Fig. 2: The activation of vCA1 neurons projecting to the BA, paired with aversive stimuli, generated a new conditioned fear memory.

a An optical cannula was implanted to photostimulate vCA1 neurons projecting to the BA (vCA1: BA projectors) expressing Chronos-GFP (Chronos group) or eYFP (eYFP group). Retrograde CAV2-Cre was injected into the BA, and AAV-DIO-Chronos-GFP or AAV-DIO-eYFP was injected into the vCA1. b Left: image showing eYFP expression (green) in vCA1 neurons and optical cannula tip (arrow). Right: diagrams showing optical cannula implantation sites in d and e. c Experimental setup for d, e. Mice were habituated to 20 Hz photostimulation in Context C on days 1–3. After a 3-min acclimatization period and baseline recording of freezing behavior for 1 min, 20 Hz photostimulation was applied to the vCA1 through an optical cannulae (blue). On day 4, the mice received 20 Hz photostimulation (blue bars) 6 times, each co-terminating with a footshock, in Context A. On days 5–6, the mice were tested for freezing behavior in Context C in the presence and absence of 20 Hz photostimulation. d The time course of freezing behavior in Context C during habituation (HB) and test sessions in the Chronos (7 mice) and eYFP groups (6 mice). Freezing time on Days 1–3 (HB) and Days 5–6 (test) were averaged in each mouse for each time bin. e Summary plot showing the difference in the average freezing time in the presence and absence of photostimulation (ON – OFF freezing) during habituation and test sessions. *p < 0.05 (two-way ANOVA with post hoc comparisons, group × behavioral session interaction, p < 0.01). f Experimental setup for g. Mice in the Chronos: unpaired group underwent surgery with AAV-DIO-Chronos-GFP injected into the vCA1 as in a. On the training day, the mice received 20 Hz photostimulation 6 times in Context A and then received 6 shocks in Context A 30 min later. The mice were tested for photostimulation-induced freezing behavior in Context C. g Left: the time course of freezing behavior during habituation and test sessions in the Chronos: unpaired group (9 mice). Right: summary plot of the difference in the average freezing time in the presence and absence of photostimulation (ON – OFF freezing). p = 0.39, habituation vs. test sessions (two-sided paired t-test). Error bars represent the SEM. Source data are provided as a Source Data file. See also Supplementary Fig. 3.