Pre-existing and emergent cortical neuronal assembly sequences during learning

Neuronal assemblies -- groups of co-active neurons -- support memory consolidation and retrieval. In the hippocampus, assemblies can pre-exist learning and contribute to memory through sequential activation. Whether similar principles apply to higher cortical areas for flexible memory storage like the orbitofrontal cortex (OFC) remains unclear. Using a novel ground truth-validated clustering approach, we investigated the activity of longitudinally tracked mouse OFC neurons during cue-reward memory acquisition and maintenance. Assemblies active after learning pre-existed the learning and exhibited two distinct sequential dynamics consistent with memory consolidation or retrieval. Consolidation sequences emerged during learning, while retrieval sequences partly recruited pre-existing reward sequences. These findings demonstrate that OFC learning recruits pre-existing networks flexibly repurposed for new associations, revealing circuit motifs that may enable cortical memory storage.