Investigating the role of neuromodulators in mice during associative learning with a 50% reward schedule

We aimed to investigate the role of dopamine (DA), acetylcholine (ACh), norepinephrine (NE) and serotonin (5-HT) in associative learning and the correlation of their release with the reward prediction error (RPE). To address this, we trained mice on a sound detection Pavlovian conditioning task with a 50% reward schedule that allowed examining clean representations of postivite (rewarded trials) and negative RPE (reward omission trials), while we measured DA, ACh, NE and 5-HT release by fiber photometry. Behavioral updating of value representations based on the outcome of the previous trial was indexed by the licking activity of mice in the anticipation of reward. As expected, anticipatory licking during the stimulus decreased after omitted rewards but increased after rewarded trials. DA release followed a similar pattern not only in the ventral striatum (VS) but also in the prefrontal cortex (PFC). We found significant positive correlations between DA release and anticipatory lick rate (ALR) difference in both brain areas. The release of ACh and 5-HT also followed the RPE in both the basolateral amygdala (BLA) and in the PFC. NE release in the PFC decreased after rewarded trials and increased after reward omissions. We observed a negative correlation in the PFC between NE release and ALR changes. These results indicate that DA, ACh and 5-HT follow the RPE in both examined brain areas. In contrast to the other three, NE release decreased after rewarded trials and increased following omitted rewards and it was negatively correlated with the intensity of the animals' anticipatory behavior.