Article Figures & Data
Figures
- Figure 1.
Induction of ΔFosB in the hippocampus by novelty exposure and spatial learning. A, Quantitation and representative Western blots (inset) of ΔFosB in the hippocampus show significant increases after novel environmental exposure (red; n = 9) compared with mice left in their home cage (white; n = 8; *p < 0.05). B, Representative paths of rats seeking a visible (cued) or hidden (spatial) platform in the MWM test. C, Escape latency from the MWM in rats (left) and mice (right) using a hidden platform significantly decreased across training days (n = 6–8; *p < 0.05 compared to day 1). D, Representative 40× images of coronal sections stained for FosB gene products from naive, cued learning, and spatial learning rats (left) and mice (right). Red arrowheads indicate FosB+ pyramidal cells in CA1.
- Figure 2.
Viral-mediated gene transfer in the hippocampus. A, Representative fluorescent coronal images of the hippocampus (approximately −2.0 mm AP) 6 d after injection of HSV–GFP or 28 d after injection of AAV–GFP. B, Coronal figures (−1.94 and −2.46 AP; Paxinos and Franklin, 2012) showing representative center fluorescence distribution from GFP (green circles), GFP–ΔJunD (orange circles), or GFP–ΔFosB (blue circles) infusions. Immunofluorescence of CA1 neurons close to the injection site of HSV–GFP and HSV–GFP–ΔFosB (C) or HSV–GFP–ΔJunD (D) reveals specific transgene expression only in cells induced with virus.
- Figure 3.
Transcriptional silencing of hippocampal ΔFosB impairs learning and memory. A, NOR in mice that received stereotaxic hippocampal injections of AAV–GFP (GFP, green; n = 8) and AAV–GFP–ΔJunD (ΔJunD, orange; n = 8). The duration of time spent with a familiar object (blank bars) and a novel object (crosshatched bars) was measured. GFP mice spent significantly more time with the novel object than the familiar object (*p < 0.05 compared with familiar), whereas the ΔJunD mice spent an equal amount of time with both objects. B, Contextually fear conditioned (Context FC) freezing (seconds) in AAV–GFP (n = 8) and AAV–GFP–ΔJunD (n = 8) mice. ΔJunD significantly decreased contextual freezing (*p < 0.05 compared with GFP). C, TDPA performance was measured in mice that received AAV–GFP (n = 16) or AAV–GFP–ΔJunD (n = 16). Latency (seconds) to crossover to the shock-paired dark side of the box was measured. ΔJunD mice had significantly lower crossover latency at days 4–5 (**p < 0.01, ***p < 0.001, respectively, compared with GFP). D, Spatial learning in the MWM was measured after AAV–GFP (n = 8) or AAV–GFP–ΔJunD (n = 8). Escape latency (seconds) did not differ between groups across training days. E, The probe test revealed that ΔJunD mice spent significantly less time in the target quadrant (*p < 0.05 compared with GFP). F, Mice received stereotaxic hippocampal infusions of HSV–GFP (n = 16) or HSV–GFP–ΔJunD (n = 15). GFP mice spent significantly more time with the novel object than the familiar object (*p < 0.001 compared with familiar), whereas the ΔJunD mice spent an equal amount of time with both objects. G, Context FC freezing after HSV–GFP (n = 8) and HSV–GFP–ΔJunD (n = 8). ΔJunD mice spent significantly less time freezing (*p < 0.05 compared with HSV–GFP). Error bars indicate mean ± SEM.
- Figure 4.
Overexpression of ΔFosB in the hippocampus impairs learning and memory. A, NOR in mice that received stereotaxic hippocampal injections of AAV–GFP (GFP, green; n = 14) or AAV–GFP–ΔFosB (ΔFosB, blue; n = 16). GFP mice spent significantly more time with the novel object than the familiar object (*p < 0.05 compared with familiar), whereas the ΔFosB mice spent an equal amount of time with both objects. B, Contextually fear conditioned (Context FC) freezing (seconds) in AAV–GFP (n = 8) and AAV–GFP–ΔFosB (n = 8) mice. ΔFosB mice had significantly greater contextual freezing (*p < 0.05 compared with GFP). C, TDPA performance was measured in mice that received AAV–GFP (n = 15) or AAV–GFP–ΔFosB (n = 16). ΔFosB mice had significantly lower crossover latency at days 4–5 (*p < 0.05, **p < 0.01, respectively, compared with GFP). D, Spatial learning in the MWM was measured after AAV–GFP (n = 7) or AAV–GFP–ΔFosB (n = 7). Escape latency (seconds) was significantly decreased by ΔFosB at days 3–4 (**p < 0.01 for both days compared with GFP). E, Probe test quadrant time revealed that ΔFosB mice spent significantly less time in the target quadrant (*p < 0.05 compared with GFP). Error bars indicate mean ± SEM.
- Figure 5.
Overexpression, but not transcriptional silencing, of ΔFosB in hippocampus produces anxiety-like behavior. A, OF behavior was assessed in AAV–GFP (green; n = 16), AAV–GFP–ΔJunD (orange; n = 8), and AAV–GFP–ΔFosB (blue, n = 8) mice. Center zone time (percentage) was reduced significantly by ΔFosB (*p < 0.001 compared with GFP), but no differences were observed between GFP and ΔJunD. B, Center zone entries were not different between groups. C, Locomotor activity in an OF, measured by distance moved (meters), was not different between groups. D, Open arm time (percentage) in an EPM was decreased in mice that received ΔFosB (*p < 0.05 compared with GFP). E, Open arm entries were also reduced in mice that received ΔFosB (**p < 0.01 compared with GFP). F, Distance moved (meters) in the EPM was not different between groups. G, Preference for sucrose (percentage) increased in all groups compared with a sucrose-free baseline (***p < 0.001 compared with baseline; B). No group differences were observed in sucrose preference. H, Body weight (grams) was measured across weeks after AAV surgery (n = 15–16 per group). Although mice steadily increased total body weight, no differences were observed between groups (dotted lines represent 95% confidence interval). Error bars indicate mean ± SEM.
- Figure 6.
ΔFosB regulates hippocampal spine formation. Mice received hippocampal infusions of HSV–GFP, HSV–GFP–ΔFosB, or HSV–GFP–ΔJunD and dendritic spine analysis was conducted on hippocampal CA1 pyramidal neurons. A, Representative dendrites showing spines from GFP+ hippocampal CA1 neurons. B, Quantitation of spines in CA1 neurons shows ΔFosB (blue; n = 30) had significantly more total spines, thin spines, and stubby spines compared with GFP alone (gray; n = 30; *p < 0.001), with no significant difference in mushroom spines. ΔJunD (red; n = 19) had significantly fewer total spines, thin spines, and mushroom spines compared with GFP alone (gray; n = 22; *p < 0.001), with no significant difference in stubby spines. Error bars indicate mean ± SEM. C, Model depicting experience-dependent changes in hippocampal cells under control conditions or when virally transduced with ΔJunD (red) or ΔFosB (blue). Green indicates changes in synaptic structure/function resulting from ΔFosB expression within a circuit that may underlie learning.
Tables
Brain region Control cells/mm2 (mean ± SEM) (n) Cued cells/mm2 (mean ± SEM) (n) Spatial cells/mm2 (mean ± SEM) (n) F (one-way ANOVA) p (one-way ANOVA) Rats CA1 2.10 ± 0.66 (5) 2.88 ± 0.43 (4) 5.10 ± 1.00 (5)* F(2,11) = 4.316 0.0413 CA3 3.50 ± 0.67 (5) 6.00 ± 1.02 (4) 4.90 ± 0.91 (5) F(2,11) = 2.038 0.176 DG 17.70 ± 0.75 (5) 30.88 ± 4.04 (4)* 40.10 ± 4.05 (5)* F(2,11) = 13.02 0.0013 NAc 58.17 ± 7.15 (6) 93.42 ± 4.17 (6)* 89.83 ± 7.95 (6)* F(2,15) = 8.582 0.0033 Mice CA1 69.15 ± 11.0 (8) 92.64 ± 12.7 (8) 143.9 ± 19.5 (7)* F(2,20) = 6.752 0.0057 CA3 533.7 ± 60.2 (8) 844.9 ± 70.9 (8)* 736.1 ± 56.3 (7) F(2,20) = 6.448 0.0069 DG 1222 ± 67.4 (8) 1893 ± 83.8 (8)* 1586 ± 131.9 (7)* F(2,20) = 13.08 0.0002 FosB+ cells as visualized by DAB staining (Fig. 1) in control, cued learning, and spatial learning mice and rats by brain region. Statistics represent one-way ANOVA with Tukey's multiple comparison post hoc test used to compare conditions.
↵*p < 0.05, significant difference from the control group.
