Novel Design and Application of High-NA Fiber Imaging Bundles for In Vivo Brain Imaging with Two-Photon Scanning Fluorescence Microscopy

Here, we provide experimental verification supporting
the use of
short-section imaging bundles for two-photon microscopy imaging of
the mouse brain. The 8 mm long bundle is made of a pair of heavy-metal
oxide glasses with a refractive index contrast of 0.38 to ensure a
high numerical aperture NA = 1.15. The bundle is composed of 825 multimode
cores, ordered in a hexagonal lattice with a pixel size of 14 μm
and a total diameter of 914 μm. We demonstrate successful imaging
through custom-made bundles with 14 μm resolution. As the input,
we used a 910 nm Ti-sapphire laser with 140 fs pulse and a peak power
of 9 × 104 W. The excitation beam and fluorescent
image were transferred through the fiber imaging bundle. As test samples,
we used 1 μm green fluorescent latex beads, ex vivo hippocampal
neurons expressing green fluorescent protein and cortical neurons
in vivo expressing the fluorescent reporter GCaMP6s or immediate early
gene Fos fluorescent reporter. This system can be used for minimal-invasive
in vivo imaging of the cerebral cortex, hippocampus, or deep brain
areas as a part of a tabletop system or an implantable setup. It is
a low-cost solution, easy to integrate and operate for high-throughput
experiments.

本项研究提供了实验验证，证实了短节段成像束在双光子显微镜成像小鼠脑部时的应用价值。该8毫米长的束由一对具有折射率对比度0.38的重金属氧化物玻璃构成，以确保达到高数值孔径NA = 1.15。该束由825个多模光纤芯组成，以14微米像素尺寸和914微米的总直径排列成六角形晶格。我们展示了通过定制的成像束实现14微米分辨率成像的成功案例。作为输入，我们使用了910纳米的Ti-sapphire激光器，其脉冲宽度为140飞秒，峰值功率为9 × 10^4瓦。激发光束和荧光图像通过光纤成像束传输。作为测试样本，我们使用了1微米的绿色荧光乳胶珠、体外表达绿色荧光蛋白的海马神经元以及体内表达荧光报告基因GCaMP6s或即刻早期基因Fos荧光报告基因的皮层神经元。该系统可作为桌面系统或植入式装置的一部分，用于大脑皮质、海马或深部脑区的最小侵入性体内成像。它是一种低成本、易于集成和操作的高通量实验解决方案。