A D1 Receptor-Dependent Dopamine Circuit Targeting VIP Interneurons in the Anterior Insular Cortex

Pain is a subjective, emotional experience arising from distributed brain activity integrating sensory and affective components. The anterior insular cortex (AIC) is central to this process, and dopaminergic signaling via D1 receptors (D1R) in the AIC has been linked to pain modulation. However, the cellular identity and functional properties of D1R-expressing neurons within AIC microcircuits remain poorly characterized. Here, we used anatomical, electrophysiological, and behavioral approaches to characterize D1R-expressing neurons in the mouse AIC. In D1R-Tomato mice, D1R- positive neurons showed a layer-dependent distribution, with superficial layers enriched in inhibitory interneurons and deeper layers containing pyramidal neurons and interneurons. Immunohistochemistry showed that most superficial D1R-expressing interneurons co-express VIP, lacking parvalbumin or somatostatin markers. Consistently, morphological reconstructions revealed bipolar and multipolar architectures typical of VIP interneurons. Ex vivo recordings showed that D1 receptor activation enhances the excitability of D1R-positive neurons, indicating a direct dopaminergic influence on this population. Finally, using c-fos as a marker, we assessed the activation of AIC D1R-expressing neurons during acute visceral pain. Together, these results provide an integrated characterization of D1R-expressing neurons in the AIC and link their recruitment to cortical processing during acute pain.