Miniaturized widefield microscope for high speed in vivo voltage imaging

Functional imaging in freely moving animals with genetically encoded voltage indicators (GEVIs) will open new capabilities for neuroscientists to study the behavioral relevance of neural activity with high spatial and temporal precision. However, miniaturization of an imaging system with sufficient collection efficiency to resolve the small changes in fluorescence yield from voltage spikes, as well as development of efficient image sensors that are sufficiently fast to capture them, has proven challenging. We present a miniaturized microscope designed for voltage imaging, with a numerical aperture of 0.6, 250 μm field of view and 1.3 mm working distance that weighs 16.4 g. We show it is capable of imaging in vivo voltage spikes from Voltron2 with a spike peak-to-noise ratio >3 at a framerate of 530 Hz.