rabbit anti fosb  (Cell Signaling Technology Inc)

Journal: Science Advances

Article Title: Transcriptional regulation of ventral hippocampus-nucleus accumbens circuit excitability drives cocaine seeking

doi: 10.1126/sciadv.adv1236

Figure Lengend Snippet: ( A ) Timeline of cocaine or saline self-administration in male mice. Arrow indicates the time of tissue collection. h, hours. ( B ) Cocaine self-administration performance. Nosepoke responses per min across cocaine (Coc) or saline (Sal) self-administration. Cocaine self-administering mice responded on the active nosepoke more than saline self-administering mice across sessions [two-way repeated measures (RM) analysis of variance (ANOVA): ME of group: F 1,11 = 19.54, P = 0.0010; ChamberXGroup: F 13,143 = 2.169, P = 0.0137], with significantly increased active nosepoke responding on days 6 and 11 to 14 (Holm-Šidák: * P < 0.05). No differences were observed between groups in inactive nosepoke responses. ME, mean error. ( C ) Cocaine or saline infusions per minute across 14 daily 2-hour sessions where responses on an active nosepoke resulted in an intravenous infusion of cocaine (Coc; 0.5 mg/kg per infusion; n = 8) or saline (Sal; n = 5). Cocaine self-administering mice received higher number of infusions across days compared to saline self-administering mice (two-way RM ANOVA: DayXGroup: F 13,143 = 2.911, P = 0.009), with significantly more infusions on days 4 to 14 (Holm-Šidák tests: * P < 0.05 for days 4 to 14). ( D ) Illustration of tissue biopsy punch (red circle) in vCA1. ( E ) Cocaine self-administration enhances ΔFosB in vHPC. vHPC tissue was taken 1 day after the last self-administration session for detection of ΔFosB mRNA or protein. Cocaine self-administration significantly increased ΔFosB mRNA compared to saline (independent samples t test: t 11 = 2.997, * P = 0.0121). F.C., fold change. ( F ) Circuit-tagging procedure. Male mice with crossed Floxed FosB and L10-GFP (KO; FosB fl/fl : Rosa26 eGFP-L10a ) or L10-GFP only (WT; Rosa26 eGFP-L10a ) received retrograde HSV vector expressing Cre (HSVrg-hEf1ɑ-Cre) into NAc to drive GFP and FosB KO in projections to accumbens, including vHPC-NAc projections. All mice were treated with cocaine (Coc; 20 mg/kg, ip) or saline (Sal) for 10 days and 1 day later vHPC coronal brain slices were taken for FosB/ΔFosB immunohistochemistry. ( G ) Representative 20× coronal images of GFP (green, top), FosB/ΔFosB (red, bottom), and a merged image (bottom) in WT-Sal ( n = 5), WT-Coc ( n = 4), and KO-Coc ( n = 3) mice. White arrows denote GFP + neurons coexpressing FosB/ΔFosB. ( H ) Cocaine increases FosB/ΔFosB in vHPC-NAc neurons. Quantification of colabeled GFP + and FosB/ΔFosB + neurons in all three groups. WT-Coc had significantly greater colabeled neurons compared to WT-Sal and KO-Coc (one-way ANOVA: F 2,9 = 18.01, P = 0.0007; Holm-Šidák: ** P < 0.01, *** P < 0.001).

Article Snippet: Immunoprecipitation was carried out overnight at 4°C using 5 μl of rabbit polyclonal FosB antibody (lot 3; 2251S, Cell Signaling Technology) or 2 μg of rabbit polyclonal IgG isotype (3900S, Cell Signaling Technology).

Techniques: Saline, Plasmid Preparation, Expressing, Immunohistochemistry

Journal: Science Advances

Article Title: Transcriptional regulation of ventral hippocampus-nucleus accumbens circuit excitability drives cocaine seeking

doi: 10.1126/sciadv.adv1236

Figure Lengend Snippet: ( A ) Illustration of dual virus CRISPR method to selectively knockout FosB/ΔFosB in vHPC-NAc neurons. ( B ) Cocaine CPP procedure and timeline. Male controls (Con; n = 15) or FosB crKO (crKO; n = 16) mice underwent cocaine CPP in a three-chamber box for twice-daily conditioning sessions of saline and cocaine (5 mg/kg, ip) followed by a drug-free posttest. ( C ) Saline-paired (Sal) and cocaine-paired (Coc) chamber time during the posttest for Con and crKO mice. Con mice spent significantly more time in the Coc chamber than in the Sal chamber; however, crKO had no differences in time spent between chambers (two-way RM ANOVA: ChamberXGroup: F 1,29 = 4.738, P = 0.0378; Holm-Šidák: ** P < 0.01). ( D ) Cocaine-paired and cocaine-unpaired (i.e., saline) chamber difference in Con and crKO mice. crKO had significantly reduced paired-unpaired time compared to Con (independent samples t test: t 29 = 2.177, * P = 0.0378). ( E ) Forced abstinence–induced seeking procedure. h, hours. ( F ) Con ( n = 6) and crKO ( n = 12) mice did not differ in the number of infusions across cocaine self-administration sessions. ( G ) After a 7-day forced abstinence in their homecage, Con increased the number of mock infusions per minute during a seeking test compared to crKO (independent samples t test: t 15 = 2.377, * P = 0.0312). Created in BioRender. Eagle, A. (2025) https://BioRender.com/tfyofsj .

Article Snippet: Immunoprecipitation was carried out overnight at 4°C using 5 μl of rabbit polyclonal FosB antibody (lot 3; 2251S, Cell Signaling Technology) or 2 μg of rabbit polyclonal IgG isotype (3900S, Cell Signaling Technology).

Techniques: Virus, CRISPR, Knock-Out, Saline

Journal: Science Advances

Article Title: Transcriptional regulation of ventral hippocampus-nucleus accumbens circuit excitability drives cocaine seeking

doi: 10.1126/sciadv.adv1236

Figure Lengend Snippet: ( A ) Illustration of ex vivo patch-clamp approach (top), representative spike traces (middle), and timeline (bottom) for patched vCA1 neurons from cocaine (Coc; n = 4 mice; n = 17 cells) and saline (Sal; n = 3; n = 13 cells) self-administering male mice. Scale bar: 50 pA, 200 ms. ( B ) Cocaine self-administration reduces vCA1 neuron spike frequency (two-way RM ANOVA: ME of group: F 1,28 = 7.320, P = 0.0115; CurrentXGroup: F 11,308 = 1.993, P = 0.0286; Holm-Šidák: * P < 0.05, # P < 0.10). ( C ) Cocaine self-administration reduces the total number of spikes (independent samples t test: t 28 = 2.705, * P = 0.0115). ( D ) There were no significant differences between groups on rheobase. ( E ) Circuit-tagging procedure to target vCA1-NAc neurons (top) and self-administration timeline (bottom) for cocaine (WT-Coc; n = 6 mice; n = 28 cells) or saline mice (WT-Sal; n = 5 mice; n = 17 cells) mice. ( F ) Cocaine self-administration significantly reduced spike frequency in vCA1-NAc neurons (two-way RM ANOVA: CurrentXGroup: F 12,516 = 4.674, P < 0.0001; Holm-Šidák: * P < 0.05, # P < 010). ( G ) Cocaine self-administration significantly reduced total spikes (independent samples t test: t 43 = 2.230, * P = 0.0310). ( H ) There was a trend toward a significant increase in rheobase in vCA1-NAc neurons of WT-Coc mice (independent samples t test: t 43 = 1.719, # P = 0.0928). ( I ) Circuit-tagging procedure to target vCA1-NAc neurons (top) and experimenter-administered injection timeline (bottom). WT mice received saline (WT-Sal; n = 8 mice; n = 23 cells) or cocaine (WT-Coc; n = 6 mice; n = 17 cells). FosB KO mice received cocaine (KO-Coc; n = 4 mice; n = 13 cells). ( J ) Cocaine significantly reduced spike frequency in WT mice (two-way RM ANOVA: CurrentXGroup: F 22,550 = 3.545, P < 0.0001; Holm-Šidák: WT-Sal versus WT-Coc, * P < 0.05). KO-Coc reduced spike frequency compared to WT-Coc (Holm-Šidák: WT-Coc versus KO-Coc, * P < 0.05 for 225 to 300 pA). ( K ) There was a trend toward a significant difference in the total number of spikes in WT-Coc compared to those in WT-Sal and KO-Coc (one-way ANOVA: F 2,50 = 2.728, P = 0.0751; Holm-Šidák: # P < 010). ( L ) There were no differences between groups on rheobase. Created in BioRender. Eagle, A. (2025) https://BioRender.com/s4xwqz2 .

Article Snippet: Immunoprecipitation was carried out overnight at 4°C using 5 μl of rabbit polyclonal FosB antibody (lot 3; 2251S, Cell Signaling Technology) or 2 μg of rabbit polyclonal IgG isotype (3900S, Cell Signaling Technology).

Techniques: Ex Vivo, Patch Clamp, Saline, Injection

Journal: Science Advances

Article Title: Transcriptional regulation of ventral hippocampus-nucleus accumbens circuit excitability drives cocaine seeking

doi: 10.1126/sciadv.adv1236

Figure Lengend Snippet: ( A ) Circuit-tagging and TRAP-seq approach [modified from ]. ( B ) Differentially regulated genes in vHPC-NAc neurons between WT-Coc versus WT-Sal mice. Gray indicates nonsignificant (N.S.) genes. ( C ) Differentially regulated genes in vHPC-NAc neurons between FosB KO-Coc versus WT-Coc mice. ( D ) vHPC tissue punches from cocaine (Coc; n = 8) and saline (Sal; n = 5) self-administering male mice were processed for Calr qPCR. Cocaine self-administration increases Calr mRNA in vHPC (independent samples t test: t 11 = 2.424, * P = 0.0338). ( E ) vHPC tissue punches from cocaine (Coc, C; n = 6) and saline (Sal, S; n = 6) self-administering male mice were processed for Western blotting for calreticulin. Cocaine increases calreticulin in vHPC (independent samples t test: t 10 = 2.428, * P = 0.0356). ( F ) Pooled vHPC tissue samples ( n = 3 mice per sample; n = 4 samples per group) from experimenter-administered cocaine (10 days; Coc; 20 mg/kg, ip; n = 12) and saline male mice (Sal; n = 12) were processed for FosB/ΔFosB ChIP-PCR for Calr . Cocaine increased FosB/ΔFosB binding (% input) to the Calr promoter (two-way ANOVA: InputXGroup: F 1,12 = 21.64, P = 0.0471; Holm-Šidák: **** P < 0.0001). ( G ) Circuit-tagged male WT and FosB KO mice received 10 days of cocaine (Coc; 20 mg/kg, ip; n = 6) or saline (Sal; n = 5). One or 14 days after the last injection, perfused vHPC slices were collected for immunohistochemistry. Representative 40× images of calreticulin (red) and GFP (green; vHPC-NAc neurons) from vHPC coronal slices. White arrows denote GFP + vHPC-NAc neurons. ( H ) Cocaine increased calreticulin intensity in GFP + neurons from WT mice at 1 day after the last injection (one-way ANOVA: F 3,1773 = 134.5; Holm-Šidák: **** P < 0.0001 for all comparisons to WT-Sal, FosB KO-Sal, and FosB KO-Coc). ( I ) Cocaine increased calreticulin intensity in GFP + neurons 14 days after the last injection both per cell (independent samples t test: t 1130 = 14.32, **** P < 0.0001) and averaged per mouse (independent samples t test: t 8 = 2.678, * P = 0.0280).

Article Snippet: Immunoprecipitation was carried out overnight at 4°C using 5 μl of rabbit polyclonal FosB antibody (lot 3; 2251S, Cell Signaling Technology) or 2 μg of rabbit polyclonal IgG isotype (3900S, Cell Signaling Technology).

Techniques: Modification, Saline, Western Blot, Binding Assay, Injection, Immunohistochemistry

Journal: The Journal of Biological Chemistry

Article Title: Discovery of small molecules and a druggable groove that regulate DNA binding and release of the AP-1 transcription factor ΔFOSB

doi: 10.1016/j.jbc.2025.111080

Figure Lengend Snippet: Effect of JPC0661 on ΔFOSB binding in dorsal hippocampus in vivo . APP mice were infused via osmotic minipumps into the dorsal hippocampus for 3 days with vehicle (“Control”) or JPC0661 (“Treatment”). Subsequently, genome-wide ΔFOSB binding was quantified with CUT&RUN sequencing performed on nuclei isolated from unilateral dorsal hippocampus tissue of individual vehicle- or JPC0661-treated mice. A , total number of ΔFOSB-bound peaks in the dorsal hippocampus of male and female APP mice receiving vehicle versus JPC0661. B , width of ΔFOSB-bound peaks from samples in ( A ). Data for ( A ) and ( B ) below are shown as mean ± SEM (n = 5 per group); ∗ p < 0.05. No sex differences were observed for ( A ) and ( B ). C , genomic distribution of ΔFOSB peaks in the dorsal hippocampus under vehicle and JPC0661 conditions, showing the percentage of peaks at the promoter, intronic, or intergenic regions. D , heatmaps showing ΔFOSB binding intensity centered around the transcription start sites (TSSs) in APP mice under vehicle ( left ) and JPC0661 ( right ) conditions, showing reduced number and amplitudes in the latter condition. E , motif analysis showing that the vast majority of ΔFOSB peaks are bound to sequences that contain AP-1 consensus sequences (TGA C/G TCA). F , number of unique genes with ΔFOSB peaks bound to gene promoter regions in APP mice treated with vehicle or JPC0661, analyzed via Venn diagrams, indicating that the predominant effect of JPC0661 is to deplete peaks seen also under vehicle-treated conditions. G , Gene Ontology (GO) enrichment analysis of genes associated with ΔFOSB promoter–binding peaks in vehicle-treated (Control_Promoter, red ) and JPC0661-treated (JPC0661_Promoter, green ) conditions. Dot size represents the significance (-log 10 of the adjusted p value), and “Odds.Ratio” indicates pathway enrichment. H , example of ΔFOSB binding at the promoter region of Creb1 . Shown are samples from the dorsal hippocampus of APP mice treated with vehicle ( top track in sky blue ) or treated with JPC0661 ( middle track in gold-green ) using an anti-ΔFOSB-antibody for CUT&RUN sequencing versus using an IgG antibody as a control for nonspecific binding ( bottom track in dark blue ; subtracted background). I , number of unique genes with ΔFOSB peaks bound to the distal intergenic regions in APP mice treated with vehicle or JPC0661, analyzed via Venn diagrams, indicating that the predominant effect of JPC0661 is to deplete peaks seen under vehicle-treated conditions. J , GO terms for the unique genes bound by ΔFOSB at intergenic regions, as described also in ( G ). K , example of ΔFOSB binding at a distal intergenic/putative enhancer region of Fos , as described in ( H ). AP-1, activator protein 1; APP, amyloid precursor protein; GO, Gene Ontology; IgG, immunoglobulin G.

Article Snippet: One subseries was stained for ΔFOSB with the avidin–biotin diaminobenzidine method, using rabbit anti-ΔFOSB (1:5000 dilution; Cell Signaling, D3S8R) as the primary antibody, biotinylated goat anti-rabbit (1:200 dilution; Vector, BA-1000) as the secondary antibody, and diaminobenzidine as the chromogen.

Techniques: Binding Assay, In Vivo, Control, Genome Wide, Sequencing, Isolation