Fig. 2. A simple kinetic model sufficed to describe CA1 pyramidal cell spine dynamics.
(a) Two-photon microendoscopy and stimulated emission depletion (STED) imaging of the same dendrites in vitro. (Top row) two-photon images depict spines closer than the resolution limit as merged entities. (Bottom) Asterisks mark example, visually scored spines, showing cases in which nearby spines do (right) or do not (left) merge.
(b) Fraction of spines (N = 151 total) seen by two-photon imaging, that were one, two or three spines as determined by STED imaging. Error bars: s.d.
(c) Separations between adjacent unmerged spines and pairs of spines that appeared merged by two-photon imaging. Open grey circles mark individual results from each of N = 150 spines. Black bars: mean ± s.d.
(d) Example, computer-simulated, time-lapse image sequence, used to quantify how resolution limits impact measured spine densities and dynamics.
(e) Computational modeling predicts the underestimation of spine density due to the finite optical resolution. Blue diagonal line: perfect detection of all spines. Black horizontal dashed lines: typical ranges of spine densities on pyramidal cells in neocortex and hippocampus. Red data: results from visually scoring simulated images of dendrites of varying spine densities. Black curve: prediction from the scoring model using 600 nm as the minimum separation between two spines correctly distinguished.
(f) Modeling predicts the overestimation of spine stability due to merging of adjacent spines in resolution-limited images. Blue data: Survival fraction values (mean ± s.e.m.) for actual spine turnover in computer simulations (spine density:2.56 μm−1). Red data: Apparent turnover for these same simulated dendrites, as scored from simulated two-photon images. Black curves: Theoretical predictions for spine survival based on the scoring model.