Fig. 7. Application of our microglia-selective gene expression method in expressing the genetically encoded Ca²⁺ indicator G-CaMP7.09 and measuring Ca²⁺ signals in cerebellar microglia.

a, b Left panels show the averaged confocal images of virally expressed G-CaMP signals in the granule cell layer of acute cerebellar slices. Regions of interest (ROIs 1–4) were set on larger compartments (possibly microglial somata or their larger processes) in a and on smaller compartments (possibly fine microglial processes) of the microglia in b. The right panels show traces of Ca²⁺ signal changes estimated from the fluorescence of the ROIs illustrated in the left panels. Bath application of 100 µM ATP (indicated in black bars) evoked Ca²⁺ transients robustly in both the larger and the smaller compartments of the transduced microglia. c A box and whisker plot showing quantified Ca²⁺ signals induced by bath-applied ATP (∆F/F_basal, see methods) in the microglial cellular compartments. Open circles indicate individual data points. The horizontal line and the box represent the median value and the interquartile range, respectively. The error bars indicate one standard deviation above and below the mean value (filled circle). d Time-lapse fluorescent images showing the movement of a microglial process expressing G-CaMP7.09 (indicated by a white circle). Note that the other G-CaMP-positive microglial compartments outside the white circle were static.