Molecular Biology

A central goal in neuroscience is to understand dynamics in the awake mammalian brain on a large scale that is also well-resolved (thousands of individual neurons). Recent optical methods for cellular-resolution fluorescence imaging using two-photon excitation (TPE) microscopy, or population-resolution optogenetic perturbation of activity, have been applied to measure or probe dynamics of neurons in behaving ro-dents. A technique allowing both fluorescence imaging and optogenetic perturbation at cellular resolution could introduce a new class of experiments to probe functional properties of the brain. Here, an approach is presented allowing all-optical stimula-tion and imaging of neurons in mice. We show that single, task-modulated neurons may be characterized and then manipulated during a behavior. We first demonstrate that channelrhodopsin-2 (ChR2), a visible-wavelength sen-sitive optogenetic cation channel, produces photocurrents under infrared TPE. An empirical estimate of ChR2’s two-photon absorption cross-section at λ=920 nm is presented, with a value (260±20 GM) indicating that TPE stimulation of ChR2 i