到环境电气噪声的影响，并且在纵向监测方面面临挑战。另一方面，光学记录以其非侵入性、记录并行性和时空可伸缩性的固有优势成为测量神经活动的一种有吸引力的方法。光学电压成像包括两个主要组成部分：电压指示器和光学成像仪器。生物化学的最新进展使快速响应、高灵敏度的荧光电压指示器成为可能。然而，现有的光学仪器仍然缺乏足够的速度、可扩展性和灵敏度。因此，神经活动的实时、多尺度光学成像尚未实现。该发现计划的总体目标是开发用于实时、多尺度光学神经成像的独特成像技术和设备。我们的长期目标是绘制大脑的功能连接体。在接下来的五年里，我们提出了三个项目，从技术发展和神经科学应用两个方面合作研究光电压成像。具体地说，这些项目的目标是(1)开发用于实时、多尺度光电压成像的压缩超快显微镜(CUMIC)(2)在体外使用10 kHz2 MHz的压缩超快显微镜(CUMIC)研究病理条件下轴突起始段和Ranvier结节的生物物理性质(3)在110 kHZ的自由行为动物中使用CUMIC确定神经编码和神经可塑性。最先进的CUMIC系统将极大地帮助神经科学家理解神经生物物理学、电路神经科学和行为输出中的未决问题。CUMIC还将为未来大脑皮层的实时高时空分辨率神经成像铺平道路。此外，在该计划中开发的先进成像技术将得到广泛的应用，包括纳米技术和分子生物学。最后，该项目将培养3名博士、1名硕士和10名暑期学生。获得从光学工程到神经科学应用的宝贵专业知识，这些高素质的人员将贡献他们在光子学、医学物理和生物化学领域的知识，这些领域对加拿大未来在全球知识经济中的成功至关重要。

A prevalent goal in neuroscience is to record fast, spontaneous neural activities occurring at varied spatial and temporal scales in real time. Conventional electrophysiology relied on microelectrodes to record neuron's membrane potentials. However, in general, this invasive approach is limited in the number of recording sites, vulnerable to environmental electrical noises, and challenged for longitudinal monitoring. Optical recording, on the other hand, has emerged as an attractive approach to measuring neural activities with inherent advantages in non-invasiveness, recording parallelism, and spatiotemporal scalability. Optical voltage imaging encompasses two major constituents: voltage indicators and optical imaging instruments. Recent advances in biochemistry have enabled fast-response, high-sensitivity fluorescent voltage indicators. However, existing optical instruments still lack sufficient speed, scalability, and sensitivity. Thus, real-time, multi-scale optical imaging of neural activities has not been achieved.The overall objective of this Discovery program is to develop unique imaging techniques and devices for real-time, multi-scale optical neuroimaging. Our long-term goal is to map the functional connectome of the brain. For the next five years, we propose three projects to investigate optical voltage imaging from the technological development and neuroscience applications in a collaborative effort. Specifically, these projects aim (1) To develop compressed ultrafast microscope (CUMIC) for real-time, multi-scale optical voltage imaging(2) To investigate biophysical properties of the axon initial segment and the node of Ranvier under pathological conditions in vitro using CUMIC at 10 kHz2 MHz (3) To determine neural encoding and neuroplasticity to sensory stimulations in freely behaving animals using CUMIC at 110 kHz The results of the proposed program will represent a unique contribution in biophotonics by significantly enhancing our imaging capability of neurons from sub-cellular to organism levels. The state-of-the-art CUMIC system will greatly assist neuroscientists in understanding open questions in neuronal biophysics, circuit neuroscience, and behavioral outputs. CUMIC will also pave the way for real-time high-spatiotemporal-resolution neuroimaging in the brain cortex in the future. In addition, the advanced imaging technique developed in this program will find a diverse range of applications, including nanotechnology and molecular biology. Finally, this program will train 3 Ph.D., 1 M.Sc., and 10 summer students. Gaining valuable expertise ranging from optical engineering to neuroscience applications, these highly qualified personnel will contribute their knowledge in areas of photonics, medical physics, and biochemistry that are critical for Canada's future success in the global knowledge-based economy.