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Genetically encoded voltage indicators (GEVIs) enable monitoring of neuronal activity at high spatial and temporal resolution. However, the utility of existing GEVIs has been limited by the brightness and photostability of fluorescent proteins and rhodopsins. We engineered a GEVI, called Voltron, that uses bright and photostable synthetic dyes instead of protein-based fluorophores, thereby extending the number of neurons imaged simultaneously in vivo by a factor of 10 and enabling imaging for significantly longer durations relative to existing GEVIs. We used Voltron for in vivo voltage imaging in mice, zebrafish, and fruit flies. In the mouse cortex, Voltron allowed single-trial recording of spikes and subthreshold voltage signals from dozens of neurons simultaneously over a 15-minute period of continuous imaging. In larval zebrafish, Voltron enabled the precise correlation of spike timing with behavior.
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This content will become publicly available on January 1, 2027
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 the 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, a 250 µm field of view, and a 1.3-1.6 mm working distance that weighs 16.4 g. We show it is capable of imagingin vivovoltage spikes from Voltron2 with a spike peak-to-noise ratio >3 at a framerate of 530 Hz.
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- Award ID(s):
- 2319406
- PAR ID:
- 10710932
- Publisher / Repository:
- Optica
- Date Published:
- Journal Name:
- Biomedical Optics Express
- Volume:
- 17
- Issue:
- 1
- ISSN:
- 2156-7085
- Page Range / eLocation ID:
- 1
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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