Fig. 6: ACC-to-PAG circuit regulates observational fear and affective pain behaviors.

a, b Injection and implantation strategy for optogenetic inhibition of the ACC terminals projecting to the PAG a, d, g; n = 18 (eGFP); n = 24 (NpHR)) or BLA b, e, h; n = 18 (eGFP); n = 25 (NpHR)). Representative confocal images showing virus expression in the right ACC and bilateral placement of optic fibers in the PAG or BLA. Scale bar, 200 μm. Green, eGFP; Blue, DAPI. c Experimental paradigm for optogenetic inhibition during OFC. A yellow laser was delivered bilaterally during the 4-minute conditioning period. d Optogenetic silencing of the ACC → PAG circuit significantly impaired the acquisition of vicarious freezing (%) in observer mice during the conditioning period of OFC (two-way repeated measures ANOVA, Bonferroni’s post hoc test, F_{(1, 40)} = 5.130, p = 0.0290). e Optogenetic inhibition of the ACC → BLA pathway significantly reduced vicarious freezing response (two-way repeated measures ANOVA, Bonferroni’s post hoc test, F_{(1, 41)} = 13.11, p = 0.0008). See also Supplementary Table. 1. f Schematic of the experimental paradigm for optogenetic inhibition in the PEAP. A yellow laser was delivered bilaterally for 30 minutes. Mice received noxious mechanical stimulus (pin prick) on the dark side and innocuous stimulation on the light side. g, h The mean time (%) spent on the light side for the optogenetic inhibition of the ACC → PAG circuit (g) and the ACC → BLA circuit (h). g Optogenetic inhibition of the ACC → PAG circuit significantly reduced the time spent on the light side compared to the control group (two-way repeated measures ANOVA, Bonferroni’s post hoc test, F_{(1, 40)} = 21.21, p < 0.0001). h Optogenetic suppression of the ACC → BLA circuit did not result in significant alterations in the time spent on the light side. Source data are provided as a Source Data file. * p < 0.05, ** p < 0.01, *** p < 0.001. All data are presented as means ± S.E.M.