Rowlands, Christopher JPark, DemianBruns, Oliver TPiatkevich, Kiryl DFukumura, DaiJain, RakeshBawendi, Moungi GBoyden, Edward SSo, Peter TC2017-12-062017Rowlands, Christopher J, Demian Park, Oliver T Bruns, Kiryl D Piatkevich, Dai Fukumura, Rakesh K Jain, Moungi G Bawendi, Edward S Boyden, and Peter TC So. 2017. “Wide-field three-photon excitation in biological samples.” Light, science & applications 6 (1): e16255. doi:10.1038/lsa.2016.255. http://dx.doi.org/10.1038/lsa.2016.255.http://nrs.harvard.edu/urn-3:HUL.InstRepos:34493099Three-photon wide-field depth-resolved excitation is used to overcome some of the limitations in conventional point-scanning two- and three-photon microscopy. Excitation of chromophores as diverse as channelrhodopsins and quantum dots is shown, and a penetration depth of more than 700 μm into fixed scattering brain tissue is achieved, approximately twice as deep as that achieved using two-photon wide-field excitation. Compatibility with live animal experiments is confirmed by imaging the cerebral vasculature of an anesthetized mouse; a complete focal stack was obtained without any evidence of photodamage. As an additional validation of the utility of wide-field three-photon excitation, functional excitation is demonstrated by performing three-photon optogenetic stimulation of cultured mouse hippocampal neurons expressing a channelrhodopsin; action potentials could reliably be excited without causing photodamage.en-USbiophotonicsmultiphoton microscopyoptogeneticstemporal focusingthree-photonJournal Article2017-12-0610.1038/lsa.2016.255