A multiple (mArc) tagging system to uncover the organizational principles of multiple memories.

Unlabelled: Engrams or memory traces are the neuronal ensembles that collectively store individual experiences. Genetic strategies based on immediate early genes (IEGs), such as , allow us to tag the ensembles active during memory encoding and compare them to those active during retrieval. However, these strategies only allow for the tagging of one neural ensemble. Here, we developed a multiple (mArc) system that allows for the tagging of two ensembles. We validated this system by investigating how context, time, and valence influence neuronal ensemble reactivation in the dentate gyrus (DG). We show that similar contextual and valenced experiences are encoded in overlapping DG ensembles. We also find that ensembles are modulated by time, where experiences closer in time are encoded in more similar ensembles. These results highlight the dynamic nature of DG ensembles and show that the mArc system provides a powerful approach for investigating multiple memories in the brain.

Highlights: The mArc system allows for the tagging of two ensembles in the same mouse DG ensembles labeled by the mArc system receive increased excitatory inputContext, valence, and time influence DG ensemble reactivationDG neural ensembles are reactivated less with increasing time.