Abstract Chronic social stress (CSS) is a significant public health challenge that negatively impacts behavior and immune function through brain‐spleen interactions. Oxytocin (OT), a neuropeptide critical for social behavior and immune regulation, is upregulated during CSS, though its underlying mechanisms remain unclear. This study investigates the role of OT in splenic immune modulation using a murine model of CSS. Behavioral evaluations, serum oxytocin quantification, and splenic immunophenotypic analysis were performed. Splenic denervation confirmed OT’s neuromodulatory role, whereas OTR antagonism revealed its endocrine function. CSS‐induced OT elevation was associated with immunosuppression, characterized by increased Foxp3⁺ regulatory T cells and reduced CD4⁺ T and CD19⁺ B cells. OT also modulated macrophage polarization, inhibiting M1‐like (pro‐inflammatory) and enhancing M2‐like (anti‐inflammatory) phenotypes. Denervation or pharmacological blockade of OT signaling partly reversed CSS‐induced splenic immunosuppression but adversely affected survival in CSS‐exposed mice. Additionally, denervation or OTR antagonism reduced the mice's response to social defeat, as shown by decreased social avoidance behavior. These findings suggest that OT‐mediated immunosuppression likely represents a compensatory mechanism in response to chronic social stress. Targeting the OT–immune axis could offer innovative therapeutic approaches for stress‐associated disorders by restoring immune homeostasis while maintaining behavioral integrity. Keywords: brain‐spleen interaction, chronic social stress, immune regulation, macrophage polarization, oxytocin __________________________________________________________________ Chronic social stress elevates oxytocin levels, leading to immunosuppression by enhancing regulatory T‐cell activity and promoting anti‐inflammatory macrophage polarization in the spleen. Blocking oxytocin signaling reverses these effects but worsens stress‐induced behavioral disorders and reduces survival rates. This highlights oxytocin's dual role in balancing immune responses and protecting against stress‐related disorders via neuroendocrine pathways. graphic file with name ADVS-12-2500849-g007.jpg 1. Introduction In contemporary society, individuals are increasingly subjected to a range of social pressures. The epidemiological link between elevated stress levels and disease incidence underscores the intricate interplay between mental and physiological health within modern medicine.^[ [34]^1 , [35]^2 ^] Prolonged or recurrent adverse social experiences, such as social exclusion, failure, and perceived threats, can result in Chronic Social Stress (CSS).^[ [36]^3 , [37]^4 ^] In this study, we employed a standardized CSS model using large and aggressive Kunming mice (KM) as the aggressors against C57BL/6 mice. Mice in the CSS group were subjected to 10 min of social aggression daily, followed by confinement in an isolation cage where they had only visual, olfactory, and auditory contact with the aggressor mice for 24 h. This prolonged psychological stress alters the bidirectional communication between the brain and the immune system, contributing to immune dysregulation in many stress‐related psychiatric disorders.^[ [38]^5 , [39]^6 , [40]^7 ^] Oxytocin (OT), a critical neuropeptide, has garnered significant attention for its roles in modulating social behavior, stress responses, and immune functions.^[ [41]^8 , [42]^9 ^] Although it is established that OT neurons project extensively throughout the brain and that Oxytocin Receptors (OTRs) are widely distributed in peripheral organs,^[ [43]^10 , [44]^11 , [45]^12 ^] the extent to which brain activity directly modulates immune response via OT neuronal projections remains unclear. The spleen, a major lymphatic organ involved in immune responses to psychological stress, plays a key role in monocytosis through granulocyte aggregation and ectopic bone marrow formation, leading to elevated circulating monocyte levels.^[ [46]^13 , [47]^14 ^] Research into the brain‐spleen axis has identified it as a direct neural pathway regulating cognition and emotion through peripheral immune response during stress. Studies have shown that activation of CRH^+ neurons in the PVN and central amygdala (CeA) can appropriately increase the production of splenic plasma cells.^[ [48]^14 , [49]^15 ^] Conversely, disruption of these neural connections reduces plasma cell numbers by lowering glucocorticoid production via the hypothalamic‐pituitary‐adrenal (HPA) axis.^[ [50]^16 ^] This finding suggests that immune cells, distributed throughout various tissues, are dynamically regulated by nervous system activity. Investigating potential interactions between OT neurons and the brain‐spleen axis could provide novel insights into the immunoregulatory role of oxytocin. Research on OT in CSS has primarily focused on its effects on maladaptive behaviors in mice, such as social avoidance, depression, and anxiety, which are associated with dysregulated OT expression.^[ [51]^17 , [52]^18 , [53]^19 , [54]^20 ^] OT, acting through OTR, modulates synaptic plasticity and reduces central sensitization in chronic migraine models.^[ [55]^21 , [56]^22 ^] Additionally, OT has demonstrated immunomodulatory effects, including inhibition of lipopolysaccharide (LPS)‐induced microglial activation and reduced release of pro‐inflammatory factors.^[ [57]^23 ^] It also enhances CD8^+ T cell efficacy against tumor cells.^[ [58]^24 , [59]^25 ^] However, OT and OTR functions vary across brain regions and cell types, displaying a high degree of spatial and temporal specificity.^[ [60]^26 ^] Investigating the roles of OT and OTR in the spleen is crucial for understanding the complex functions of oxytocin in immune regulation. 2. Results 2.1. Behavioral and Physiological Impact of Chronic Social Stress in Mice To investigate the effects of CSS on mice, we established a CSS model (Figure [61]1A). Body weight in the Stress group decreased significantly compared to pre‐CSS levels, while no significant changes were observed in the Control group (Figure [62]1B). Additionally, Stress group mice exhibited a significantly reduced survival rate (Figure [63]1C). H&E staining at the end of the CSS period showed no structural abnormalities in the spleen (Figure [64]S1, Supporting Information). Behavioral responses were assessed using the Open Field Test (OFT) and the Three‐Chamber Social Test (TST). In the OFT, stressed mice demonstrated reduced activity in the central zone, increased freezing duration, and decreased overall locomotion, indicating diminished spontaneous exploration and elevated anxiety levels (Figure [65]1D–G). The TST indicated that CSS did not impair social ability, preference, or memory, as Stress mice interacted more with other mice than with empty cages, preferred strangers over familiar mice, and chose familiar over attacker mice (Figure [66]1H–J). Notably, the Control mice, lacking CSS exposure, showed social avoidance toward aggressive mice likely due to their physical presence rather than fearful memories. In summary, CSS significantly increased anxiety levels and reduced spontaneous exploratory behavior in mice, while preserving their social abilities, preferences, and memory.