Figure 1.

Dynamic motility of resting microglial processes and connection with synapses. A, Images of a single resting microglial cell in the intact cortex of an Iba1–EGFP mouse. Each is a stack image composed of 70–80 focal planes, each advancing 0.5 μm along the z-axis. Scale bar, 20 μm. Resting microglia have extensive processes which extend in all directions. These processes are dynamic, moving back and forth. B, Continuous monitoring of the motility of three representative microglial processes (from 3 different cells) using free-scanning imaging over 20 min. The absolute position of each process was expressed relative to a fixed starting point and summed into 12 s bins. Note a brief (∼4–5 min) pause, indicated by the red arrows, in process motility. C, Contacts between microglial processes (yellow arrow in each image) and a presynaptic bouton (blue arrows in top row) and a postsynaptic spine (blue arrow in bottom row). Images in each row (-pre, -post) were obtained from single-plane free-scanning images at a higher resolution (scale bars: -pre, 2 μm; -post, 4 μm) (see supplemental Videos 1-Presynapse and 2-Postsynapse, available at www.jneurosci.org as supplemental material). Note that the tip of the microglial process is enlarged during the contact. Left, middle, and right panels in each row were obtained before, during, and after the contact of the microglial process with the synaptic structure, respectively. D, Plots of fluorescence intensity as a function of distance along a straight line between a bouton and adjacent microglial process, indicated by the red broken line in C, -pre, left. The fluorescent intensity plot before contact (left) has two peaks, the left peak associated with the microglial process and the right with the presynaptic bouton. A region of minimum intensity, an intensity trough, is indicated by the black arrow and signifies a clear separation between the two structures. As the microglial process and the bouton came into contact, there was a loss of this fluorescence intensity trough (center). After the 5 min contact, the fluorescence intensity trough reappeared (right).