Capturing dynamic neuronal responses to dominant and subordinate social hierarchy members with catFISH

Dominance hierarchies are key to social organization in group-living species, requiring individuals to recognize their own and others' ranks. This is particularly complex for mid-ranking animals, who navigate interactions with higher- and lower-ranking individuals. Using in situ hybridization, we examined how mid-ranked mice's brains respond to dominant and subordinate stimuli by labeling activity-induced immediate early genes and neuronal markers. We show that distinct neuronal populations in the amygdala and hippocampus respond differentially across social contexts. In the basolateral amygdala and dorsal endopiriform, glutamatergic Slc17a7⁺ neurons, particularly dopamine-receptive Slc17a7⁺Drd1⁺ neurons, show elevated IEG expression in response to social stimuli, with a higher response to dominant over subordinate animals. Similar response patterns are observed among GABAergic Slc32a1⁺Oxtr⁺ neurons in the medial amygdala. We also identified distinct neural ensembles selectively active in response to dominant and subordinate animals by examining cell reactivation over repeated stimulus presentations. We find a higher degree of reactivation among Slc17a7⁺Oxtr⁺ ensembles in the endopiriform when the same individual was presented twice in succession. A similar pattern was observed among Oxtr⁺ neurons in the dentate gyrus hilus, while the inverse was observed among Slc17a7⁺Avrp1b⁺Oxtr⁺ neurons in distal CA2CA3, suggesting distinct encoding or recollection mechanisms across hippocampal subregions. We also highlight methodological advances showing that IEG responses are shaped by stimulus duration and the identity of the IEG and timepoint at which expression is measured. This work lays the foundation for further precise, cell type-resolved investigation into how the brain processes social information.