Figure 2. Density of spines on basal dendrites of CA1 pyramidal neurons in vivo.
(A) CA1 basal dendrite imaged with 2P (top) or 2P-STED (bottom) microscopy. Filled arrowheads highlight spines discerned in 2P-STED, but not 2P mode. Open arrowheads indicate spines that could only be visualized in 2P-STED mode as they were otherwise masked by the blurry fluorescence of the dendrite in the 2P image (MIP of two z-sections). (B) Measured spine densities in consecutive 2P and 2P-STED acquisitions of the same dendrites (p<0.0001, paired t-test; n = 82 basal dendrites, 6 mice). (C) Image of a basal dendrite obtained from fixed brain tissue acquired on a confocal STED. (D) Dendritic spine density on basal dendrites in fixed hippocampal tissue (n = 37 basal dendrites, 6 mice). (E) Geometric model to extrapolate the spine density in 3D. Spines cannot be detected when they are inside the ‘blind zone’, depending on the dimensions of the morphology and microscope PSF. (F) Extrapolation of spine density in 3D following the model in (E) for in vivo (black line) and fixed tissue measurements (red line).
Figure 2—figure supplement 1. Methodology of dendritic spine density analysis in vivo.
