Supplementary Figure 10: 4-Hz analog stimulation–mediated freezing behavior.

(a) Percentage of freezing displayed by ChR2 mice in context A during (Day 1 Stim. Ctx A), or 24 hours after (Day 2 No Stim. Ctx A) 4 Hz optogenetic analog stimulation, or 48 hours later in a neutral context (Day 2 No Stim. Ctx B) (n = 8 mice, Mann-Whitney U test, Day 1 or Day 2 versus Day 3: U = 2 and 0, ** P = 0.002; *** P < 0.001). (b) Left, representative micrograph of the motor cortex (M1) in a PV-cre mouse infected with ChR2. Scale bar = 0.5 mm. Right, Percentage of freezing displayed by PV-Cre mice infected in the motor cortex with ChR2 during motor cortex 4 Hz analog stimulation (n = 4 mice; Paired t-test, t(3) = 0.623, P = 0.578). (c) Left, representative micrograph of BLA in a PV-cre mouse infected with ChR2. Scale bar = 0.4 mm. Right, Averaged peri-stimulus time histograms (PSTH) showing that light activation of BLA PV interneurons can suppress the activity of BLA putative principal neurons (n = 4 cells from 2 mice). (d) Percentage of freezing displayed by PV-Cre mice infected in the BLA with ChR2 during baseline conditions (n = 5 mice; Paired t-test, t(4)= -0.721, P = 0.511). (e, f) Percentage of dmPFC (e, left) and BLA (f, left) neurons significantly phase-locked to 4 Hz oscillations and circular distribution of the 4 Hz oscillation preferred phase for populations of dmPFC (e, right) and BLA (f, right) phase-locked putative excitatory principal neurons (PNs) and inhibitory interneurons (INs) during dmPFC 4 Hz analog optogenetic stimulation (dmPFC: 25 PNs, 9 INs; BLA: 3 PNs, 1 INs). (g) Left, behavioral protocol. Right, Percentage of freezing behaviour to CS^-, and CS⁺ presentations during no-light or light exposure on Test 1 (n = 5 mice, Paired t-test, CS⁺ no light versus CS⁺ light, t(4)= -4.265, P = 0.013). Freezing levels to CS⁺ presentations during Test 2 normalized to CS⁺ evoked freezing levels during Test 1 (n = 5 mice, Paired t-test, CS⁺ Day 2 versus CS⁺ Day3, t(4)= -1.080, P = 0.341). Error bars: mean ± s.e.m. For the representative micrographs in (b and c), similar results were obtained for the animals used in these experiments (4 mice in b, and 5 mice in c). Data for panels (a) and (g) overlap with the data represented in Figure 7c and 7f, respectively.