Sex-dependent effects of angiotensin type 2 receptor expressing medial prefrontal cortex (mPFC) interneurons in fear extinction learning

Background The renin-angiotensin system (RAS) has been identified as a potential therapeutic target for PTSD, though its mechanisms are not well understood. Brain angiotensin type 2 receptors (AT2Rs) are a subtype of angiotensin II receptors located in stress and anxiety-related regions, including the medial prefrontal cortex (mPFC), but their function and mechanism in the mPFC remain unexplored. We therefore used a combination of imaging, cre/lox, and behavioral methods to investigate mPFC-AT2R-expressing neuron involvement in fear learning. Methods To characterize mPFC-AT2R-expressing neurons in the mPFC, AT2R-Cre/td-Tomato male and female mice were used for immunohistochemistry (IHC). mPFC brain sections were stained with glutamatergic or interneuron markers, and density of AT2R+ cells and colocalization with each marker was quantified. To assess fear-related behaviors in AT2R-flox mice, we selectively deleted AT2R from mPFC neurons using an AAV-Cre virus. Mice then underwent Pavlovian auditory fear conditioning, approach/avoidance, and locomotion testing. Results IHC results revealed that AT2R is densely expressed in the mPFC. Furthermore, AT2R is primarily expressed in somatostatin interneurons in females but not males. Following fear conditioning, mPFC-AT2R deletion impaired extinction in female but not male mice. Locomotion was unaltered by mPFC-AT2R deletion in males or females, while AT2R-deleted females had increased exploratory behavior. Conclusion These results lend support for mPFC-AT2R+ neurons as a novel subgroup of somatostatin interneurons that influence fear extinction in a sex-dependent manner. This furthers underscores the role of mPFC in top-down regulation and a unique role for peptidergic (ie., angiotensin) mPFC regulation of fear and sex differences. ### Competing Interest Statement The authors have declared no competing interest.