Social transfer of pain in mice

Abstract

A complex relationship exists between the psychosocial environment and the perception and experience of pain, and the mechanisms of the social communication of pain have yet to be elucidated. The present study examined the social communication of pain and demonstrates that "bystander" mice housed and tested in the same room as mice subjected to inflammatory pain or withdrawal from morphine or alcohol develop corresponding hyperalgesia. Olfactory cues mediate the transfer of hyperalgesia to the bystander mice, which can be measured using mechanical, thermal, and chemical tests. Hyperalgesia in bystanders does not co-occur with anxiety or changes in corticosterone and cannot be explained by visually dependent emotional contagion or stress-induced hyperalgesia. These experiments reveal the multifaceted relationship between the social environment and pain behavior and support the use of mice as a model system for investigating these factors. In addition, these experiments highlight the need for proper consideration of how experimental animals are housed and tested.

Keywords: Chronic pain; alcohol withdrawal; empathy; olfactory communication; social environment.

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 (F_2,21 = 30.0, P < 0.0001) across time (F_4,84 = 27.6, P < 0.0001) and a significant interaction between these variables (F_8,84 = 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 (F_2,19 = 7.4, P = 0.004) and a significant effect of time (F_2,38 = 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 (F_3,138 = 26.16, P < 0.0001), treatment (F_3,46 = 6.69, P = 0.0008), and a significant interaction (F_9,138 = 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 (F_2,19 = 13.0, P = 0.0003), week (F_2,38 = 7.1, P < 0.002), and a significant interaction (F_4,38 = 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 (F_2,21 = 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 (F_3,37 = 7.3, P = 0.0006) and test session (F_2,74 = 26.7, P < 0.0001), as well as a significant interaction (F_6,74 = 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 (F_4,30 = 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.