Voltage imaging: A practical guide.

Recent advances in genetically encoded voltage indicators (GEVIs), optical instrumentation, and analysis software have brought voltage imaging within reach of most neuroscience labs. Voltage imaging can resolve subthreshold dynamics and action potentials across scales, from the flickering of single ion channels to large-scale neural population dynamics in behaving animals. This primer provides a practical guide to the choices involved in setting up a voltage imaging experiment, including selecting a voltage indicator, choosing excitation wavelengths and microscopy configurations, expressing the constructs in the desired cell types, and extracting information from the data. We discuss the physical principles that govern signal-to-noise ratio, the trade-offs between one-photon and two-photon excitation, and the use of structured illumination to improve contrast in tissue. Our aim is to provide a reference for new and seasoned voltage imagers on how to design effective voltage imaging experiments.