Light-sheet functional imaging in fictively behaving zebrafish

The processing of sensory input and the generation of behavior involves large networks of neurons 1,2, which necessitates new technology 3–7 for recording from many neurons in behaving animals. In the larval zebrafish, light-sheet microscopy can be used to record the activity of almost all neurons in the brain simultaneously at single-cell resolution 3,4. Existing implementations, however, cannot be combined with visually driven behavior because the light sheet scans over the eye, interfering with presentation of controlled visual stimuli. Here we describe a system that overcomes the confounding eye stimulation through the use of two light sheets and combines whole-brain light-sheet imaging 3 with virtual reality for fictively behaving 1 larval zebrafish.