Articles

We investigated how targeted reactivation of cortical memory engrams during sleep influences systems consolidation of contextual fear memory. Using a mouse model of contextual fear conditioning (CFC), we tagged retrosplenial cortex (RSC) neurons activated during learning with channelrhodopsin-2 (ChR2) via a c-Fos-based TRAP system. Twenty-four hours post-training, animals received optogenetic stimulation of these RSC engram cells selectively during non-rapid eye movement (NREM) sleep or REM sleep. Immediately after, hippocampal activity was transiently blocked (muscimol infusion) to test hippocampal dependence. We found that NREM-specific reactivation of RSC engrams preserved contextual fear recall despite hippocampal inactivation, whereas REM stimulation or no stimulation failed to rescue memory. These results indicate that engram reactivation during NREM accelerates the transfer of memory to cortical storage. We present high-level diagrams of experimental workflow and results, and report a timeline of revisions. Abbreviations and keywords are provided. This comprehensive revision addresses all reviewer comments and renders the manuscript submission-ready.