Two-photon Volumetric Endoscopic System for Deep and High-speed 3D Brain Imaging

Jyun-Yi Lin^1,2*, Yu-Feng Chien¹, Po-Ting Yeh³, Shih-Kuo Chen³, Shi-Wei Chu^1,2,4

¹Department of Physics, National Taiwan University, Taipei, Taiwan
²Brain Research Center, National Tsing Hua University, Hsinchu, Taiwan
³Department of Life Science, National Taiwan University, Taipei, Taiwan
⁴Molecular Imaging Center, National Taiwan University, Taipei, Taiwan

* Presenter:Jyun-Yi Lin, email:r07222004@ntu.edu.tw

The study of in vivo neural connections and activities is the essential step to understand how the brain functions, and two-photon microscopy (2PM) has been a powerful tool in neuroscience with its ~ μm spatial resolution, optical sectioning ability, and remarkable penetration depth. However, in typical 2PM, volumetric images are acquired by slow axial scan, and the best imaging depth it could reach is only ~ 1 mm, which are the major limitations when capturing rapid 3D neuron dynamics in cm-sized mouse brains. In this work, a tunable acoustic gradient-index (TAG) lens and a rod-like gradient-index (GRIN) lens are integrated into 2PM, respectively providing axial scan at 100 kHz ~ 1 MHz and extending imaging depth to ~ 1 cm, thus enabling deep brain imaging with 1~10 Hz volume rate in a ~380 × 380 × 150 µm³ volume. This technique allows, for the first time, the observation of volumetric calcium dynamics in an arbitrary region within a mouse brain at sub-cellular and sub-second spatiotemporal resolution, which paves the way towards studying the functionality in deep-brain regions.

Keywords: Gradient-index lens, TAG lens, Two-photon microscopy, Volumetric imaging, Functional imaging