Optimizing strategies for developing genetically encoded voltage indicators

Genetically encoded optical indicators of neuronal activity enable unambiguous recordings of input-output activity patterns from identified cells in intact circuits. Among them, genetically encoded voltage indicators (GEVIs) offer additional advantages over calcium indicators as they are direct sensors of membrane potential and can adeptly report subthreshold events and hyperpolarization. Here, we outline the major GEVI designs and give an account of properties that need to be carefully optimized during indicator engineering. While designing the ideal GEVI, one should keep in mind aspects such as membrane localization, signal size, signal-to-noise ratio, kinetics and voltage dependence of optical responses. Using ArcLight and derivatives as prototypes, we delineate how a probe should be optimized for the former properties and developed along other areas in a need-based manner. Finally, we present an overview of the GEVI engineering process and lend an insight into their discovery, delivery and diagnosis.