Figure 2. Optical inhibition of VTA_DA→BLA projections during cue-reward pairing attenuates the encoding of identity-specific cue-reward memories.

(a-i) Optical inhibition of VTA_DA→BLA projections during Pavlovian long-delay conditioning with outcome-specific Pavlovian-to-instrumental transfer test. (a) Bottom: Representative fluorescent image of ArchT-tdTomato expression in VTA_DA neurons. Middle: Strategy for bilateral optogenetic inhibition of VTA_DA→BLA projections. Top: Representative image of fiber placement in the vicinity of immunofluorescent ArchT-tdTomato-expressing VTA_DA axons and terminals in BLA. (b) Schematic representation of ArchT-tdTomato expression in VTA and (c) placement of optical fiber tips in BLA for all subjects. (d) Pavlovian long-delay conditioning and Pavlovian-to-instrumental transfer procedure. A, action (left or right lever press); CS, 30-s conditioned stimulus (aka, “cue”, white noise or click) followed immediately by reward outcome (O, sucrose solution or grain pellet). (e) Lever-press rate averaged across levers and across the final 2 instrumental sessions. (f) Food-port entry rate during across Pavlovian conditioning. Three-way RM ANOVA, Training × Cue: F_{(4.09, 77.71)} = 5.73, P = 0.0004. (g-i) Outcome-specific Pavlovian-to-instrumental transfer test. (g) Lever-press rates on the lever that earned the “Same” outcome as predicted by the forthcoming or current cue or on the other available lever (Different). *P < 0.05, planned comparisons cue same presses v. preCue same presses and cue different presses v. preCue different presses. (h) Elevation in pressing [(Presses during cue)/(Presses during cue + preCue presses)]. Two-way RM ANOVA, Virus × Lever: F_{(1, 19)} = 9.22, P = 0.007. (i) Food-port entry rate. Two-way RM ANOVA, Cue: F_{(1, 19)} = 15.18, P = 0.001. ArchT, N = 11, 6 male rats; tdTomato, N = 10, 5 male rats. (j-q) Optical inhibition of VTA_DA→BLA projections during Pavlovian trace conditioning with outcome-specific devaluation test. (j) Bottom: Representative fluorescent image of ArchT-tdTomato expression in VTA_DA neurons. Middle: Strategy for bilateral optogenetic inhibition of VTA_DA→BLA projections. Top: Representative image of fiber placement in the vicinity of immunofluorescent ArchT-tdTomato-expressing VTA_DA axons and terminals in BLA. (k) Schematic representation of ArchT-tdTomato expression in VTA and (l) placement of optical fiber tips in BLA for all subjects. (m) Pavlovian trace conditioning and outcome-specific devaluation procedure. CS, 10-s conditioned stimulus (white noise or tone) following by 1.5-s trace interval before reward outcome (O, chocolate or unflavored purified pellets); LiCl, lithium chloride 0.3M, 1.5% volume/weight. (n) Percentage of time in the food-delivery port during Pavlovian conditioning. Three-way RM ANOVA, Training × Cue: F_{(1.61, 16.13)} = 31.49, P <0.0001. (o-p) Outcome-specific devaluation probe test. (o) Percentage of time in the food port during baseline, cue signaling the devalued reward and cue signaling the non-devalued (valued) reward. *P<0.05, **P<0.01, planned comparisons cue valued % time in port v. preCue % time in port and cue devalued % time in port v. preCue % time in port. (p) Elevation in percent time in food port [(CS % time in port)/(CS % time in port + preCue % time in port)]. Two-way RM ANOVA, Virus × Cue: F_{(1, 10)} = 5.20, P = 0.046. (q) Amount out of 100 available pellets consumed during post-test consumption choice. Two-way RM ANOVA, Value: F_{(1, 10)} = 249.00, P < 0.0001. ArchT, N = 5, 4 male rats; Control, N = 7, 4 male rats (3 WT/cre-dependent ArchT; 4 Th-cre/cre-dependent tdTomato). Data presented as trial-averaged, between-subject mean ± s.e.m. with individual data points. ^P = 0.059, *P<0.05, **P<0.01, ***P<0.001 Bonferroni-corrected post-hoc comparisons. See Supplemental Table 1 for full statistical reporting.