Pharmacological antagonism of kainate receptor rescues dysfunction and loss of dopamine neurons in a mouse model of human parkin-induced toxicity

摘要

a Representative Western blot showing the levels of GluK2 subunit of KAR and GluA1 subunit of AMPAR in lysates prepared from the substantia nigra of WT or parkinQ311X mice at 1 month of age. Membranes were also probed for TH as DA neuron marker. Samples were run in triplicate, with each lane loaded with lysate from an individual mouse. The histograms on the right show the means ± SEM from densitometer quantifications. For GluK2/KAR analysis, all the bands between 80 and 120 kDa were quantified. GluK2/KAR: WT 1.00 ± 0.07 n = 5 mice, parkinQ311X 1.35 ± 0.10 n = 6 mice, two-tailed unpaired Student t test *p = 0.012, t = 2.786, df = 9; GluA1: WT 1.00 ± 0.09 n = 6 mice, parkinQ311X 0.94 ± 0.06 n = 6 mice, two-tailed unpaired Student t test p > 0.05. b Representative confocal images showing TH (red) and GluK2 (green) double immunofluorescence in the SNc of the WT and the parkinQ311X mice at 1 month of age. The image is representative of 12 sections derived from n = 3 WT mice and 12 sections from n = 3 parkinQ311X mice (GluK2 antibody Millipore 04-921). The bar is 30 µm. The histogram shows the means ± SEM of fluorescence intensity. Each data point on the graph represents one neuron. Twelve sections per genotype were examined, derived from n = 3 WT mice and n = 3 parkinQ311X mice. GluK2/TH: WT 1.00 ± 0.10 n = 48 neurons, parkinQ311X 2.24 ± 0.08 n = 53 neurons, two-tailed unpaired Student t test ****p = 3.87E–16, t = 9.749, df = 99. Data from non-normalized GluK2 intensity: WT 1.00 ± 0.12 n = 48 neurons, parkinQ311X 3.22 ± 0.13 n = 53 neurons, two-tailed unpaired Student t test ****p = 5.03E−22, t = 12.47, df = 99. c Representative images showing TH immunoperoxidase labeling in the SNc of the WT and the parkinQ311X mice at 1 and 6 months of age. The histograms just below the picture show DA neuron quantification performed by stereological count. Data are the means ± SEM of TH-positive neurons: WT 1 month (1 M) 4817 ± 146 n = 10 mice, parkinQ311X 1 M 5160 ± 182 n = 10 mice, WT 6 months (6 M) 5075 ± 270 n = 11 mice, and parkinQ311X 6 M 3913 ± 293 n = 10 mice, one-way ANOVA followed by Tukey’s test. F = 5.933, WT 6 M vs. parkinQ311X 6 M. **p = 0.0054, parkinQ311X 1 M vs. parkinQ311X 6 M, **p = 0.0033.