512. Oxytocin-receptor neurons in mPFC in stress-induced anxiety behaviors

Abstract

Abstract Background The hypothalamic neuropeptide oxytocin (Oxt) modulates diverse social and anxiety-related behaviors. These biological actions are mediated by specific Oxt receptors (Oxtr), members of the G protein-coupled receptor (GPCR) family. While the medial prefrontal cortex (mPFC) is fundamental in regulating anxiety behavior, the mechanisms by which Oxtr-expressing (Oxtr+) neurons in the mPFC orchestrate anxiety-related phenotypes through downstream molecular signaling pathways and neural circuit mechanisms remain poorly understood. Aims & Objectives To delineate the pathways downstream of the Oxtr in mPFC in controlling anxiety-like behavior. Method We first employed flow cytometry to identify cell-type-specific alterations in Oxtr expression induced by stress exposure. To define downstream neural substrates, we performed retrograde viral tracing from Oxtr-expressing mPFC neurons to map efferent projections and combined chemogenetic approaches with behavioral assays to functionally validate the roles of mPFC-BLA (basolateral amygdala) and mPFC-aIC (anterior insular cortex) circuits in anxiety-like behaviors. Subsequently, we assessed stress-induced changes in mitochondrial pathways, ultrastructure, and function through integrated analyses, including transcriptomic profiling (identifying OXTR-dependent mitochondrial gene networks), electron microscopy (visualizing mitochondrial morphology) and seahorse extracellular flux analysis (quantifying mitochondrial respiration and ATP production). Results Here, through flow cytometry, AAV-based manipulations, transcriptomic profiling, electron microscopy, seahorse extracellular flux analysis and fiber photometry, we identified that Oxtr in the mPFC modulates anxiety-like behaviors primarily by impairing mitochondrial ATP production and Ornithine metabolomics pathway. Critically, exogenous ATP administration, overexpression of mfn2 or significantly rescues anxiety phenotypes in Oxtr-knockdown models, confirming the causal role of energy metabolism dysregulation. Combining HSV-based anterograde viral tracing with chemogenetic manipulation and fiber photometry, we dissected the functional divergence of OXTR+ neuron subpopulations in the mPFC. Layer 5 neurons projecting to the anterior insular cortex (aIC) exert anxiolytic effects, layer 2/3 neurons projecting to the basolateral amygdala (BLA) promote anxiogenic responses. During acute restraint stress, these two populations exhibit opposing activity patterns. Electrophysiological evidence further reveals that anxiolytic circuits (layer 5-aIC) directly inhibit anxiogenic pathways (layer 2/3-BLA) via GABAergic synaptic transmission. Discussion & Conclusions Collectively, these results elucidate a multi-tiered mechanistic framework linking Oxtr signaling: mitochondrial-ornithine axis dysfunction (molecular level) and cross-circuit modulation between mPFC-aIC and mPFC-BLA pathways (circuit level) in the generation of anxiety-like behaviors. Such antagonistic interplay may represent a central principle in how the mPFC orchestrates complex behaviors through top-down control.