Fig. 4: Neuron and astrocyte immunohistochemical analysis and mRNA expression of GABA_A receptor subunits.

a Outline of sagittal section of mouse brain, showing the brain regions analyzed using immunohistochemical staining. b–i Coronal sections stained through fluorescence-labeling of: b NeuN in anterior cingulate cortex (ACC); c NeuN in hippocampus (HC); d DCX in dentate gyrus (DG); e parvalbumin (PV) in ACC; f PV in piriform cortex (PC); g PV in DG; h GFAP in CA1 region; and i GFAP in DG. The DG, CA1, and CA3 regions in hippocampus are indicated in part (c). NeuN and PV immunofluorescence are displayed in green (scale bar = 300 μm), and that of DCX and GFAP and DCX displayed in red (scale bar = 120 μm). The plots show immunofluorescence densities for WT (green dots) and KO (red dots) estimated in terms of average area optical density (in parts b, c, e, h, and i), or in terms of the number of immunoreactive cells (in parts d, f, g); in each instance, five randomly selected images from each of five WT or KO male mice were examined. The levels of mRNA expression for 13 different GABA_A receptor subunits in WT and KO mouse cerebrum (j) and cerebellum (k) were measured using quantitative RT-PCR, and the mRNA levels in KO mice were normalized to the average expression level in WT (n = 10 in each group). The measured expression levels in WT, KO as well as HT brains are given in Supplementary Table S4. Statistical analysis was performed using unpaired t-test for immunohistochemical staining and one-way ANOVA with Newman–Keuls post-hoc test for mRNA quantitation. Average y values ± SEM in the different plots are represented by horizontal bars; WT is represented by green dots and KO by red dots. N.D. represents non-detectable; *p < 0.05, **p < 0.01, ***p < 0.001