Striatal Histopathological Changes in Huntington's Disease Mouse Model

To examine the histopathological changes in the striatum, 10 weeks (14 weeks after birth) after AAV-DJ-82Q injection, the mice (n = 7 per group) were deeply anesthetized with diethyl ether and then intracardially perfused with saline and cold 4% paraformaldehyde in 0.1 M phosphate buffer (PB, pH 7.4). Brains were removed and post-fixed for 24 h and cryoprotected in 10, 20 and 30% sucrose in PBS serially at 4°C. Serial coronal slices (30-μm thickness) of striatum were acquired on a model CM3050S freezing microtome (Leica, Wetzlar, Germany) and collected in sequence as free-floating sections on PBS. The sections from the mid-coronal level of the striatum (Franklin and Paxinos, 2008 ) were processed for immunofluorescence stain as previously described (Lee et al., 2016a (link)). Briefly, the brain sections from each group (n = 3 per brain) were incubated with mouse anti-HTT protein (1:400; catalog MAB5374, clone EM48; Millipore), rabbit anti-NeuN (1:5,000; Abcam), rabbit anti-cleaved caspase-3 (1:500; Cell Signaling Technology), rabbit anti-Iba-1 (1:2,000; WAKO), mouse anti-GFAP (1:5,000; Millipore) and rabbit anti-Olig2 (1:500; Abcam) overnight with gentle agitation at room temperature. The sections were followed with a mixture of secondary antibodies and examined with confocal imaging system (LSM five PASCAL; Carl Zeiss, Germany).