Fig. 4

SLC7A10 immunoreactivity is detected in Olig2-astrocytes. a The specificity of the primary antibody was confirmed by pre-adsorption assay. Protein from mouse brainstem (15 μg) was separated on by SDS-PAGE and reacted with anti-SLC7A10, pre-adsorbed anti-SLC7A10 and anti-GAPDH antibody. Pre-adsorption was performed with excess SLC7A10 recombinant peptide. The 33-kDa band labeled with SLC7A10 antibody (arrow) was eliminated by incubation with pre-adsorbed antibody. b Immunohistochemistry was performed on the mouse brain and spinal cord sections. Immunoreactivity of SLC7A10 was detected in the caudal brain, brainstem and gray matter of the spinal cord. c Immunoreactivity of SLC7A10 was completely eliminated in the staining with pre-adsorbed antibody. d–f Immunofluorescence staining was performed on brain sections of Olig2-Ai27 mice. Strong SLC7A10 immunoreactivity (d) was observed in the LGP where tdTomato-labeled Olig2-lineage cells (e) were localized (f merged image). The border between the caudo-putamen (CPu) and the LGP is marked with a dashed line. g–i SLC7A10-immunoreactive cells in the LGP. The immunoreactive cells have bushy morphology (g demarcated with dashed lines) and show overlapping distribution with Olig2-astrocytes labeled with tdTomato (h). Merged image (i). j–m 3D-deconvolution of stacked images of astrocytic processes. SLC7A10 immunoreactivity (j) was colocalized on Olig2-astrocytic processes labeled with tdTomato (k). Merged image (l). m Enlarged image of the boxed region in l. Note the clear co-localization of SLC7A10 immunoreactivity and tdTomato fused with channelrhodopsin (H134R) on the plasma membrane (arrowheads). Scale bars: f 500 μm also for d and e; i 50 μm also for g and h; l 5 μm also for j and k; m 2 μm