Protocol for imaging and analyzing the synaptic coverage of astrocytes with super-resolution STED microscopy in mouse brain slices

Super-resolution has allowed us to image areas previously inaccessible due to the limits of standard microscopy techniques. Here, we present a protocol to determine the astroglial coverage of synapses. We detail the steps to obtain and label mouse brain slices followed by imaging using 3D stimulated emission depletion (STED) microscopy and analysis using a custom-made Fiji plug-in, with imaging at a lateral resolution of 80 nm. This technique ascertains the interaction between astrocytes and the glial cell's role in signaling and brain region function.