standard mouse conditioning chamber  (Coulbourn Instruments)

Journal: Frontiers in Cellular Neuroscience

Article Title: Acute hyperexcitability differentially affects hippocampal neurogenesis features and spatial memory

doi: 10.3389/fncel.2026.1833859

Figure Lengend Snippet: Contextual fear conditioning performance. (A) Schematic diagram of the CFC paradigm showing the single-shock training session and the memory test 24 h later. (B) Learning curve across training days (D1–D3). Saline- and 25 mg/kg KA treated mice showed a progressive increase in freezing behavior, whereas 5 mg/kg KA-administered mice showed a learning delay as they displayed a similar % in freezing time as on Day 1 (* p < 0.05 vs. Sal and 25 mg/kg KA at D2; gray box indicates this time point). (C) Freezing behavior on D2. Animals from the 5 mg/kg KA group exhibited significantly lower freezing compared to the saline and 25 mg/kg groups (* p < 0.05). (D) Freezing levels during the memory test. No significant differences were observed among groups. Data are expressed as mean ± SEM. n = 16 (Sal), n = 14 (5 mg/kg KA), n = 15 (25 mg/kg KA).

Article Snippet: The test was performed in a mouse conditioning chamber (25 cm × 25 cm × 20 cm) (San Diego Instruments, San Diego, CA, USA) with transparent acrylic walls and a metal rod floor.

Techniques: Saline

Journal: iScience

Article Title: Melanocortin 4 receptor-expressing neurons in the lateral stripe of the striatum regulate affect and motor control

doi: 10.1016/j.isci.2025.112456

Figure Lengend Snippet: MC4R-LSS neurons are activated by aversive foot shocks in a D1-independent manner (A) Timeline of experiments. (B) Representative picture of GCaMP8m-expression and fiber placement in the lateral stripe of the striatum (LSS) of an MC4R-cre mouse. Fiber tract and anterior commissure (aca) marked with dashed line; LSS marked with arrow. LV, lateral ventricle. Scale bar 200 μm. (C) GCaMP8m signal measured with fiber photometry from MC4R-LSS neurons shows activation of MC4R-LSS neurons in response to aversive shocks, but not to the onset of the predictive tone cue. No change in neural activity was seen in response to shock omission in the fear test on day 2. Mean and SEM from 6 mice, 6 trials per mouse. (D) Heatmap of the data shown in (C) but split out per trial showing that the neural response is similar across all trials. Each row in the heatmap represents the mean from 6 mice. (E) Dopamine measurements from the LSS during the same behavioral paradigm as in (C). Dopamine is released in the LSS in response to both the aversive foot shocks and their predictive tone cues. No clear change in dopamine release was seen in response to shock omission in the fear test on day 2. Mean and SEM from 7 mice, 6 trials per mouse. (F) Heatmap of the data shown in (E) but split out per trial showing that the tone-induced dopamine release develops during the session. Each row in the heatmap represents the mean from 7 mice. (G) iGluSnFR-measurements of glutamate release onto D1-expressing neurons in the LSS during fear conditioning. Glutamate is released onto D1-expressing neurons in response to the aversive foot shock. A small release can be seen in response to the tone onset. Mean and SEM from 10 mice, 6 trials per mouse. (H) Heatmap of the data shown in (G), but split out per trial, showing that the shock-induced glutamate release is similar across trials and that the tone-induced release is developed within the session. Each row in the heatmap represents the mean from 10 mice. (I) Timeline of the crossover experiment testing if shock-induced activity of MC4R-LSS neurons is D1-dependent. (J) Group mean peri-event traces (± SEM) from the experiment shows a robust increase in neural activity in response to the foot shock after both saline-injection and pretreatment with the D1-antagonist SCH23390 (0.2 mg/kg; n = 7). (K) Mean peak size during the shock for each animal showing that SCH23390 does not affect the shock-induced neural response ( n = 7, paired two-tailed t-test). (L) Analysis of the inactivity time during the sessions showing that mice spend significantly more time inactive after SCH23390 injection compared to saline injection ( n = 11, paired two-tailed t-test). ∗∗∗ p < 0.001.

Article Snippet: Standard mouse operant conditioning chamber , Med Associates , ENV-307W.

Techniques: Expressing, Activation Assay, Activity Assay, Saline, Injection, Two Tailed Test

Journal: bioRxiv

Article Title: A hippocampal astrocytic sequence emerges during learning and memory

doi: 10.1101/2024.09.06.611660

Figure Lengend Snippet: (A) Astrocyte one-photon (1p) recordings in dorsal CA1 (dCA1) of the hippocampus. Wild type C57BL/6J mice were injected with 1000nl of the astrocyte-specific genetically encoded calcium indicator, AAV5-GfaABC1D-cyto-GCaMP6f-SV40, into dCA1 and allowed two weeks for recovery before implantation of a gradient-index (GRIN) lens above the pyramidal cell layer. After 10 days of post-surgical recovery, mice were imaged using the nVista System (Inscopix, Inc.). On Day 1, all mice (n=7) underwent contextual fear conditioning (CFC) for 330 seconds in Context A (Cxt A; blue throughout figure), where they received four foot shocks (0.75mA, 2s duration) at the 120, 180, 240 and 300 second time points. On Day 2, mice underwent a 330 second contextual recall session in the same mouse conditioning chamber (Cxt A) or were instead exposed to a novel Context B (Cxt B; orange throughout figure) for the same duration of time as a memory-specific control. (B) Representative 20x confocal microscopy image of the dCA1 pyramidal cell layer expression of GfaABC1D-GCaMP6f (green; astrocytes) and DAPI (blue; nuclei) with an approximate location of the GRIN lens implantation. (C) Representative 20x confocal microscopy images of the dCA1 pyramidal cell layer visualizing DAPI+ cells (blue; nuclei), GfaABC1D-GCaMP6f (green; viral expression), glial fibrillary acidic protein (GFAP) (red; astrocytes) or NeuN (red; neurons) to ensure selective expression of our virus in astrocytes. (D) Average percent (%) freezing for Day 1 (CFC) and Day 2 (recall/novel exposure) across Cxt A and B groups shows both groups display similar levels of freezing on Day 1, and Cxt A mice freeze more during Day 2, as expected. (E) Representative regions-of-interest (ROIs) map shows all astrocytes active during CFC from a mouse that were detected using manual ROI selection and curation. (F) CellReg output that shows a representative set of tracked astrocytes across Days 1-2, where green ROIs are detected across both sessions. (G) Number of astrocytes active on Days 1-2 across Cxt A and B groups indicates no significant difference in the total number of cells across days for either group. (H) Percentage of ‘reactivated’ astrocytes (# reactivated astrocytes during Day 2/total # astrocytes during Day 1) across Cxt A and B groups shows no significant difference. For 1D and 1G, Two-way RM ANOVA was performed with Sidak’s multiple comparisons test (post-hoc) where applicable. For 1H, an independent t-test was performed. Significance was indicated by: *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001. Error bars in all plots indicate mean ± SEM. All data was tested for normality and no outliers were removed. Cxt A (n = 4) and Cxt B (n = 3). See Supplemental Statistical Table for details.

Article Snippet: Cxt A was a standard mouse conditioning chamber (18.5 x 18.5 x 21.5 cm; Coulbourn Instruments) with metal-panel side walls, plexiglass front and rear walls and a stainless-steel grid-floor (16 bars) that were connected to a precision animal shocker (Actimetrics).

Techniques: Injection, Control, Confocal Microscopy, Expressing, Virus, Selection

Journal: bioRxiv

Article Title: A hippocampal astrocytic sequence emerges during learning and memory

doi: 10.1101/2024.09.06.611660

Figure Lengend Snippet: (A) Freezing percentage across contextual fear conditioning (CFC) 1-minute time bins indicate that mice in Cxt A (blue) and Cxt B (black) acquired fear at similar rates, as expected. Significant main effect of Time (Mixed-effects model (REML); [Time x Group] F(5,12) = 0.04285, p = 0.1948; [Time] F(5,18) = 7.242, p = 0.0007; [Group] F(1,5) = 0.008831, p = 0.09267). This was driven by significant differences in freezing levels across time bins in both groups (Sidak’s multiple comparisons: [Cxt A] 0 vs 300s, p = 0.0067; 60 vs. 300s, p = 0.0057; 120 vs. 300s, p = 0.0215. [Cxt B] 0 vs. 300s, p = 0.0145; 60 vs. 300s, p = 0.0148; 120 vs 300s, p = 0.0256). (B) Freezing percentages across contextual recall in 1-minute time bins indicate that mice in Cxt A froze significantly more than Cxt B, because they were re-exposed to the fearful context. Significant main effect of Group when freezing was analyzed across time within the recall session (Two-way RM ANOVA; [Time x Group] F(5,25) = 0.5339, p = 0.7485; [Time] F(5,25) = 1.478, p = 0.2324; [Group] F(1,5) = 25.76, p = 0.0039; [Subject] F(5,25) = 1.742, p = 0.1618). Post-hoc multiple comparisons revealed this effect was driven by significant differences at the 180 and 240s time bins (Sidak’s multiple comparisons [Cxt A vs. B]: 180s; p = 0.0109, 240s; p = 0.0144). Error bars in all plots indicate mean ± SEM. All data was tested for normality and no outliers were removed. Cxt A (n = 4) and Cxt B (n = 3). Significance was indicated by: *p ≤ 0.05, **p≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001.

Article Snippet: Cxt A was a standard mouse conditioning chamber (18.5 x 18.5 x 21.5 cm; Coulbourn Instruments) with metal-panel side walls, plexiglass front and rear walls and a stainless-steel grid-floor (16 bars) that were connected to a precision animal shocker (Actimetrics).

Techniques:

Journal: bioRxiv

Article Title: A hippocampal astrocytic sequence emerges during learning and memory

doi: 10.1101/2024.09.06.611660

Figure Lengend Snippet: ( A ) Representative calcium time series for Cxt A (blue) and Cxt B (black) showing almost all astrocytes display a robust response to one or more shocks during the contextual fear conditioning (CFC) session. ( B ) Peri-event analysis for astrocytes in Cxt A (blue) and Cxt B (orange) show increased activity on average (z-scored % dF/F) at the time of each 0.75 mA foot shock (dashed line; 120, 180, 240, 300s). Shock response is averaged for each group with Cxt A (n = 4) and Cxt B (n = 3). Here, we used a 95% t-confidence interval (tCI) method where a significant event is indicated by the colored bars above the time series. These intervals indicate any time >1.0 seconds that did not include the mean-shifted baseline of 0. ( C ) Representative cosine-similarity plot of each pair of astrocytes in time for an animal (Astro4). Here, we observe block-like structured activity after the onset of the first foot shock across all mice during CFC. Dashed white lines indicate the times of administered foot shocks (120, 180, 240, and 300s). ( D ) Representative heatmap of all astrocytic calcium activity for an animal (Astro4) (z-scored % dF/F) that was averaged across shocks #1 and 3 (Odds) and sorted by their argmax during CFC. Time = 0 indicates the onset of each foot shock. To assess the stability of the sequential response, the same activity for Odds was sorted based on the argmax of the average response of each astrocyte to shocks #2 and 4 (Evens). Here, we observe a stable sequence across all animals across foot shocks (n = 7). ( E ) Spearman’s correlation between the true peak times (Odd sorted on Odd) and the cross-validated peak times (Odd sorted on Even) shows a significant difference between the average rho value and the hypothesized value of zero (n = 7) (One-sample t-test: t=13.19, p<0.0001). ( F ) Representative linear regression of the true peak times (i.e., Odd Sort on Odd Peaks against the cross-validated peak times (i.e., the Odd Peaks sorted on the Even Peak times) for an animal (Astro4) (slope=0.256; intercept=2.71; p_value_slope=0.00017; p_value_bias=3.73E-21; R =0.065). ( G ) Density histogram and kernel density estimation plot of predicted peak times (µ) at the population level (n = 7) show a skewed distribution after the onset of foot shock on average. ( H ) Plots of the empirical cumulative distribution function and the theoretical cumulative distribution functions (CDFs) of the fit models (i.e., Power-Law (purple; WAIC= 7879), Exponential (red; WAIC=8837), Gaussian (teal; WAIC=9854), Lognormal (green; WAIC=8015), Actual Data (grey). Power-Law is the best fit for our data, indicating temporally compressed structure. Kolmogorov-Smirnov (K-S) test for Power-Law shows that even the best fitting model does not perfectly capture the peak distribution (KS distance = 0.46, p<0.001). ( I ) Scatter plot of the within-trial time field width (σ) as a function of the peak location (µ) for each astrocyte from all mice. Here, the width of a calcium event (individual data points shown) increases linearly with time after foot shock (peak location) as determined by a linear mixed effects model. The solid line indicates the fixed effects best fit line that predicts Y from X with the associated 95% CI are shown on the plot (slope = 0.364, p<0.001). Significance was indicated by: *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001. All data was tested for normality and no outliers were removed. All mice (n = 7). See Supplemental Statistical Table for details.

Article Snippet: Cxt A was a standard mouse conditioning chamber (18.5 x 18.5 x 21.5 cm; Coulbourn Instruments) with metal-panel side walls, plexiglass front and rear walls and a stainless-steel grid-floor (16 bars) that were connected to a precision animal shocker (Actimetrics).

Techniques: Activity Assay, Blocking Assay, Sequencing

Journal: bioRxiv

Article Title: A hippocampal astrocytic sequence emerges during learning and memory

doi: 10.1101/2024.09.06.611660

Figure Lengend Snippet: (A) Cosine similarity plots for all animals (astro3-9) for contextual fear conditioning, showing the structured temporal dynamics induced by repeated foot shocks. Dashed lines indicate the times of administered foot shocks (120, 180. 240. 300 seconds). (B) Cosine similarity plots for all animals (astro3-9) for recall, showing the structured temporal dynamics maintained in Cxt A (astro3, astro4, astro5, astro9), but not Cxt B (astro6, astro8, astro8) mice. Animal label colors (e.g. astro3) indicate Cxt A (blue) or Cxt B (black) groups. Dashed lines indicate the times of spontaneously detected events for each animal.

Article Snippet: Cxt A was a standard mouse conditioning chamber (18.5 x 18.5 x 21.5 cm; Coulbourn Instruments) with metal-panel side walls, plexiglass front and rear walls and a stainless-steel grid-floor (16 bars) that were connected to a precision animal shocker (Actimetrics).

Techniques:

Journal: bioRxiv

Article Title: A hippocampal astrocytic sequence emerges during learning and memory

doi: 10.1101/2024.09.06.611660

Figure Lengend Snippet: (A) Detected shock sequences for all individual animals (astro3-9) during contextual fear conditioning as a confirmation of the ‘sequence detector’ described in . Individual calcium activity was summed over each time index to generate the summed activity trace for a given mouse (solid line; Cxt A = blue, Cxt B = black). Temporal thresholding was performed by finding the time indices where the summed activity was > 1.5 standard deviations above the mean and lasted for > 1 second. These time periods qualified as detected ‘sequences’ (grey shading). (B) Using the same method above, detected spontaneous sequences for all individual animals (astro3-9) during recall.

Article Snippet: Cxt A was a standard mouse conditioning chamber (18.5 x 18.5 x 21.5 cm; Coulbourn Instruments) with metal-panel side walls, plexiglass front and rear walls and a stainless-steel grid-floor (16 bars) that were connected to a precision animal shocker (Actimetrics).

Techniques: Sequencing, Activity Assay

Journal: bioRxiv

Article Title: A hippocampal astrocytic sequence emerges during learning and memory

doi: 10.1101/2024.09.06.611660

Figure Lengend Snippet: (A) Representative summed activity trace for an animal in Cxt A during recall showing an inverse relationship between detected epochs of high activity (grey) and freezing periods (black). (B) Quantification of the average percent freezing in Cxt A animals (n = 4) during high activity ‘sequence’ periods compared to ‘non-sequence’ periods. Cxt B mice were not included in this analysis as they did not freeze during recall ( see ; Supplemental Figure 1B). During sequence periods, Cxt A animals froze significantly less than when outside a detected sequence (Paired t-test: t=-3.25, p=0.0473). (C) Spearman’s Rho values for each group during recall. Correlations were calculated for the ‘reactivated’ cells (e.g. astrocytes that were present across days), between their peak time on average during fear conditioning (FC) at the times of foot shock (FC Sort on FC; not shown, see Supplemental Figure) and the cross-validated peak times on average at the time of spontaneously detected high activity events (Recall Sort on FC). Points represent individual animals (Independent t-test: t=6.37, p=0.003107). (D) Representative heatmaps of calcium activity from ‘reactivated’ astrocytes in Cxt A (left; Astro5) and Cxt B (right; Astro7), sorted by their ordering in the foot shock sequence during FC (Recall Sort on FC). Significance was indicated by: *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001. All data was tested for normality and one animal (Astro9) was removed from the Cxt A group due to having a low number of reactivated cells ( 4B-C ; data from this animal is shown in Supplemental Figure ). Cxt A (n = 4) and Cxt B (n = 3).

Article Snippet: Cxt A was a standard mouse conditioning chamber (18.5 x 18.5 x 21.5 cm; Coulbourn Instruments) with metal-panel side walls, plexiglass front and rear walls and a stainless-steel grid-floor (16 bars) that were connected to a precision animal shocker (Actimetrics).

Techniques: Activity Assay, Sequencing

Journal: bioRxiv

Article Title: A hippocampal astrocytic sequence emerges during learning and memory

doi: 10.1101/2024.09.06.611660

Figure Lengend Snippet: A-G) Left Panel: Heatmap of cells in fear conditioning that were also present during recall sorted on the argmax of the peaks during shocks in fear conditioning. Right Panel: Heat map of the same reactivated cells averaged over spontaneously detected events, but, during re-exposure to Cxt A or exposure to Cxt B. Heatmap is sorted on the argmax values from FC. Animals who were re-exposed to Cxt A (e.g., sub panels A-D) show more sequence stability compared to animals exposed to Cxt B (e.g., E-G). Astro9 was removed from statistical analysis as it only had 6 ‘reactivated’ cells.

Article Snippet: Cxt A was a standard mouse conditioning chamber (18.5 x 18.5 x 21.5 cm; Coulbourn Instruments) with metal-panel side walls, plexiglass front and rear walls and a stainless-steel grid-floor (16 bars) that were connected to a precision animal shocker (Actimetrics).

Techniques: Sequencing

Journal: eLife

Article Title: Chronic activation of a negative engram induces behavioral and cellular abnormalities

doi: 10.7554/eLife.96281

Figure Lengend Snippet: ( a ) C57BL/6J mice were bilaterally injected with a 1:1 viral cocktail of AAV9-c-Fos-tTA and AAV9-TRE-eYFP into the vHPC. This activity-dependent labeling strategy captures sufficiently active neurons expressing the immediate-early gene, cfos . This system couples the cfos promoter to the expression of the tetracycline transactivator (tTA), which binds to the tetracycline response element (TRE) in its protein form. When doxycycline (DOX) is present, such as in the animal’s diet, this inhibits the binding of tTA to TRE, preventing eYFP labeling of active cells. ( b ) Experimental schematic used to label, isolate, and analyze the two groups of vHPC cells labeled by eYFP upon shock (negative) or male-to-female interaction (positive). Prior to surgery, mice were placed on a Dox diet to inhibit the labeling of active neurons. On day 0, mice were injected with the activity-dependent viral strategy to enable labeling of active neurons during a salient experience. On day 10 after viral expression and recovery, the mice were taken off of Dox prior to engram tagging to open the labeling window. On day 11, mice were subjected to contextual fear conditioning (CFC) or male-to-female interaction to label negative or positive neurons, respectively. Mice were placed immediately back on Dox to close this tagging window. Then, 24 hr later, mice were sacrificed and brains were obtained for sequencing experiments. ( c ) List of gene abbreviations and full names used for the GSEA of anti-inflammatory genes. ( d ) GSEA with the enrichment score for negative vs. untagged cells and positive vs. untagged cells using the anti-inflammatory gene set. ( e ) List of gene abbreviations and names used for the GSEA of pro-inflammatory genes. ( f ) GSEA with the enrichment score of negative vs. untagged cells and positive vs. untagged cells using the pro-inflammatory gene set.

Article Snippet: For CFC, all time points took place in mouse conditioning chambers (Coulbourn Instruments, Holliston, MA).

Techniques: Injection, Activity Assay, Labeling, Expressing, Binding Assay, Sequencing

Journal: eLife

Article Title: Chronic activation of a negative engram induces behavioral and cellular abnormalities

doi: 10.7554/eLife.96281

Figure Lengend Snippet: ( a ) Schematic representation of tagging a negative engram in young (3 months) and old (11 months) TRAP2 mice followed by 3 months of chronic engram stimulation and a battery of behavioral tests. AAV9-hSyn-DIO-hM3Dq-mCherry or AVV9-hSyn-DIO-mCherry control vector was bilaterally injected into the ventral hippocampus (vHPC) of young and old mice. After viral expression and surgical recovery, all mice were subjected to contextual fear conditioning (CFC) in context A and subsequent 4-hydroxytamoxifen (4-OHT) intraperitoneal (IP) injection to induce negative engram tagging. For the next 3 months, mice received the water-soluble Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) agonist deschloroclozapine dihydrochloride (DCZ) in their home cage water. After 3 months of stimulation wherein the young and old groups reached 6 months and 14 months of age, the mice underwent open field, y-maze, zero maze, remote recall, extinction, and generalization in a novel context B. ( b ) Representative image of hSyn-DIO-hM3Dq-mCherry expression in the vHPC (red) and DAPI+ cells (blue) after 3 months of hM3Dq activation. ( c ) Weights of all groups were recorded over the 3-month stimulation protocol, once per month. (Left) Young 6-month-old and (right) older 14-month-old mice. Values are given as a mean + SEM. Statistical analysis was performed with two-way (RM) ANOVA followed by Sidak’s post hoc test. n = 9–17 per group after outlier removal. p≤0.05, **p≤0.01, ***p≤0.001, ****p≤0.0001, no label = not significant.

Article Snippet: For CFC, all time points took place in mouse conditioning chambers (Coulbourn Instruments, Holliston, MA).

Techniques: Battery, Control, Plasmid Preparation, Injection, Expressing, Activation Assay

Journal: eLife

Article Title: Chronic activation of a negative engram induces behavioral and cellular abnormalities

doi: 10.7554/eLife.96281

Figure Lengend Snippet: ( a ) Schematic of contextual fear conditioning (CFC) in context A wherein 3- and 11-month-old mice received four, 1.5 mA, 2 s foot shocks. Following the CFC session, the negative engram was tagged in all mice by intraperitoneal (IP) injection of 4-hydroxytamoxifen (4-OHT). ( i ) Average percent freezing levels and total percentage freezing across the 300 s CFC session for ( ii ) 3-month groups and ( iii ) 11-month groups (two-way ANOVA RM; [3 months] interaction: F(5,130) = 1.131, p=0.3473; time: F(5,130) = 115.6, p<0.0001; group: F(1,26) = 2.737, p=0.1101; subject: F(26,130) = 3.056, p<0.0001. [11 months] Interaction: F(5,90) = 1.218, p=0.3074; time: F(5,90) = 234.4, p<0.0001; group: F(1,18) = 1.862, p=0.1892; subject: F(18,90) = 2.887, p=0.0005). ( b ) Schematic of remote recall performed after 3 months of negative engram stimulation. Mice were returned to context A for 5 min in the absence of foot shocks. ( i ) Average percent freezing levels and total percentage freezing across the 300 s remote recall session for ( ii ) 6-month groups and ( iii ) 14-month groups two-way ANOVA RM; [6 months] interaction: F(5,85) = 15.72, p=0.0115; time: F(5,85) = 15.72, p<0.001; group: F(1,17) = 6.588, p=0.0200; subject: F(17,85) = 2.458, p=0.0035. [14 months] Interaction: F(5,130) = 0.9027, p=0.4815; time: F(5,130) = 14.18, p<0.0001; group: F(1,26) = 0.3967, p=0.5343; subject: F(26,130) = 4.107, p<0.0001. ( c ) Schematic of extinction wherein mice were placed in context A for 30 min in the absence of foot shocks. ( i ) Average percent freezing levels and total percentage freezing across the 1800s extinction session for ( ii ) 6-month groups and ( iii ) 14-month groups (two-way ANOVA RM; [6 months] interaction: F(5,85) = 0.4133, p=0.8383; time: F(5,85) = 1.979, p=0.0899; group: F(1,17) = 5.494, p=0.0315; subject: F(17,85) = 7.352, p<0.0001. [14 months] interaction: F(5,130) = 1,362, p=0.2570; time: F(5,130) = 5.387, p=0.0002; group: F(1,26) = 0.3836, p=0.5411; subject: F(26, 130)=11.16, p<0.0001). ( d ) Schematic of the novel context B used to assess generalization. ( i ) Average percent freezing levels and total percentage freezing across the 300 s generalization session for ( ii ) 6-month groups and ( iii ) 14-month groups (two-way ANOVA RM; [6 months] interaction: F(4,72) = 2.257, p=0.0713; time: F(4,72) = 3.373, p=0.0138; group: F(1,18) = 4.151, p=0.0566; subject: F(18,72) = 2.537, p=0.0028. [14 months] interaction: F(4,100) = 2.575, p=0.0421; time: F(4,100) = 5.717, p=0.0003; group: F(1,25) = 6.343, p=0.0186; subject: F(25,100) = 3.175, p<0.0001). Statistical analysis utilized a two-way ANOVA with time point and group as factors and two-way repeated measures (RM) ANOVA with time (seconds) and group as factors across 6- and 14-month-old mice. Tukey’s or Sidak’s post hoc tests were performed where applicable. Error bars indicate SEM. p≤0.05, **p≤0.01, ***p≤0.001, ****p≤0.0001, ns = not significant. n = 8–11 per group after outlier removal.

Article Snippet: For CFC, all time points took place in mouse conditioning chambers (Coulbourn Instruments, Holliston, MA).

Techniques: Injection