Fig. 3

Optogenetic microglia depolarization decelerates chemotactic response kinetics. a Graphic illustration and representative images of microglia chemotaxis towards an induced laser-damage. b Two-photon maximum projections of the chemotactic response 0, 9, and 24 min after the laser-damage. c Two-photon z-projection of a patched microglia during chemotaxis. d Voltage-clamp recordings of patched microglia during chemotaxis. Gray: Individual microglial responses from four experiments, red: Average of all experiments. Left: no light stimulation during laser-damage, right: with light stimulation during laser damage. e Automated MATLAB analysis of chemotaxis quantified as the reduction in microglia-free area around the laser damage (black polygon) at different time points of the experiment. f Relative laser damage response measured as microglia-free area. Black: Control slices (no construct) with light stimulation (n = 8 areas, 5 slices). Gray: Experiments with ChETA expression in microglia, but without light stimulation (n = 7 areas, 4 slices). Blue: Slices with ChETA expression in microglia combined light stimulation (n = 11 areas, 7 slices). Insert: Graphic representation of light stimulation protocol between stack acquisitions. 2-way ANOVA (‡ control480 – ChETA480, p < 0.001, († ChETA no light – ChETA480, p < 0.01). g Time to 50% engulfment was prolonged by optogenetic depolarization. h 9 min after injury, the microglia-free area was larger when microglia were depolarized. g, h One-way ANOVA with Tukey’s post hoc comparison (*p < 0.05, **p < 0.01, ***p < 0.001)