Anterior cingulate inputs to nucleus accumbens control the social transfer of pain and analgesia

Abstract

Empathy is an essential component of social communication that involves experiencing others' sensory and emotional states. We observed that a brief social interaction with a mouse experiencing pain or morphine analgesia resulted in the transfer of these experiences to its social partner. Optogenetic manipulations demonstrated that the anterior cingulate cortex (ACC) and its projections to the nucleus accumbens (NAc) were selectively involved in the social transfer of both pain and analgesia. By contrast, the ACC→NAc circuit was not necessary for the social transfer of fear, which instead depended on ACC projections to the basolateral amygdala. These findings reveal that the ACC, a brain area strongly implicated in human empathic responses, mediates distinct forms of empathy in mice by influencing different downstream targets.

Fig. 1.. Rapid social transfer of pain to bystander mice.

(A) Timeline of social transfer of pain protocol. (B) Timecourse of mechanical von Frey (VF) sensitivity at 0, 4, 24 and 48 h post 1h social interaction in Con/Con and CFA/BY pairs. (C) Mechanical thresholds immediately after the 1 h social interaction. (D) Mechanical thresholds of the ipsilateral vs. contralateral hindpaws. (E) Tail withdrawal latencies in the tail immersion test. (F) Time (%) spent on the warm floor (40° C) in the thermal place test (TPT). (G) Time (s) a stranger conspecific spent sniffing each group. Data are means ± s.e.m. Dotted lines (---) represents mean baseline thresholds for all groups (B, C, D); Statistical tests included two-way repeated measures (A), one-way (B, E, F, G) and two-way (D) ANOVA with Holm-Sidak post hoc tests; *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 representative of post hoc comparisons. All statistical measure details are presented in Table S1A.

Fig. 2.. Activation of an ACC→NAc core circuit by the social transfer of pain.

(A) Timeline to “TRAP” activated neurons during social transfer of pain. (B) Representative photomicrographs of TdTomato positive cells (white arrows) in the ACC of Con, CFA and BY mice after social transfer of pain (C) Quantification of Ai14-positive/TRAPed cells across 13 brain regions (n = 6–9/group). (D) Schematic of AAV injection and representative photomicrographs of ACC injection site and fibers in the NAc core. (E) Schematic and timeline of monosynaptic rabies tracing. (F) Representative photomicrographs of NAc core RVdG/RG injection site and ACC GFP expression in CFA and BY mice. Data are means ± s.e.m. One-way ANOVA with Holm-Sidak post hoc tests comparing Con vs. BY, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. All statistical measure details are presented in Table S1B. Anterior cingulate (ACC), insula (INS), nucleus accumbens core (NAc), mediodorsal thalamus (MD Thal), interlaminar thalamus (IL Thal), ventrobasal thalamus (VB Thal), lateral habenula (L Hb), central amygdala (CeA), basolateral amygdala complex (BLAc), bed nucleus of the stria terminalis (BNST), zona inserta (ZI), ventral tegmental area (VTA), periaqueductal gray (PAG).

Fig. 3.. ACC→NAc projections bidirectionally control social transfer of pain.

(A) Schematic of viral injection and experimental timeline. Light stimulation (~5–15 mW, 8 s on/2 s off) periods represented by yellow-green boxes. (B) Schematic shows fiber optic implant above the ACC. First light off test of mechanical sensitivity (VF) of YFP and NpHR injected mice immediately after 1 h social transfer with ACC inhibition. (C) Mechanical sensitivity during averaged (n=2) light off and light on sessions of ACC inhibition. (D) Schematic shows fiber optic implant above the ACC. First light off test of mechanical sensitivity of YFP and NpHR injected TRAP2 mice immediately after 1 h social transfer with ACC-TRAP inhibition. (E) Mechanical sensitivity during averaged (n=2) light off and light on sessions of ACC-TRAP inhibition. (F) Schematic of bilateral fiber optic implants above the NAc core. First light off test of mechanical sensitivity of YFP and NpHR injected mice immediately after 1 h social transfer with ACC→NAc input inhibition. (G) Mechanical sensitivity during averaged (n=2) light off and light on sessions of ACC→NAc input inhibition. (H) First light off test of mechanical sensitivity at 0, 4, 24, 48, 74 h and 1 wk post 1 h social interaction with ACC→NAc input inhibition. Data are means ± s.e.m. Dotted line (---) represents mean baseline thresholds for all groups. *P < 0.05, **P < 0.01, ***P < 0.001, **** P < 0.0001; One-way ANOVA with Holm-Sidak post hoc tests comparing YFP to matched NpHR groups (B–G) and two-way repeated measures ANOVA with Holm-Sidak post hoc tests comparing treatment groups to baseline at each timepoint, notation is the least significant p value of all comparisons (H). All statistical measure details are presented in Table S1C.

Fig. 4.. Distinct ACC projections control the social transfer of pain and fear.

(A) Schematic and timeline of social transfer of fear (Shock Experience: 2 × 0.7 mA, 2 s duration, 1 min intervals; Shock Conditioning: 24 × 0.7 mA for 2 s duration, 10 s intervals, total of 4 min). (B) Freezing behavior of Shock and Bystander (BY) during conditioning phase of socially transferred fear. (C) Freezing behavior of BY mice during retrieval phase of socially transferred fear. (D) Schematic of YFP and NpHR injection and experimental timeline of optogenetic stimulation in NAc or BLA. Light stimulation (~10–15 mW, 8 s on/2 s off) periods represented by green boxes. (E) Conditioning session with inhibition of ACC→NAc projections in YFP- and NpHR-BY mice during shock observation. (F) Freezing behavior of YFP- and NpHR-BY mice during retrieval phase. (G) First light off session measuring mechanical sensitivity following NpHR inhibition of ACC→NAc projections during social transfer of pain in the same mice from E, F. Dotted line (---) represents mean baseline of all groups. (H) Conditioning session with inhibition of ACC→BLA projections in BY mice during shock observation. (I) Freezing behavior of YFP- and NpHR-BY mice during retrieval phase. (J) First light off session measuring mechanical sensitivity following NpHR inhibition of ACC→BLA projections during social transfer of pain in the same mice from H, I. Dotted line (---) represents mean baseline of all groups. Data are means ± s.e.m. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Two-way repeated measures ANOVA with Holm Sidak post hoc tests comparing freezing to the first minute, notation is the least significant p value of all comparisons (B, E, H), paired t-test (C) or unpaired t-test (F,I) comparing average baseline to average freezing during retrieval, or one-way ANOVA with Holm-Sidak post hoc tests (G,J). All statistical measure details are presented in Table S1D.

Fig. 5.. ACC→NAc projections regulate social transfer of analgesia.

(A) Schematic of social transfer of analgesia protocol and timeline. (B) Mechanical sensitivity at 0, 4, 24, 48, 72 h and 1 week post 1 h social interaction in CFA-Con/CFA-CON pairs and CFA-Mor/CFA-Analg-BY pairs. (C) Time (%) on the warm (40° C) floor in the thermal place test (TPT) prior to and 1 h and 1 week (wk) post CFA injection. (D) Time (%) on the warm (40° C) floor in the TPT immediately after the second 1 h social transfer of analgesia, 1 week post CFA injection. (E) Schematic of YFP and NpHR injection and optogenetic stimulation in ACC or NAc. Light stimulation (~10–15 mW, 8 s on/2 s off) periods represented by yellow-green boxes. (F) First light off session measuring mechanical sensitivity following light stimulation of ACC in YFP- and NpHR-expressing CFA-Con and CFA-Analg-BY mice during social transfer of analgesia. (G) Time (%) on the warm (40° C) floor in the TPT following light stimulation of ACC in YFP- and NpHR-expressing CFA-Con, CFA-Mor, and CFA-Analg-BY mice during the second social transfer of analgesia test. (H) First light off session measuring mechanical sensitivity following light stimulation of ACC→NAc projections in YFP- and NpHR-expressing CFA-Con and CFA-Analg-By mice during social transfer of analgesia. (I) Time (%) on the warm (40°C) floor in the TPT following ACC→NAc light stimulation in YFP- and NpHR-expressing CFA-Con, CFA-Mor, and CFA-Analg-BY mice during the second social transfer of analgesia test. Data are means ± s.e.m. *P < 0.05, **P < 0.01, ***P < 0.001, **** P < 0.0001. Two-way repeated measures (B) or one-way (C,D,F–I) ANOVA with Holm-Sidak post hoc tests comparing between groups. Not all significant post hoc analyses are displayed. All statistical measure details are presented in Table S1E.