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. 2020 Dec 15;9:e60812. doi: 10.7554/eLife.60812

Figure 3. Inhibition of mPFC→BNST neurons increases PRF.

(A) Schematic depiction of experimental procedure for assessing effects of chemogenetic inhibition of mPFC→BNST neurons during retrieval. (B) Cartoon of viral strategy and representative images of hM4Di–mCherry labeling in BNST neurons receiving mPFC input (scale bars = 200 µm). (C) Lower CS-related freezing during retrieval in PRF mice than in FRF mice transfected with mCherry, not hHM4Di. Data are means ± SEM. *p<0.05.

Figure 3.

Figure 3—figure supplement 1. Freezing during conditioning prior to mPFC→BNST inhibition on retrieval.

Figure 3—figure supplement 1.

(A) Schematic depiction of viral strategy to selectively inhibit BNST-projecting mPFC neurons during retrieval. (B) Freezing increased across CS trials, irrespective of virus group. (C) Trial-by-trial breakdown of freezing during each CS of retrieval indicated a non-significant trend for decreasing freezing across trials in the mCherry PRF group. n = 8–9 mice group/virus. Freezing data are means ± SEM.
Figure 3—figure supplement 2. Electrode placements and virus localization for combined chemogenetic/single-unit recordings.

Figure 3—figure supplement 2.

Estimated location of tips of the electrodes at the center of the multi-array in the IL of PRF (A) and FRF (B) mice. Estimated extent of virus, as indicated by mCherry expression, in the BNST of PRF (C) and FRF (D) mice (darker shading represents areas of greater overlap across mice).
Figure 3—figure supplement 3. CS and freezing-related IL unit activity and effects of mPFC→BNST inhibition.

Figure 3—figure supplement 3.

(A) Schematic depiction of experimental procedure for in vivo IL single-unit recordings, combined with chemogenetic inhibition of mPFC→BNST neurons during retrieval (n = 8–9 mice per group/virus). (B) Cartoon of viral strategy and representative image of hM4Di-mCherry labeling in the mPFC and electrode array placement in the IL (scale bar = 500 µm). (C) Raster plot of a representative IL unit firing in response to CS onset (CS-ON neuron). Baseline-normalized population trace of CS-ON neuronal activity during retrieval; average of all groups (D) and split by group (E). (F) Higher percentage of CS-ON units during PRF than FRF retrieval in mice transfected with mCherry, not hHM4Di (n = 17 recorded units in PRF/mCherry, n = 25 units in PRF/hM4Di, n = 20 units in FRF/mCherry, n = 17 units in FRF/hM4Di, from three mice per group/virus). Baseline-normalized population trace of freeze cessation (Freeze-OFF neurons) IL unit activity during retrieval; average of all groups (G) and split by group (H) (n = 3 CS units in PRF/mCherry, n = 6 units in PRF/hM4Di, n = 1 units in FRF/mCherry, n = 1 units in FRF/hM4Di). (I) Higher percentage of Freeze-OFF units during PRF than FRF retrieval in mice transfected with mCherry, not hHM4Di (n = 17 recorded units in PRF/mCherry, n = 25 units in PRF/hM4Di, n = 20 units in FRF/mCherry, n = 17 units in FRF/hM4Di, from three mice per group/virus, n = 4 Freeze-OFF units in PRF/mCherry, n = 6 units in PRF/hM4Di, n = 3 units in FRF/mCherry, n = 0 units in FRF/hM4Di). Baseline-normalized population trace of freeze onset (Freeze-ON neurons) IL unit activity during retrieval; average of all groups (J) and split by group (K). (L) No differences in the percentage of Freeze-OFF units during retrieval between groups (n = 17 units in PRF/mCherry, n = 25 units in PRF/hM4Di, n = 20 units in FRF/mCherry, n = 17 units in FRF/hM4Di, from three mice per group/virus, n = 4 Freeze-ON units in PRF/mCherry, n = 6 units in PRF/hM4Di, n = 4 units in FRF/mCherry, n = 2 units in FRF/hM4Di). Data are means ± SEM. *p<0.05.
Figure 3—figure supplement 4. Heat maps illustrating IL unit activity.

Figure 3—figure supplement 4.

(A) Heat plots of unit activity aligned to CS onset (left columns), freeze cessation (center columns), and freeze onset (right columns). The same data shown as peri-event histograms and % event-related activity can be found in Figure 3—figure supplement 3.