丰富环境通过调控前扣带回皮层锥体神经元兴奋性缓解束缚应激诱导的小鼠焦虑样行为

doi:10.12122/j.issn.1673-4254.2025.05.08

南方医科大学学报 ›› 2025, Vol. 45 ›› Issue (5): 962-968.doi: 10.12122/j.issn.1673-4254.2025.05.08

丰富环境通过调控前扣带回皮层锥体神经元兴奋性缓解束缚应激诱导的小鼠焦虑样行为

陈昌锋¹(), 方勤¹, 高银环¹, 王烈成¹, 陈磊²()

^1.安徽医科大学基础医学院，安徽合肥 230032
^2.安徽中医药大学第一附属医院药学部，安徽合肥 230032

收稿日期:2024-11-28 出版日期:2025-05-20 发布日期:2025-05-23
通讯作者: 陈磊 E-mail:chenchangfeng89@126.com;azyfychenlei@ahtcm.edu.cn
作者简介:陈昌锋，硕士，实验师，E-mail: chenchangfeng89@126.com
基金资助:
安徽省自然科学基金(2408085MH189);安徽省高等学校科学研究项目(2024AH051017);安徽省高校中青年教师培养行动项目(YQYB2024031);安徽中医药大学第一附属医院临床科学研究项目(2021yfylc14);安徽省生物医学基础与应用科技公共服务平台2023年大型仪器设备开放基金(2023-04);安徽省级大学生创新创业训练计划项目(S202410366078);安徽医科大学科研平台基地升级计划(2021xkjT048)

Enriched environment reduces pyramidal neuron excitability in the anterior cingulate cortex to alleviate restraint stress-induced anxiety-like behaviors in mice

Changfeng CHEN¹(), Qin FANG¹, Yinhuan GAO¹, Liecheng WANG¹, Lei CHEN²()

^1.School of Basic Medicine Sciences, Anhui Medical University, Hefei 230032, China
^2.Department of Pharmay, First Affiliated Hospital of Anhui University of Chinese Medicine, Hefei 230032, China

Received:2024-11-28 Online:2025-05-20 Published:2025-05-23
Contact: Lei CHEN E-mail:chenchangfeng89@126.com;azyfychenlei@ahtcm.edu.cn

摘要/Abstract

摘要：

目的探究前扣带回皮层锥体神经元调控丰富环境缓解束缚应激致焦虑样行为的机制。方法 C57BL/6J小鼠分为3组：正常组（CON）、束缚组（RS）和丰富环境组（RS+EE），8只/组。CON组正常实验室环境饲养，RS组给与2 h/d连续3 d的束缚应激，RS+EE组在给与2 h/d连续3 d的束缚应激的同时EE下饲养。利用高架十字迷宫实验和旷场实验评估小鼠焦虑样水平，通过免疫荧光实验检测小鼠ACC区c-Fos的变化，采用膜片钳实验检测小鼠Pyn^ACC兴奋性的变化，进一步检测Pyn^ACC的mEPSC和mIPSC的发放频率和幅度，以评估突触传递的变化。结果高架十字迷宫实验中，与CON组比较，RS组小鼠在开臂的停留时间减少（P<0.01），进入开臂的次数减少（P<0.05）；与RS组比较，RS+EE组小鼠在开臂的停留时间增加（P<0.01），进入开臂的次数增加（P<0.05）；旷场实验中，与CON组比较，RS组小鼠在中心区域的移动时间减少（P<0.01），进入中心区域的次数减少（P<0.01）；与RS组比较，RS+EE组小鼠在中心区域的移动时间增加（P<0.01），进入中心区域的次数增加（P<0.05）。免疫荧光实验中，与CON组比较，RS组小鼠ACC区c-Fos表达增加（P<0.001）；与RS组比较，RS+EE组小鼠前扣带回皮层c-Fos表达减少（P<0.01）。膜片钳实验中，与CON组比较，RS组小鼠Pyn^ACC动作电位发放增多（P<0.01）；与RS组比较，RS+EE组小鼠Pyn^ACC动作电位发放减少（P<0.05）；与CON组比较，RS组小鼠Pyn^ACC的mEPSC频率增加（P<0.05）；与RS组比较，RS+EE组小鼠Pyn^ACC的mEPSC频率降低（P<0.05），而幅度均没有变化；mIPSC的频率和幅度均没有变化。结论丰富环境降低前扣带回皮层锥体神经元兴奋性缓解束缚应激致焦虑样行为。

关键词: 丰富环境, 前扣带回皮层, 锥体神经元, 焦虑, 兴奋性

Abstract:

Objective To investigate the mechanism by which the pyramidal neurons of the anterior cingulate cortex (ACC) modulate the effects of enriched environment (EE) for relieving anxiety-like behaviors in mice. Methods C57BL/6J mice were randomly divided into control group, restraint stress (RS) group, and RS+EE group (n=8). The mice in the latter two groups were subjected to RS for 2 h daily for 3 days, and those in RS+EE group were housed in an EE during modeling. Anxiety-like behaviors of the mice were evaluated using the elevated plus-maze tests (EPM) and open field test (OFT). Changes in c-Fos expression in the ACC of the mice were detected with immunofluorescence assay, and pyramidal neuron excitability in the ACC (Pyn^ACC) was measured using patch-clamp technique. The miniature excitatory and inhibitory postsynaptic currents (mEPSC and mIPSC, respectively) were analyzed to assess synaptic transmission changes. Results Behavioral tests showed obvious anxiety-like behaviors in RS mice, and such behavioral changes were significantly improved in RS+EE mice. Immunofluorescence staining revealed significantly increased c-Fos expression in the ACC in RS mice but lowered c-Fos expression in RS+EE group. Compared with the control mice, the RS mice showed increased action potential firing rate of Pyn^ACC, which was significantly reduced in RS+EE group. Compared with the RS mice, the RS+EE mice showed also decreased frequency of mEPSCs of Pyn^ACC, but the amplitude exhibited no significant changes. No obvious changes in the frequency or amplitude of mIPSCs were observed in RS+EE mice. Conclusion EE reduces excitability of Pyn^ACC to alleviate anxiety-like behaviors induced by RS in mice.

Key words: enriched environment, anterior cingulate cortex, pyramidal neuron, anxiety, excitability

陈昌锋, 方勤, 高银环, 王烈成, 陈磊. 丰富环境通过调控前扣带回皮层锥体神经元兴奋性缓解束缚应激诱导的小鼠焦虑样行为[J]. 南方医科大学学报, 2025, 45(5): 962-968.

Changfeng CHEN, Qin FANG, Yinhuan GAO, Liecheng WANG, Lei CHEN. Enriched environment reduces pyramidal neuron excitability in the anterior cingulate cortex to alleviate restraint stress-induced anxiety-like behaviors in mice[J]. Journal of Southern Medical University, 2025, 45(5): 962-968.

图/表 4

图1 EE可改善RS诱导的焦虑样行为

Fig.1 Enriched environment (EE) alleviates anxiety-like behaviors induced by restraint stress (RS) in mice. A: Heatmaps of 5-minute walking paths in the 3 groups in elevated plus-maze (EPM) tests. B: Time spent in open arms. C: Entries in open arms. D: Total distance in EPM tests. E: Heatmaps of 5-minute walking paths in the 3 groups in open field test (OFT). F: Entries in the central area. G: Time in the central area in OFT. *P<0.05, **P<0.01 (n=8).

图2 EE可降低RS诱导的ACC区c-Fos高表达

Fig.2 EE prevents the high c-Fos expression of the ACC induced by RS. A: The location of ACC in the brain atlas (Bregma 0.02 mm). B: The original diagram of c-Fos expression in ACC from each group. Scale bar=100 μm. C: Statistical graphs of c-Fos expression in each group. CON: n=3, RS: n=4, RS+EE: n=4. **P<0.01, ***P<0.001.

图3 EE可降低RS诱导的PynACC动作电位发放增多

Fig.3 EE attenuates RS-induced increases in action potentials of PynACC. A: Representative image of a PynACC under whole-cell recording (Scale bar=100 μm). B: Groups of typical sequential spikes. C: Quantification of the numbers of spikes with different stimuli. CON: 10 cells from 4 mice; RS: 11 cells from 4 mice; RS+EE: 10 cells from 3 mice. *P<0.05, **P<0.01.

图4 EE可降低RS诱导的PynACC mEPSC的发放频率增加

Fig.4 EE attenuates RS-induced increase of mEPSC frequency of PynACC. A: Representative traces of mEPSCs of PynACC. B: Cumulative plots of inter-event interval of mEPSC. C: Statistical results of mEPSC frequency. D: Cumulative plots of amplitude of mEPSC. E: Statistical results of mEPSC amplitude. CON: 12 cells from 4 mice; RS: 13 cells from 3 mice; RS+EE: 14 cells from 3 mice. F: Representative traces of mIPSCs of PynACC. G: Cumulative plots of inter-event interval of mIPSC. H: Statistical results of mIPSC frequency. I: Cumulative plots of amplitude of mIPSC. J: Statistical results of mIPSC amplitude. CON: 12 cells from 4 mice; RS: 11 cells from 4 mice; RS+EE: 11 cells from 3 mice. *P<0.05.

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丰富环境通过调控前扣带回皮层锥体神经元兴奋性缓解束缚应激诱导的小鼠焦虑样行为

Enriched environment reduces pyramidal neuron excitability in the anterior cingulate cortex to alleviate restraint stress-induced anxiety-like behaviors in mice

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