Subregional and Laminar Specializations of Pyramidal Neurons in the Macaque Anterior Cingulate Cortex

The anterior cingulate cortex (ACC)-a component of both the limbic system and the frontal-executive network-is composed of three anatomically distinct subregions with diverse roles in processing cognitive and emotional information: rostral area 32 (A32), dorsal area 24 (A24), and ventral area 25 (A25). Pyramidal neurons (PYRs) as the neural substrates of cortical communication govern signal processing and integration differently across brain regions, layers, and pathways. Their properties and diversity across the heterogeneous ACC subregions have yet to be comprehensively characterized in primates. Here, we compared the biophysical and morphological properties of PYRs in layers 2-3 (L2-3) and 5-6 (L5-6) across ACC subregions of young adult rhesus monkeys (8 females, 11 males) using in vitro whole-cell patch-clamp recording with intracellular filling. Results showed the more pronounced laminar differences in biophysical properties of PYRs in A24 and A32 than in A25, with L2-3 PYRs exhibiting lower excitability than L5-6 PYRs. Further, A32 contained a higher proportion of PYRs receiving high-frequency-sEPSC and an enrichment of mushroom spines in L2-3 compared with L5-6. This heterogeneity in laminar differentiation aligns with cytoarchitectural differences across ACC subregions and suggests layer-specific signaling and excitatory drive in A24 and A32. In contrast, A25 showed cellular heterogeneity in firing patterns as well as different inhibitory signaling dynamics from other ACC subregions. Together, these results highlight the heterogeneity in laminar differentiation of PYR properties across ACC subregions, contributing to their diverse circuits and roles in cognitive-emotional integration and disruption in disease.