Prefrontal Neurons Projecting to the Dorsomedial Striatum and Insular Cortex Support Reward Learning

The medial prefrontal cortex (mPFC) contains projection-defined neuronal populations that route information to multiple downstream targets. However, their role in reward learning remains unclear. Here, we identified and functionally characterized a subpopulation of mPFC neurons collateralizing to the dorsomedial striatum (DMS) and the contralateral anterior insular cortex (aIC). Fiber photometry and single-cell calcium imaging revealed that mPFC^DMS+aIC neurons responded robustly to reward stimuli and exhibited learning-enhanced, unconditioned stimulus (US)-evoked activity during reward conditioning, whereas conditioned stimulus (CS) responses remained stable. This enhancement reflected an increased response magnitude and a greater proportion of US-responsive neurons. Importantly, optogenetic inhibition of these neurons during the US period, but not during the CS period, impaired the acquisition of conditioned anticipatory licking. Taken together, our findings identify a finely projection-defined mPFC neuronal subpopulation that preferentially contributes to reinforcement-related updating during reward learning.