Research ArticleNEUROPSYCHOLOGY

Social transfer of pain in mice

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Science Advances  19 Oct 2016:
Vol. 2, no. 10, e1600855
DOI: 10.1126/sciadv.1600855
  • Fig. 1 Social transfer of CFA and morphine withdrawal–induced pain.

    (A) Experimental timeline of experiments presented in (B) and (C). Von Frey (VF, thick orange arrows) injections of morphine/CFA (Mor/CFA, black syringes) or naloxone (NLX, green syringe). (B) Mice subjected to intraplantar CFA injection showed a robust and persistent decrease in mechanical sensitivity for all test sessions (CFA/Co-Housed; n = 8) compared to vehicle-injected mice housed in a separate room (PBS/Separate; n = 8). Vehicle-injected mice housed in the same room as CFA-injected mice (Veh/Co-Housed; n = 8) demonstrated significantly decreased mechanical thresholds compared to Veh/Separate mice during the last three test sessions. This resulted in significant differences between groups (F2,21 = 30.0, P < 0.0001) across time (F4,84 = 27.6, P < 0.0001) and a significant interaction between these variables (F8,84 = 9.1, P = 0.003) according to repeated-measures analysis of variance (ANOVA). (C) Co-Housed mice injected with either a slow-release morphine emulsion (Mor/Co-Housed/WD; n = 7) or vehicle emulsion (Veh/Co-Housed; n = 8) every other day demonstrated significant decreases in mechanical thresholds on the two test sessions compared to vehicle-injected mice housed in a separate room (Veh/Separate; n = 7). Repeated-measures ANOVA showed a significant effect of treatment (F2,19 = 7.4, P = 0.004) and a significant effect of time (F2,38 = 5.7, P = 0.006). Following a significant interaction, Bonferroni’s post hoc analyses were conducted. Differences compared to control are represented by *, and differences compared to baseline are represented by #. Mean basal responses of all groups are represented by dotted lines.

  • Fig. 2 Social transfer of alcohol withdrawal–induced mechanical sensitivity to nearby water-drinking controls.

    (A) Experimental timeline of experiments presented in (B) to (E). Von Frey (thick orange arrows); tail immersion (TI, small maroon arrows); ethanol [EtOH, 3 to 10% (v/v)]. h, hours. (B) Ethanol-drinking mice (EtOH/Co-Housed/WD; n = 14 males per group) demonstrate a significant decrease in mechanical thresholds following one withdrawal session that is matched by water-drinking control mice housed in the same room (H2O/Co-Housed; n = 10 males) by the second withdrawal session. Ethanol-drinking control mice housed in an adjacent room (EtOH/Separate/WD; n = 12 males) also demonstrate enhanced mechanical sensitivity between 1 and 3 withdrawal sessions. Water-drinking mice in an adjacent room (H2O/Separate; n = 14 males) display stable mechanical thresholds across the time course. Repeated-measures ANOVA that compared mechanical sensitivity of male mice over time revealed significant main effects of week (F3,138 = 26.16, P < 0.0001), treatment (F3,46 = 6.69, P = 0.0008), and a significant interaction (F9,138 = 4.97, P < 0.0001). Bonferroni’s post hoc analysis revealed significant differences between H2O/Separate and H2O/Co-Housed, EtOH/Co-Housed/WD, and EtOH/Separate/WD. (C) In a separate experiment that used female mice (n = 7 to 8 per group), H2O/Separate mice (n = 8) never significantly deviated from baseline. Both Co-Housed groups demonstrated decreased mechanical thresholds during the first and second withdrawal sessions, with the bystander group (H2O/Co-Housed; n = 7) reaching the lowest level. Repeated-measures ANOVA demonstrated significant main effects of treatment (F2,19 = 13.0, P = 0.0003), week (F2,38 = 7.1, P < 0.002), and a significant interaction (F4,38 = 4.4, P < 0.005). Bonferroni’s post hoc analysis revealed significant differences between H2O/Separate and H2O/Co-Housed and EtOH/Co-Housed/WD. (D) When tested for thermal sensitivity by immersing the tail into a hot water bath, Co-Housed EtOH mice (n = 8) and H2O mice (n = 8) demonstrate significantly shorter withdrawal latencies on the second withdrawal session compared to H2O/Separate mice according to one-way ANOVA on the second withdrawal session (F2,21 = 9.8, P = 0.001). (E) Ethanol-drinking mice with continuous access/no withdrawal sessions (EtOH/Co-Housed/NoWD; n = 7) and H2O mice housed in the same room (H2O/Co-Housed/NoWD; n = 7) did not demonstrate any alterations in mechanical sensitivity following 2 weeks of ethanol exposure. There were no significant differences between groups according to repeated-measures ANOVA (P > 0.05). Significant changes (P < 0.05) from baseline according to Bonferroni’s post hoc analyses are represented by #. Significant differences compared to control (P < 0.05) are represented by *. Mean basal responses of all groups are represented by a dotted line.

  • Fig. 3 Social transfer occurs via alcohol withdrawal–specific olfactory cues, and this state leads to chemical and thermal hyperalgesia.

    (A) Experimental timeline for (B) to (G). Von Frey (thick orange arrows); ethanol [EtOH, 3 to 10% (v/v)]. (B) When a group of mice housed in a separate room (H2O/Olfactory-WD; n = 8) was exposed to bedding from the cages of H2O/Co-Housed mice (n = 9) and EtOH/Co-Housed/WD mice (n = 8), they demonstrated significant decreases in mechanical thresholds within 24 hours. Mice exposed to bedding from naïve water-drinking mice maintained baseline levels of sensitivity (H2O/Olfactory-CTRL; n = 16). H2O/Co-Housed and EtOH/Co-Housed/WD mice began the experiment 1 day before H2O/Olfactory-WD mice, and transfer of bedding is represented by thin blue arrows. Repeated-measures ANOVA revealed a significant effect of treatment (F3,37 = 7.3, P = 0.0006) and test session (F2,74 = 26.7, P < 0.0001), as well as a significant interaction (F6,74 = 3.3, P = 0.0068). (C) The mechanical hypersensitivity in groups of mice from the olfactory experiment (H2O/Co-Housed, EtOH/Co-Housed, and H2O/Olfactory-WD) and the no withdrawal experiment (Fig. 1D) manifests as hyperalgesia following a low concentration (1.5%) of formalin (black syringe) in a pattern that was significant during the second phase of the formalin test according to one-way ANOVA (F4,30 = 10.19, P <.0001). (D) There were no significant differences in the percent of time spent on closed or open arms for any group (H2O/Separate, n = 9; EtOH/Co-Housed, n = 9; and H2O/Co-Housed, n = 9) according to ANOVA (P > 0.05). (E) H2O/Co-Housed mice (n = 14) and EtOH/Co-Housed/WD mice (n = 14) were treated with diazepam (Diaz; 1.0 mg/kg; maroon syringe; n = 7) or vehicle (Veh; n = 7) 20 min before the second von Frey test. Diazepam had no effect on mechanical thresholds in any group, according to ANOVA (P > 0.05). (F) H2O/Co-Housed and EtOH/Co-Housed/WD were treated with metyrapone (Met; 50.0 mg/kg; maroon syringe) or vehicle (Veh) 20 min before the second von Frey test. Metyrapone had no effect on mechanical thresholds in any group (EtOH/Co-Housed, n = 5; H2O/Co-Housed, n = 7) compared to vehicle (EtOH/Co-Housed, n = 4; H2O /Co-Housed, n = 8), according to ANOVA (P > 0.05). (G) Acoustic startle responses did not differ between Co-Housed (n = 8/group) and Separate (n = 8) mice according to repeated-measures ANOVA (P > 0.05). Significant changes (P < 0.05) from baseline according to Bonferroni’s post hoc analyses are represented by #. Significant differences compared to control (P < 0.05) are represented by *. Nonsignificant differences are represented by NS. Mean basal responses of all groups are represented by a dotted line.

  • Table 1 No changes between groups in plasma CORT levels at several time points.

    When examining plasma CORT (taken immediately postmortem) in separate groups of mice, there were no changes in the mean (±SEM) plasma CORT levels (P > 0.05) between groups (n = 5 to 12) following 1 week of drinking and one withdrawal session (WD 1), 3 weeks of drinking and three withdrawal sessions (WD 3), following restored access to EtOH during the fourth withdrawal session or after 4 weeks of drinking and four withdrawal sessions followed by 7 days of extended withdrawal (xtend), or following 30 min of restraint stress on the eighth day after recovery from hyperalgesia.

    Time of sacrificeH2O/Co-HousedEtOH/Co-Housed/WDEtOH/SeparateH2O/Separate
    WD 1204.4 ± 15.63192.9 ± 24.91
    WD 3279.3 ± 44.57310.7 ± 63.68337.4 ± 37.2381.3 ± 44.33
    WD 4/restored202.5 ± 14.07178 ± 25.67
    xtend WD288.4 ± 22.4319.8 ± 45.5271.5 ± 30.39314.8 ± 41.06
    After restraint629.4 ± 67.39615.9 ± 55.47569.3 ± 46.65

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/2/10/e1600855/DC1

    fig. S1. Naloxone precipitates withdrawal behaviors in morphine-treated mice.

    fig S2. Recovery of mechanical thresholds returns to baseline after 4 days of extended withdrawal.

    fig S3. Stranger mice develop socially transferred hypersensitivity.

    fig S4. Nonsynchrony of nocifensive behavior in primary and bystander mice.

    fig S5. Alcohol access reverses mechanical hypersensitivity in EtOH-withdrawn mice.

    fig. S6. No differences in behavior on EPM.

    fig. S7. Diazepam attenuates HICs following acute EtOH withdrawal.

    fig S8. Four to 5 weeks of isolation/individual housing leads to mechanical hypersensitivity.

    table S1. Average mechanical thresholds and alcohol intake.

  • Supplementary Materials

    This PDF file includes:

    • fig. S1. Naloxone precipitates withdrawal behaviors in morphine-treated mice.
    • fig. S2. Recovery of mechanical thresholds returns to baseline after 4 days of extended withdrawal.
    • fig. S3. Stranger mice develop socially transferred hypersensitivity.
    • fig. S4. Nonsynchrony of nocifensive behavior in primary and bystander mice.
    • fig. S5. Alcohol access reverses mechanical hypersensitivity in EtOH-withdrawn mice.
    • fig. S6. No differences in behavior on EPM.
    • fig. S7. Diazepam attenuates HICs following acute EtOH withdrawal.
    • fig. S8. Four to 5 weeks of isolation/individual housing leads to mechanical hypersensitivity.
    • table S1. Average mechanical thresholds and alcohol intake.

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