No abstract

Astrocytes are key cellular regulators within the brain. The basolateral amygdala (BLA) is implicated in fear memory processing, yet most research has entirely focused on neuronal mechanisms, despite a significant body of work implicating astrocytes in learning and memory. In the present study, we usedin vivofiber photometry in C57Bl/6J male mice to record from amygdalar astrocytes across fear learning, recall, and three separate periods of extinction. We found that BLA astrocytes robustly responded to foot shock during acquisition, that their activity remained remarkably elevated across days in comparison to unshocked control animals, and that their increased activity persisted throughout extinction. Further, we found that astrocytes responded to the initiation and termination of freezing bouts during contextual fear conditioning and recall, and this behavior-locked pattern of activity did not persist throughout the extinction sessions. Importantly, astrocytes do not display these changes while exploring a novel context, suggesting that these observations are specific to the original fear-associated environment. Chemogenetic inhibition of fear ensembles in the BLA did not affect freezing behavior or astrocytic calcium dynamics. Overall, our work presents a real-time role for amygdalar astrocytes in fear processing and provides new insight into the emerging role of these cells in cognition and behavior.SIGNIFICANCE STATEMENT:We show that basolateral amygdala astrocytes are robustly responsive to negative experiences, like shock, and display changed calcium activity patterns through fear learning and memory. Additionally, astrocytic calcium responses become time-locked to the initiation and termination of freezing behavior during fear learning and recall. We find that astrocytes display calcium dynamics unique to a fear-conditioned context and chemogenetic inhibition of BLA fear ensembles does not impact freezing behavior or calcium dynamics. These findings show that astrocytes play a key, real-time role in fear learning and memory.

Memory engrams are both necessary and sufficient to mediate behavioral outputs. Defensive behaviors such as freezing and avoidance are commonly examined during hippocampal-mediated fear engram reactivation, yet how reactivation of these cellular populations across different contexts engages the brain to produce a variety of defensive behaviors is relatively unclear. To address this, we first optogenetically reactivated a tagged fear engram in the dentate gyrus (DG) subregion of the hippocampus across three distinct contexts. We found that there were differential amounts of light-induced freezing depending on the size of the context in which reactivation occurred: mice demonstrated robust light-induced freezing in the most spatially restricted of the three contexts but not in the largest. We then utilized graph theoretical analyses to identify brain-wide alterations in cFos co-activation during engram reactivation across the smallest and largest contexts. Our manipulations conferred greater positive cFos correlations and recruited regions spanning putative fear and defense systems as hubs in the respective networks. Moreover, reactivating DG-mediated engrams generated network topologies across experimental conditions, emphasizing both shared and distinct features. By identifying and manipulating the circuits supporting memory function, as well as their corresponding brain-wide activity patterns, it is thereby possible to resolve systems-level biological mechanisms mediating memory's capacity to modulate behavioral states.

Acknowledgements: Thank you to Bret Judson and the Boston College Imaging Core for infrastructure and support.

Network dysfunction has been implicated in numerous diseases and psychiatric disorders, and the hippocampus serves as a common origin for these abnormalities. In this study, we tested the hypothesis that chronic induction of local changes in neurons and astrocytes is sufficient to induce impairments in cognition and behavior. We chronically activated the hM3D(Gq) pathway in CaMKII+ neurons or GFAP+ astrocytes within the ventral hippocampus across 3, 6 and 9 months. We observed that CaMKII-hM3Dq activation impaired fear acquisition, decreased anxiety and social interaction, and modified spatial odor memory and novel environment exploration, while GFAP-hM3Dq activation impaired fear acquisition and enhanced recall. CaMKII-hM3Dq activation modified the number of microglia, while GFAP-hM3Dq activation impacted microglial morphological characteristics, but neither affected astrocytes. Manipulation of both cell types increased the presence of phosphorylated tau at the earliest time point. Overall, our study provides evidence for how each of these cell types are uniquely engaged in disorders that have characteristic network dysfunction while adding a more direct role for glia in modulating behavior.