Figure 3:

Histone modification is involved in the increase of NMDAR protein expression in mPFC of D2R-GSK3^−/− mice. A, representative Western blots and summary histograms showed that total protein levels of NMDAR and AMPAR subunits from the mPFC of D2R-GSK-3β^+/+ and D2R-GSK-3β^−/− mice. Both NR2A and NR2B were significantly increased in D2R-GSK-3β^−/− mice (n = 8 for each group, *p < 0.05 for both), but NR1, GluR1, and GluR2 were unchanged (n = 8 for each group, p > 0.05 for all). B, representative Western blots and summary histograms showed that total protein levels of HDAC2 & 4, as well as three acetylation sites of H3K from mPFC tissue of D2R-GSK-3β^+/+ mice and D2R-GSK-3β^−/− mice. HDAC2 was decreased while HDAC4 was increased in D2R-GSK-3β^−/− mice (vs. D2R-GSK-3β^−/− mice, n = 6 for each group, *p < 0.05 for both). Both H3K18ac and H3K27ac, but not H3K9ac, were increased in D2R-GSK-3β^−/− mice (vs. D2R-GSK-3β^−/− mice, n = 6 for each group, *p < 0.05). C, chromatin was pooled from the PFC of 2 animals from D2R-GSK-3β^+/+ or D2R-GSK-3β^−/− mice and immunoprecipitated using antibodies against H3K27ac (left) and HDAC2 (right). qPCR was performed using a primer set specific to the promoter region of Grin1, Grin2a, and Grin2b. The signal of the amplified DNA was normalized to input. H3K27ac enrichment at either Grin2a or Grin2b but not Grin1 were significantly enhanced in D2R-GSK-3β^−/− (vs. D2R-GSK-3β^+/+, n = 6 mice yielding 3 data points for each group, *p < 0.05, **p < 0.01). On the contrary, HDAC2 enrichment at Grin2b, but not Grin1 and Grin2a, was significantly reduced in D2R-GSK-3β^−/− mice (vs. D2R-GSK-3β^+/+, n = 6 mice yield 3 data points for each group, *p < 0.05).