Dynamics of ACC excitatory neurons during reversal learning: Implications for cognitive flexibility

Cognitive flexibility, a core component of adaptive behavior, enables individuals to adjust their actions when environmental contingencies change and is frequently impaired in neuropsychiatric conditions such as obsessive-compulsive disorder (OCD) and autism spectrum disorder (ASD). The anterior cingulate cortex (ACC), known to support error monitoring, action-outcome evaluation, and rule updating, has been implicated in cognitive flexibility, yet the precise role of its excitatory neurons remains unclear. Using fiber photometry in a touchscreen-based task, we monitored ACC excitatory neuron activity in mice during discrimination and reversal learning to test whether their activity varies across learning stages and relates to behavioral adaptation. By analyzing calcium dynamics in a ± 1-second window around the reward-seeking nosepoke, we found that ACC activity increased significantly during incorrect trials compared to correct trials, specifically in the middle phase of reversal learning. Moreover, the magnitude of ACC activity in both correct and incorrect trials correlated positively with task accuracy and negatively with the number of correction trials, respectively, highlighting a strong link between ACC engagement and adaptive performance during this critical transition phase. These results suggest that ACC excitatory neurons may contribute to performance monitoring and rule updating rather than merely encoding trial outcomes. Together, our findings underscore a temporally specific role for ACC excitatory neurons in cognitive flexibility and may inform future therapeutic strategies targeting ACC-related dysfunction in disorders of behavioral rigidity.