TIME- AND SEX-DEPENDENT CHARACTERIZATION OF ENGRAM SPINE MORPHOLOGY IN FEAR MEMORY PROCESSING

Fear memories are dynamic and undergo time-dependent changes that allow organisms to adapt their behavior. While fear can be reduced through extinction training, extinguished fear may relapse over time. Despite the well-characterized neural circuitry, how fear memories are processed in the brain, especially at the level of their neural correlates, across time and sex to optimally guide behavior remains unclear. Engram neurons, i.e., neurons activated during training, are believed to store memories and to undergo long-lasting synaptic plasticity, including changes in dendritic spine structure. However, little is known about whether and how dendritic spine features of fear-related engram neurons are shaped across time and sex.
Here, we combined activity-dependent engram tagging with fear-related behavioral paradigms to characterize dendritic spine morphology in hippocampal neuronal ensembles in male and female mice over time. Separate cohorts of mice underwent either (i) contextual fear conditioning (CFC), (ii) CFC followed by extinction training and spontaneous recovery testing, or (iii) remained in the home-cage as controls, and were tested for memory retention at recent (3 days) or remote (3 weeks) timepoints. In addition to CFC-tagged neurons, ensembles of randomly tagged neurons were analyzed to distinguish CFC-associated spine morphology from pre-existing structural variability across sexes. Dendritic spines were imaged using super-resolution confocal microscopy, and quantified for density and morphological features. Estrous cycle stage in female mice was monitored throughout.
Overall, this work provides a systematic characterization of dendritic spine properties in fear memory-relevant engram neurons across fear behavioral state, time, and sex.