Fig. 6

Double staining of CFs and microglia in the developing cerebellum. a Orthogonal fluorescence images of Iba1-positive microglia (green) and CFs anterogradely labeled with Alexa Fluor 568-dextran (magenta) in C57BL/6 mice. The xy-image is a single plane with its position represented as white lines in the xz and zy images. b Serial enlarged images (∆z = 1 μm, from the left top to the right bottom) of a region surrounded by a dotted line in (a). Alexa Fluor 568-labeled CFs were very closely associated with Iba1-positive microglial processes but were not internalized in microglia (arrows, arrowheads, and double arrowheads). c–f Orthogonal fluorescence images of other microglia. g One of the rare examples in which an isolated CF fragment was internalized in Iba1-positive microglia. h Orthogonal fluorescence images similar to a but taken from C57BL/6 mice administrated 3-AP, harmaline and nicotinamide. i Serial enlarged images of a region surrounded by a dotted line in h. j, k Orthogonal fluorescence images of other 3-AP treated microglia. Fragments of Alexa Fluor 568-labeled CFs are included in Iba1-positive microglia (arrows and arrowheads in h–k), indicating that the fluorescence of Alexa Fluor 568 can be detected after being engulfed by microglia. Scale bars, 10 μm