Simultaneous multiplane imaging with reverberation two-photon microscopy

Multiphoton microscopy has gained enormous popularity because of its unique capacity to provide high-resolution images from deep within scattering tissue. Here, we demonstrate video-rate multiplane imaging with two-photon microscopy by performing near-instantaneous axial scanning while maintaining three-dimensional micrometer-scale resolution. Our technique, termed reverberation microscopy, enables the monitoring of neuronal populations over large depth ranges and can be implemented as a simple add-on to a conventional design. Reverberation two-photon microscopy enables video-rate multiplane neuroimaging by performing near-instantaneous axial scanning over large depth ranges while maintaining 3D micrometer-scale resolution.