Neural Estimates of Imagined Outcomes in Basolateral Amygdala Depend on Orbitofrontal Cortex

Abstract

Reciprocal connections between the orbitofrontal cortex (OFC) and the basolateral nucleus of the amygdala (BLA) provide a critical circuit for guiding normal behavior when information about expected outcomes is required. Recently, we reported that outcome signaling by OFC neurons is also necessary for learning in the face of unexpected outcomes during a Pavlovian over-expectation task. Key to learning in this task is the ability to build on prior learning to infer or estimate an amount of reward never previously received. OFC was critical to this process. Notably, in parallel work, we found that BLA was not necessary for learning in this setting. This suggested a dissociation in which the BLA might be critical for acquiring information about the outcomes but not for subsequently using it to make novel predictions. Here we evaluated this hypothesis by recording single-unit activity from BLA in rats during the same Pavlovian over-expectation task used previously. We found that spiking activity recorded in BLA in control rats did reflect novel outcome estimates derived from the integration of prior learning, however consistent with a model in which this process occurs in the OFC, these correlates were entirely abolished by ipsilateral OFC lesions. These data indicate that this information about these novel predictions is represented in the BLA, supported via direct or indirect input from the OFC, even though it does not appear to be necessary for learning.

Significance statement: The basolateral nucleus of the amygdala (BLA) and the orbitofrontal cortex (OFC) are involved in behavior that depends on knowledge of impending outcomes. Recently, we found that only the OFC was necessary for using such information for learning in a Pavlovian over-expectation task. The current experiment was designed to search for neural correlates of this process in the BLA and, if present, to ask whether they would still be dependent on OFC input. We found that although spiking activity in BLA in control rats did reflect the novel outcome estimates underlying learning, these correlates were entirely abolished by OFC lesions.

Keywords: amygdala; extinction; orbitofrontal; over-expectation; rat; single-unit.

Figure 1.

Task design and histology. a, Shown is the task design and experimental timeline. A1, A2, and A3 are auditory cues (tone, white noise, and clicker, counterbalanced). V is a visual cue (a cue light). Two differently flavored sucrose pellets were used as reward (banana- or grape-flavored sucrose pellets, represented by solid or empty circles, counterbalanced). Training began with 12 conditioning sessions (CD1–CD12), in which each cue was presented eight times. A1 and V cues were paired with the same reward (3 pellets) and A2 was paired with the other reward (3 pellets). A3 was paired with no reward. After completion of the last conditioning session, rats underwent a single compound probe session (CP1) followed by three compound training sessions (CP2–CP4). During the first half of the compound probe session (CP 1/2), rats continued to receive simple conditioning. During the second half (CP 2/2), rats began compound training in which A1 and V were presented together as a compound (A1/V), followed by delivery of the same reward (3 pellets). A2, A3, and V continued to be presented as in simple conditioning. During the compound training sessions (CP2–CP4), rats received presentations of A1/V, A2, A3, and V. After completion of the last compound training session, rats underwent a single extinction probe session (PB). The first half of the session (PB 1/2) consisted of further compound training. During the second half of the session (PB 2/2), rats received eight non-reinforced presentations of A1, A2, and A3 with the order mixed and counterbalanced. b, Drawing illustrating the location of recording sites in BLA in control group and OFC-lesioned animals. Boxes indicate approximate location of recording sites, taking into account any vertical distance traveled during training and the approximate lateral spread of the electrode bundle. c, Minimum (black) and maximum (line) OFC lesion extent are shown for bregma +4.7, +3.7, +2.7 (adapted from Paxinos and Watson, 1998).

Figure 2.

Conditioned responding and cue-evoked activity during simple conditioning. a, b, Plot illustrating increase in conditioned responding as a percentage of time in the food cup during each of the four cues across sessions in control (a), and in OFC-lesioned (b) trained rats. Red diamond, A1; blue square, A2; green circle, A3; and yellow triangle, V. c, d, Proportions of neurons that were significantly responsive to any of the four cues in control (c), and in OFC-lesioned (d) trained rats. Bars indicate percentage of cue-responsive neurons within each pair of sessions and separated by those that increased (white) or decreased (black) firing rate compared with baseline. *p < 0.05; ns, Not significant.

Figure 3.

Cue-evoked activity summates at the start of compound training in control but not in OFC-lesioned rats. a, b, Conditioned responding in control (a) and OFC-lesioned (b) trained rats at the end of conditioning (CP 1/2) and through compound training (CP 2/2 and CP2-CP4). Error bars indicate SEM. c–f, Population activity across all cue-responsive neurons to A1, V (c, d), and A2 (e, f) during the compound probe session; dark and light lines illustrate activity during the conditioning and compound phases of the session, respectively. Gray shading indicates SEM, and gray horizontal bars indicate the period of cue presentation. g–j, Distribution of summation index scores for firing to A1 (g, i) and A2 (h, j) in the compound probe. Index scores were computed for each neuron based on the change in mean normalized firing to the relevant cue between conditioning and compound training, using the following formula: (firing CP 2/2 − firing CP 1/2)/(firing CP 2/2 + firing CP 1/2). Black bars represent neurons in which the difference in firing was statistically significant (t test, p < 0.05). k, l, Scatter plots showing relationship between the change in behavior and neural activity to A1 in the compound probe session. Neural summation index scores were computed for firing to A1 as described above; behavioral summation index scores were computed similarly, for each session in which a cue-responsive neuron was recorded, but using conditioned responding instead of firing. m, n, Line plots showing the ratio between normalized firing to A1 and A2 during each compound training session (CP–CP4). N values indicate number of cue-responsive neurons in each session. Error bars indicate SEM. *p < 0.05.

Figure 4.

Cue-evoked activity spontaneously declined at the start of extinction training in control but not in OFC-lesioned rats. a, b, Conditioned responding in control (a) and OFC-lesioned (b) trained rats as a percentage of time in the food cup during each cue at the end of compound training (PB 1/2) and during the eight trials of extinction (Trials 1–8 and bar graph showing means). Error bars indicate SEM (*p < 0.05). c–f, Population activity across all cue-responsive neurons to A1, V(c, d) and A2 (e, f) during the extinction probe session; light and dark lines illustrate activity during the compound phase and on the first trial (1T) of extinction during the session, respectively. Gray shading indicates SEM, and gray horizontal bars indicate the period of cue presentation. g–j, Distribution of over-expectation index scores for firing to A1 (g, i) and A2 (h, j) in the extinction probe. Index scores were computed for each neuron based on the change in mean normalized firing to the relevant cue between compound training and the first trial of extinction, using the following formula: (firing PB 1T − firing PB 1/2)/(firing PB 1T + firing PB 1/2). Black bars represent neurons in which the difference in firing was statistically significant (t test, p < 0.05). k, l, Scatter plots showing relationship between the change in behavior and neural activity to A1 on the first trial of extinction training. Neural over-expectation index scores were computed for firing to A1 as described above; behavioral over-expectation index scores were computed similarly, for each session in which a cue-responsive neuron was recorded, but using conditioned responding instead of firing. m, n, Scatter plots showing relationship between the change in behavior on the first trial of extinction and neural activity to A1 at the start of compound training. Neural summation index scores were computed for firing to A1 as described in Figure 3.