Fig. 3.
In vivo volumetric three-photon fluorescence microscopy of micebrains. Maximum intensity projection of a (a) 62-μm and (c) 52-μm thick volume at ∼80 μm below dura, color-coded by depth, imaged with a Gaussian focus, at a2-μm axial step size, under 1300 nm excitation. Post-objective powers: 3 and 4 mW, respectively. (b,d) Image of the same volume as (a,c), respectively, obtained with a Bessel focus of NA 0.6 and axial FWHM of 38 μm.White arrowheads label dendritic spines, and yellow arrowheads label axonal boutons.Post-objective powers: 30 and 40 mW, respectively.(e) Maximum intensity projection of a 30-μm thick volume at ∼290 μm below dura, color-coded by depth, imaged with a Gaussian focus, at a2-μm axial step size, under 1700 nm excitation. Post-objective power: 15 mW. (f) Image of the same volume as (e) obtained with a Bessel focus of NA 0.6 and axial FWHM of 27 μm. Post-objective power: 73 mW.A Thy1-GFP line M mouse was used for (a-d), and a Gad2-IRES-Cre × Ai14 for(e,f).