Data_Sheet_1_GluN2A/ERK/CREB Signaling Pathway Involved in Electroacupuncture Regulating Hypothalamic-Pituitary-Adrenal Axis Hyperactivity.docx

The hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis caused by stress will inevitably disrupt the homeostasis of the neuroendocrine system and damage physiological functions. It has been demonstrated that electroacupuncture (EA) can modulate HPA axis hyperactivity during the perioperative period. As the initiating factor of the HPA axis, hypothalamic corticotrophin-releasing hormone (CRH) is the critical molecule affected by EA. However, the mechanism by which EA reduces CRH synthesis and secretion remains unclear. Activated N-methyl-D-aspartate receptor (NMDAR) has been linked to over-secretion of hypothalamic CRH induced by stress. To determine whether NMDAR is involved in EA regulating the over-expression of CRH, a surgical model of partial hepatectomy (HT) was established in our experiment. The effect of EA on hypothalamic NMDAR expression in HT mice was examined. Then, we investigated whether the extracellular regulated protein kinases (ERK)/cyclic adenosine monophosphate response element-binding protein (CREB) signaling pathway mediated by NMDAR was involved in EA regulating HPA axis hyperactivity. It was found that surgery enhanced the expression of hypothalamic CRH and caused HPA axis hyperactivity. Intriguingly, EA effectively suppressed the expression of CRH and decreased the activation of GluN2A (NMDAR subunit), ERK, and CREB in HT mice. GluN2A, ERK, and CREB antagonists had similar effects on normalizing the expression of CRH and HPA axis function compared with EA. Our findings suggested that surgery enhanced the activation of the hypothalamic GluN2A/ERK/CREB signaling pathway, thus promoting the synthesis and secretion of CRH. EA suppressed the phosphorylation of GluN2A, ERK, and CREB in mice that had undergone surgery, indicating that the GluN2A/ERK/CREB signaling pathway was involved in EA alleviating HPA axis hyperactivity.

应激所致下丘脑-垂体-肾上腺（hypothalamic-pituitary-adrenal, HPA）轴功能亢进，势必干扰神经内分泌系统稳态并损害生理功能。已有研究证实，电针（electroacupuncture, EA）可在围手术期调节HPA轴亢进状态。作为HPA轴的启动因子，下丘脑促肾上腺皮质激素释放激素（hypothalamic corticotrophin-releasing hormone, CRH）是电针调控的关键分子。然而，电针下调CRH合成与分泌的具体机制仍未阐明。活化的N-甲基-D-天冬氨酸受体（N-methyl-D-aspartate receptor, NMDAR）与应激诱导的下丘脑CRH过度分泌密切相关。为明确NMDAR是否参与电针调控CRH的过度表达，本实验构建了部分肝切除术（partial hepatectomy, HT）小鼠模型。本研究检测了电针对部分肝切除术小鼠下丘脑NMDAR表达的影响，随后进一步探究了NMDAR介导的细胞外调节蛋白激酶（extracellular regulated protein kinases, ERK）/环磷酸腺苷反应元件结合蛋白（cyclic adenosine monophosphate response element-binding protein, CREB）信号通路是否参与电针调控HPA轴亢进的过程。结果显示，手术可上调下丘脑CRH的表达并引发HPA轴功能亢进。值得注意的是，电针可有效抑制部分肝切除术小鼠的CRH表达，并降低GluN2A（NMDAR亚基）、ERK及CREB的活化水平。GluN2A、ERK及CREB拮抗剂均可起到与电针相似的效果，使CRH表达及HPA轴功能恢复正常。本研究结果表明，手术可激活下丘脑GluN2A/ERK/CREB信号通路，进而促进CRH的合成与分泌。电针可抑制手术小鼠体内GluN2A、ERK及CREB的磷酸化水平，提示GluN2A/ERK/CREB信号通路参与了电针缓解HPA轴功能亢进的过程。