Data_Sheet_1_Chronic Hippocampal Abnormalities and Blunted HPA Axis in an Animal Model of Repeated Unpredictable Stress.DOCX

Incidence of post-traumatic stress disorder (PTSD) ranges from 3 to 30% in individuals exposed to traumatic events, with the highest prevalence in groups exposed to combat, torture, or rape. To date, only a few FDA approved drugs are available to treat PTSD, which only offer symptomatic relief and variable efficacy. There is, therefore, an urgent need to explore new concepts regarding the biological responses causing PTSD. Animal models are an appropriate platform for conducting such studies. Herein, we examined the chronic behavioral and neurobiological effects of repeated unpredictable stress (RUS) in a mouse model. 12 weeks-old C57BL/6J male mice were exposed to a 21-day RUS paradigm consisting of exposures to a predator odor (TMT) whilst under restraint, unstable social housing, inescapable footshocks and social isolation. Validity of the model was assessed by comprehensive examination of behavioral outcomes at an acute timepoint, 3 and 6 months post-RUS; and molecular profiling was also conducted on brain and plasma samples at the acute and 6 months timepoints. Stressed mice demonstrated recall of traumatic memories, passive stress coping behavior, acute anxiety, and weight gain deficits when compared to control mice. Immunoblotting of amygdala lysates showed a dysregulation in the p75NTR/ProBDNF, and glutamatergic signaling in stressed mice at the acute timepoint. At 6 months after RUS, stressed mice had lower plasma corticosterone, reduced hippocampal CA1 volume and reduced brain-derived neurotrophic factor levels. In addition, glucocorticoid regulatory protein FKBP5 was downregulated in the hypothalamus of stressed mice at the same timepoint, together implicating an impaired hypothalamus-pituitary-adrenal-axis. Our model demonstrates chronic behavioral and neurobiological outcomes consistent with those reported in human PTSD cases and thus presents a platform through which to understand the neurobiology of stress and explore new therapeutic interventions.

创伤后应激障碍（post-traumatic stress disorder, PTSD）在暴露于创伤事件的个体中发病率为3%至30%，其中暴露于战斗、酷刑或强奸的群体患病率最高。迄今为止，仅有少数经美国食品药品监督管理局（Food and Drug Administration, FDA）批准的药物可用于治疗PTSD，此类药物仅能缓解症状，且疗效参差不齐。因此，亟需探索与PTSD发生相关的生物学应答新机制。动物模型是开展此类研究的合适平台。

本研究在小鼠模型中探究了重复不可预测应激（repeated unpredictable stress, RUS）的长期行为学与神经生物学效应。实验选用12周龄的C57BL/6J雄性小鼠，采用为期21天的RUS造模范式，造模应激源包括束缚状态下暴露于捕食者气味（TMT）、不稳定群居环境、不可逃避足底电击以及社会孤立。

通过在造模后急性时点、3个月及6个月时全面评估行为学结局，以验证该模型的有效性；同时在急性时点与6个月时点采集脑组织与血浆样本进行分子谱分析。

与对照组小鼠相比，应激组小鼠表现出创伤记忆唤醒、被动应激应对行为、急性焦虑以及体重增长缺陷。对杏仁核裂解物进行免疫印迹分析发现，急性时点应激组小鼠的p75神经营养因子受体（p75NTR）/前脑源性神经营养因子（ProBDNF）信号通路以及谷氨酸能信号通路均存在失调。

造模后6个月，应激组小鼠血浆皮质酮水平降低、海马CA1区体积减小以及脑源性神经营养因子（brain-derived neurotrophic factor, BDNF）水平下降。此外，同期应激组小鼠下丘脑内糖皮质激素调节蛋白FKBP5表达下调，这提示下丘脑-垂体-肾上腺轴（hypothalamus-pituitary-adrenal-axis, HPA轴）功能受损。

本模型所呈现的慢性行为学与神经生物学结局与人类PTSD患者的已有报道相符，因此可作为探究应激神经生物学机制、探索新型治疗干预手段的研究平台。