Futia Gregory L, Zohrabi Mo, McCullough Connor, Teel Alec, Simoes de Souza Fabio, Oroke Ryan, Miscles Eduardo J, Ozbay Baris N, Kilborn Karl, Bright Victor M, Restrepo Diego, Gopinath Juliet T, Gibson Emily A
Department of Bioengineering, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, USA.
Department of Electrical, Computer and Energy Engineering, University of Colorado, Boulder, CO 80309, USA.
bioRxiv. 2024 Oct 22:2024.10.21.619528. doi: 10.1101/2024.10.21.619528.
Multiphoton microscopy combined with optogenetic photostimulation is a powerful technique in neuroscience enabling precise control of cellular activity to determine the neural basis of behavior in a live animal. Two-photon patterned photostimulation has taken this further by allowing interrogation at the individual neuron level. However, it remains a challenge to implement imaging of neural activity with spatially patterned two-photon photostimulation in a freely moving animal. We developed a miniature microscope for high resolution two-photon fluorescence imaging with patterned two-photon optogenetic stimulation. The design incorporates a MEMS scanner for two-photon imaging and a second beam path for patterned two-photon excitation in a compact and lightweight design that can be head-attached to a freely moving animal. We demonstrate cell-specific optogenetics and high resolution MEMS based two-photon imaging in a freely moving mouse. The new capabilities of this miniature microscope design can enable cell-specific studies of behavior that can only be done in freely moving animals.
多光子显微镜与光遗传学光刺激相结合是神经科学中的一项强大技术，能够精确控制细胞活动，以确定活体动物行为的神经基础。双光子图案化光刺激通过允许在单个神经元水平进行研究，进一步推动了这一技术的发展。然而，在自由移动的动物中，利用空间图案化双光子光刺激实现神经活动成像仍然是一项挑战。我们开发了一种微型显微镜，用于高分辨率双光子荧光成像以及图案化双光子光遗传刺激。该设计集成了一个用于双光子成像的微机电系统（MEMS）扫描仪和一个用于图案化双光子激发的第二光束路径，采用紧凑且轻便的设计，可以头戴在自由移动的动物身上。我们在自由移动的小鼠中展示了细胞特异性光遗传学和基于高分辨率MEMS的双光子成像。这种微型显微镜设计的新功能能够实现只能在自由移动动物中进行的细胞特异性行为研究。
