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. 2010 Sep 1;35(13):2521–2537. doi: 10.1038/npp.2010.117

Figure 4.

Figure 4

CBP-dependent regulations in the rat dorsal hippocampus during the formation of a spatial memory. (a) Representative images of CBP immunoreactive staining revealed with DAB staining in regions CA1 (A, B) and in the dentate gyrus (dg; C, D) of the dorsal hippocampus in representative VPf rats (A, C) and HPf (B, D) rats. Pyr: pyramidal neuron layer; sr: stratum radiatum; dg: dentate gyrus. Scale bar=250 μm. (b) Mean (±SEM) optical density of CBP immunoreactive staining in the CA1 pyramidal cell layer (CA1) and in the dentate gyrus granular layer (DG). In HPf rats (n=5), whatever the region, the optical density of CBP immunoreactivity was significantly larger than in VPf rats (n=5). *p<0.05. (c) Chromatin immunoprecipitation of acetylated histones were performed on chromatin obtained from 3-day-trained rats from the VPf and HPf groups (n=3). Semiquantitative PCR were performed to analyze the presence of cbp proximal promoter in the immunoprecipitates. Controls for immunoprecipitation were performed with an unrelated control antibody (Ct-IgG) or no antibody (noAb, not shown). Representative results are shown in duplicate for each gene tested. Bands were quantified (ImageJ) and fold inductions (f.i.) are indicated in the table below. acH3: acetylated K9, K14 H3 histone; acH4: acetylated K12 H4 histone; and acH2B: acetylated K12, K15 H2B histone. (d) Chromatin immunoprecipitation of CBP were performed on chromatin obtained from 3-day-trained rats from the VPf and HPf groups (n=3). Semiquantitative PCR were performed to analyze the presence of bdnf-pIV, cFos, FosB, zif268, and cbp proximal promoters in the immunoprecipitates. Controls for immunoprecipitation were performed with an unrelated control antibody (Ct-IgG, shown in simplicate) or no antibody (noAb, not shown). Representative results are shown in duplicate for each gene tested. Bands were quantified (ImageJ) and fold inductions (f.i.) are indicated in the table on the right. Bdnf-eIV: bdnf exon IV; bdnf-pIV: bdnf promoter IV. (c, d) Input DNA corresponding to the total of chromatin before immunoprecipitation was run in parallel as positive control. An activation score was arbitrarily affected by the HPf vs VPf condition: (+++) when f.i. >2.00; (++) when f.i.>1.50; (+) when f.i. >1.25; (=) when 1.25<f.i.>0.75; and (−) when f.i.<0.75.