Supplementary Material for: Involvement of the AVP/OT system and lateral septum in the modulation of anxiety/depression caused by 14-week and 20-week social isolation

Introduction: Chronic social isolation (CSI) stress leads to numerous maladaptive changes in physiology and psychology, however, very little is known about the effects of longer time-scale CSI and there are divergent views regarding the underlying mechanisms. This study aimed to elucidate the common neuroendocrine mechanisms underlying the maladaptive changes caused by 14-week (14wk-) and 20-week (20wk-) CSI. Methods: We investigated the impacts of 14wk- and 20wk- CSI on anxiety/depression-like behaviors in male C57BL/6N mice with classical behavioral tests, and the immunofluorescence method was used for quantification of the arginine vasopressin (AVP)/Oxytocin (OT) positive neurons in the PVN and screening out the differential activation brain regions (DABrs) on exposure of tail suspension. The expression of Avpr1a and Oxtr in the lateral septum dorsal (LSD) was assessed by quantitative RT-PCR (qPCR), and the function of LSD Avpr1a+/+ neurons in emotion regulation was verified with pharmacological approaches. The concentration of AVP/OT in plasma was examined with the Enzyme-linked immunosorbent assay (ELISA). Results: 14wk- and 20wk- CSI increased anxiety/depression-like behaviors and altered levels of exploratory locomotion in opposite directions, which are likely due to an imbalance of the AVP/OT system within the PVN and peripheral plasma, differential activation of LSD and thalamic brain regions, and abnormal expression of Avpr1a in LSD. Pharmacological results demonstrated that Avpr1a receptor in the LSD were involved in emotion regulation. Conclusion: This study suggests that the imbalance of the AVP/OT system and the dysfunction of Avpr1a+/+ neurons in the LSD were engaged in common neuroendocrinology mechanisms for anxiety/depression induced by long-term CSI.