Subarachnoid Hemorrhage Induces mPFC CRF Neuronal Dysfunction Leading to Anxiety and Depression in Male Mice

BACKGROUND: Anxiety and depression are common neuropsychiatric sequelae after subarachnoid hemorrhage (SAH), yet the underlying neural mechanisms remain poorly understood. The CRF (corticotropin-releasing factor) system in the medial prefrontal cortex (mPFC) is critical for emotional regulation and stress adaptation; however, its involvement in SAH-induced affective disorders has not been defined. METHODS: SAH was induced by endovascular perforation. Anxiety and depressive-like behaviors and mPFC electrophysiological activity were assessed using behavioral paradigms and in vivo recordings. Functional alterations of mPFC CRF neurons were evaluated by immunohistochemistry, fiber photometry, and patch-clamp recordings. Chemogenetic and optogenetic approaches were used to selectively manipulate CRF neurons. Neuronal tracing, pathway-specific optogenetics, and adeno-associated virus-mediated ablation of mPFC CRF neurons were used to dissect circuit mechanisms. Proteomics, immunogold electron microscopy, immunohistochemistry, and adeno-associated virus-based modulation of SYT (synaptotagmin) 9 were performed to investigate molecular contributors to mPFC CRF neuron dysfunction. RESULTS: SAH induced pronounced anxiety and depressive-like behaviors. mPFC CRF neurons exhibited structural disruptions, reduced calcium dynamics, and impaired excitatory synaptic transmission after SAH. Chemogenetic or optogenetic activation of mPFC CRF neurons ameliorated these behavioral deficits, whereas local CRF supplementation in the mPFC was ineffective. Approximately 35% of mPFC CRF neurons received basolateral amygdala inputs, and optogenetic activation of the basolateral amygdala→mPFC pathway improved behavioral outcomes; these effects were abolished by CRF neuron ablation. Proteomic analysis identified SYT9 as one of the most significantly downregulated proteins, predominantly enriched in CRF neurons. SYT9 overexpression restored CRF neuron activity and mitigated SAH-induced behavioral abnormalities. CONCLUSIONS: Dysfunction of mPFC CRF neurons is a key contributor to anxiety and depression after SAH. Basolateral amygdala inputs modulate mPFC CRF neuron activity, and SYT9 downregulation represents a critical molecular mechanism underlying their impairment. These findings identify mPFC CRF neuronal dysfunction as a core pathological feature of SAH-induced affective disorders and provide mechanistic insight into potential therapeutic targets.