Abstract Genetically encoded optical sensors and advancements in microscopy instrumentation and techniques have revolutionized the scientific toolbox available for probing complex biological processes such as release of specific neurotransmitters. Most genetically encoded optical sensors currently used are based on fluorescence and have been highly successful tools for single-cell imaging in superficial brain regions. However, there remains a need to develop new tools for reporting neuronal activity in vivo within deeper structures without the need for hardware such as lenses or fibers to be implanted within the brain. Our approach to this problem is to replace the fluorescent elements of the existing biosensors with bioluminescent elements. This eliminates the need of external light sources to illuminate the sensor and overcomes several drawbacks of fluorescence imaging such as limited light penetration depth, excitation scattering, and tissue heating that are all associated with the external light needed for fluorescence imaging. Here we report the development of the first genetically encoded neurotransmitter indicators based on bioluminescent light emission. These probes exhibit robust changes in light output in response to extracellular presentation of the excitatory neurotransmitter glutamate. We expect this new approach to neurotransmitter indicator design to enable the engineering of specific bioluminescent probes for multiple additional neurotransmitters in the future, ultimately allowing neuroscientists to monitor activity associated with a specific neurotransmitter as it relates to behavior in a variety of neuronal and psychiatric disorders, among many other applications.

Bioluminescent Genetically Encoded Glutamate Indicator for Molecular Imaging of Neuronal Activity