Immunohistochemical Analyses of Striatal Markers

Striatum slices (30 μm) prepared in a Leica 1000S vibratome (Nussloch, Germany) were incubated with the antibodies rabbit anti-myelin basic protein (MBP; 1:400, Abcam, Cambridge, UK), rabbit anti-ionized calcium-binding adapter molecule 1 (Iba1; 1:1000, Wako Pure Chemical Industries, Japan), and rabbit anti-neuroglycan-2 (NG2; 1:200, Abcam, Cambridge, UK) as described [12 (link)]. After mounting the slices in Fluoroshield™ (Sigma-Aldrich, MO, USA), pictures were taken using a FV300 Olympus confocal microscope (Tokyo, Japan). For each animal and staining procedure, 3 sections were stained. In a separate analysis, three brain slices were directly stained at room temperature with 300 μL of green Fluoromyelin^TM (1:300 from the stock solution) (Thermo Fisher Scientific, Waltham, MA, USA) for 30 min. Sections were rinsed, mounted in Fluoroshield™ (Sigma-Aldrich, MO, USA) and imaged using the same microscope. Quantification of fluorescence was performed as described [25 (link),26 (link)].