A Proof-of-Principle of Nanoscale Optogenetics

Optogenetics has revolutionized the study of circuit function in the brain, by allowing activation of specific ensembles of neurons by light. However, this technique has not yet been exploited extensively at the subcellular level. Here we propose a novel focal stimulation approach using STED/RESOLFT-like illumination, whereby switchable light-gated channels are focally activated by a laser beam of one wavelength and deactivated by an overlapping donut-shaped beam of a different wavelength, confining activation to a center focal region. We demonstrate the utility of current optogenetic tools to achieve highly focal depolarization using this method and further examine a proof-of-principle of nanoscale optogenetic activation using an initial macroscale approach. When employed at the nanoscale, this approach will allow unprecedented optogenetic control of nanodomains within cells.