Fig. 1: Differential responses of PVT excitatory neurons to CS- and CS+ after fear condition and safety learning.

A Injection site of GCaMP6s virus for monitoring Ca²⁺ transients in PVT neurons. Scale bar: 100 μm. B Average and heat map of Ca²⁺ transients in response to different stimuli (air puff, tail suspension, and foot shock). C Experimental protocol for fear conditioning and safety learning. D Freezing levels(Left) during CS- and CS+ retrieval and discrimination index (middle) 24 h after FC and safety learning (two-way ANOVA, interaction, F(1, 24) = 11.21, P < 0.01, stimulus, F(1, 24) = 21.20, P < 0.0001, training, F(1, 24) = 0.09946, P = 0.7552; N = 7 mice, CS-(post-FC)vs. CS+(post FC), P = 0.7663, CS-(post SL) vs. CS + (post SL), P < 0.0001, CS-(post FC) vs. CS-(post SL), P < 0.05, CS + (post FC) vs. CS + (post SL), P = 0.0846, Bonferroni’s post-test; two-tailed paired t-test, t = 4.191, df = 6; N = 7 mice, Post FC vs. Post SL, P < 0.01). Right: Freezing levels during CS-, inter-trial-interval (ITI) and CS + 24 h after SL (one-way ANOVA, N = 7 mice, CS- vs. ITI, P < 0.01, ITI vs. CS+, P < 0.01, CS- vs. CS+, P < 0.01, Bonferroni’s post-test). E Heat maps of (upper) and averaged (lower) PVT neuron responses during CS- and CS+ post-fear conditioning (two-tailed paired t-test, t = 1.509, df = 6; N = 7 mice, CS- vs. CS+, P = 0.182). F Heat maps of (upper) and averaged (lower) PVT neuron responses during CS- and CS+ after safety learning (two-tailed paired t-test, t = 3.823, df = 6; N = 7 mice, CS- vs. CS+, P < 0.01). G The weak foot shock (0.3 mA) fear conditioning protocol and freezing levels during CS- and CS+ retrieval 24 h after FC (two-tailed paired t-test, t = 4.734, df=16; N = 9 mice, CS- vs. CS+, P < 0.01). H Averaged PVT neuron responses during CS- and CS+ post-FC (two-tailed paired t-test, t = 3.534, df =8; N = 9 mice, CS- vs. CS+, P < 0.01).