Article Figures & Data
Figures
- Figure 1.
Experiment 1. A, AAV was used to express GCaMP6 in aPVT or pPVT in separate animals and fiber optic cannulae implanted into these regions. Representative images show native GCaMP expression and DAPI staining in PVT (green, native GCaMP; blue, DAPI; scale bar, 100 μm), native GCaMP expression for aPVT and pPVT (scale bar, 500 μm), as well as location of fiber tips for all animals included in the analyses. B, Representative demodulated photometry traces from aPVT (during appetitive training) and pPVT (during aversive training). C, Rats received Pavlovian appetitive conditioning and acquired magazine entries during the CS. Photometry traces show mean ± SEM %ΔF/F across trials for the 1 s prior and 3 s after CS and appetitive US presentations. D, Rats received Pavlovian aversive conditioning and acquired fear responses to the CS. Photometry traces show mean ± SEM %ΔF/F across trials for the 1 s prior and 3 s after CS and aversive US presentations. E, Rats were tested for appetitive and aversive behaviors in their respective training contexts. There was robust expression of appetitive and aversive behaviors. Photometry traces show mean ± SEM %ΔF/F across trials for the 1 s prior and 3 s after CS presentations in each context. In each panel, a significant increase in %ΔF/F was determined whenever the lower bound of the 99% CI was >0. These points of statistical significance are shown as colored lines above each %ΔF/F curve with different colors corresponding to the respective traces from sessions (C) or trials (D). aPVT, n = 6; pPVT, n = 4. F, Scatterplots of subject-normalized CS-elicited PVT activity (AUC 0.5–1.5 s from CS onset) and associated conditioned responding across initial appetitive conditioning sessions. *Indicates lower bound 99% CI > 0%ΔF/F (colored lines above traces). aPVT, n = 6; pPVT, n = 4.
- Figure 2.
Experiment 2. A, AAV was used to express GCaMP6 in aPVT or pPVT in separate animals and fiber optic cannulae implanted into these regions. Representative images show native GCaMP expression and DAPI staining in PVT (green, native GCaMP; blue, DAPI; Scale bar, 100 μm), native GCaMP expression for aPVT and pPVT (Scale bar, 500 μm), as well as location of fiber tips for all animals included in the analyses. B, Representative demodulated photometry traces from aPVT (during aversive training) and pPVT (during appetitive training). C, Rats received Pavlovian aversive conditioning and acquired fear responses to the CS. Photometry traces show mean ± SEM %ΔF/F across trials for the 1 s prior and 3 s after CS and aversive US presentations. D, Rats received Pavlovian appetitive conditioning and acquired magazine entries during the CS. Photometry traces show mean ± SEM %ΔF/F across sessions for the 1 s prior and 3 s after CS and appetitive US presentations. E, Rats were tested for appetitive and aversive behaviors in their respective training contexts. Photometry traces show mean ± SEM %ΔF/F across trials for the 1 s prior and 3 s after CS presentations in each context. F, Scatterplots of subject-normalized CS-elicited PVT activity (AUC 0.5–1.5 s from CS onset) and associated conditioned responding across initial aversive conditioning trials. *Indicates lower bound 99% CI > 0%ΔF/F (colored lines above traces). aPVT, n = 10; pPVT, n = 8.
- Figure 3.
A, AAV was used to express eYFP or hM4Di in PVT, aPVT, or pPVT. Rats then received appetitive to aversive counterconditioning. B, Experiment 3a. Representative images showing hM4Di expression for whole PVT and placement map with each animal represented at 10% opacity (eYFP, n = 8; hM4Di, n = 7). C, Mean ± SEM appetitive responses at the end of appetitive conditioning, mean ± SEM aversive responses during aversive conditioning, and mean ± SEM appetitive and aversive responses on tests. Chemogenetic silencing of PVT on test reduced appetitive behavior and increased aversive behavior. Insets, Appetitive and aversive responses across four blocks of two test trials. D, Experiment 3b. Representative images showing hM4Di expression for aPVT and pPVT for the aPVT experiment and placement map with each animal represented at 10% opacity (eYFP, n = 8; hM4Di, n = 5). E, Mean ± SEM appetitive responses at the end of appetitive conditioning, mean ± SEM aversive responses during aversive conditioning, and mean ± SEM appetitive and aversive responses on tests. Chemogenetic silencing of aPVT on test had no effect on appetitive or aversive behavior. F, Experiment 3c. Representative images showing hM4Di expression for aPVT and pPVT for the pPVT experiment and placement map with each animal represented at 10% opacity (eYFP, n = 7; hM4Di, n = 8). G, Mean ± SEM appetitive responses at the end of appetitive conditioning, mean ± SEM aversive responses during aversive conditioning, and mean ± SEM appetitive and aversive responses on tests. Chemogenetic silencing of pPVT on test had no effect on appetitive or aversive behavior. H, Experiment 4. AAV was used to express eYFP or hM4Di in PVT, rats then received aversive to appetitive counterconditioning. I, Representative images showing hM4Di expression for PVT and placement map with each animal represented at 10% opacity (eYFP, n = 8; hM4Di, n = 7). J, Mean ± SEM aversive responses during aversive conditioning, mean ± appetitive SEM responses at the end of appetitive conditioning, and mean ± SEM appetitive and aversive responses on tests. #p < 0.05.
- Figure 4.
A, Experiments 5a and 5b. AAV was used to express eYFP or hM4Di in PVT and rats then received appetitive conditioning using liquid (Experiment 5a) or pellet (Experiment 5b) reward. B, Experiment 5a. Placement map showing hM4Di expression with each animal represented at 10% opacity, eYFP (n = 8) or hM4Di (n = 8). Mean ± SEM appetitive responses at the end of appetitive conditioning and during test. Chemogenetic silencing of PVT had no effect on appetitive behaviors. C, Experiment 5b. Placement map showing hM4Di expression with each animal represented at 10% opacity in PVT, eYFP (n = 8) or hM4Di (n = 7). Mean ± SEM appetitive responses at the end of appetitive conditioning and during test. Chemogenetic silencing of PVT on test had no effect on appetitive behaviors. D, Experiment 5c. AAV was used express eYFP (n = 8) or hM4Di (n = 8) in PVT and rats then received aversive conditioning. Placement map showing hM4Di expression with each animal represented at 10% opacity. Mean ± SEM aversive responses during aversive conditioning and during test. Chemogenetic silencing of PVT had no effect on aversive behaviors. E, Experiment 5d. Chemogenetic silencing of PVT also had no effect on locomotor activity when assessed in an open field. F, Experiment 6. c-Fos immunohistochemistry was used to verify PVT chemogenetic inhibition, group hM4Di n = 8, eYFP n = 8. Example of single Fos (black arrowhead), single eYFP (gray arrowhead), and dual-labeled c-Fos/eYFP neurons (white arrowhead) in PVT. Mean ± SEM numbers of total c-Fos, total dual-labeled c-Fos/eYFP, and percentage dual-labeled c-Fos/eYFP neurons in PVT after CNO injection. #p < 0.05.
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