蛋白激酶A介导的大脑神经递质“半球偏侧化”改变在母体慢性应激所致子代行为异常的作用机制研究

In our preliminary studies, we found the maternal chronic stress would lead to the decreased coping ability of offspring rats in the forced swimming test and decreased learning and memory ability of offspring rats in the water maze test. We also found the maternal stress would affect the protein kinase A signaling pathway in the brain, the dopamine system in the prefrontal cortex and the serotonin system in the hippocampus of offspring rats. The above findings occurred in the right hemisphere, which was the "hemispheric lateralization" change. Therefore, we hypothesized that the mechanism of abnormal behavior in the offspring which was due to the maternal chronic stress may be related to the protein kinase A signal-mediated "hemispheric lateralization" change in the brain. In order to demonstrate the assumptions, the project will use the chronic unpredictable stress maternal rat model, and apply miniature high-field magnetic resonance imaging, magnetic resonance spectroscopy, neural behavioral tests and molecular biology techniques to determine the influence of maternal chronic stress on the behavior of offspring, and investigate the protein kinase A signal-mediated "hemispheric lateralization" change in the metabolism of neurotransmitters and the distribution of neuronal protein. The project will explore the mechanism of abnormal behavior in the offspring which was due to the maternal chronic stress from the interaction between genetic, environmental, brain function and the behavior of offspring. The findings of this project will expand understanding of the psychological and behavioral disorders in children.

本项目在前期研究中发现，母鼠慢性应激会导致子鼠在强迫游泳试验中的"应激应对能力"和在水迷宫试验中的"学习记忆能力"下降，并能检测到子鼠大脑的蛋白激酶A信号通路、前额叶皮质的多巴胺系统和海马的5-羟色胺系统出现异常，且这些改变主要发生在右侧大脑半球，存在"半球偏侧化"改变。因此，我们提出假设：母体慢性应激所致子代行为异常的机制可能与蛋白激酶A信号介导的大脑神经递质"半球偏侧化"改变有关。为论证该假设，本项目将在慢性不可预见性应激母鼠模型上，采用微型高场磁共振成像系统及磁共振波谱分析，并运用神经行为学试验、分子生物学技术，明确母体慢性应激对子代行为异常发生的影响，探寻慢性应激子代大脑蛋白激酶A介导的神经递质代谢以及神经蛋白分布和含量的"半球偏侧化"改变，并从遗传、环境、脑功能与行为之间相互作用的角度探讨母体慢性应激所致子代行为异常的发生机制。研究结果可拓展对儿童心理行为疾病的认识。

本项目在前期研究中发现，母鼠慢性应激会导致子鼠在强迫游泳试验中的应激应对能力下降，并能检测到子鼠右侧大脑蛋白激酶A（PKA）信号通路出现异常。因此，我们假设：母体慢性应激所致子代行为异常的机制可能与PKA介导的大脑神经递质"半球偏侧化"改变有关。本项目采用磁共振波谱（MRS）分析及其它分子生物学技术，探讨母体妊娠前慢性应激对子鼠在面对急性应激后大脑海马中的氨基酸类神经递质代谢产生的影响及其作用机制。研究结果发现：（1）母体妊娠前慢性应激所产子鼠在面对急性应激时，脑内谷氨酸（Glu）较正常组子鼠明显增加[F(7,136)=26.44,p＜0.001]；（2）脑内积聚的Glu在谷氨酰胺合成酶（PAG）的作用下合成谷氨酰胺（Gln）。应激组雄性子代右侧海马中的PAG未随着Glu的增加出现上调，破坏了Glu/Gln的平衡[F(7,136)=3.41,p=0.002]，因此，雄性子鼠在旷场试验中表现出焦虑样行为；（3）脑内的Glu还可通过谷氨酸脱羧酶(GAD)生成γ－氨基丁酸(GABA)，而GABA又可通过γ－氨基丁酸转氨酶(GABA-T)生成Glu。面对急性应激时，应激组子鼠乳酸堆积[F(3,68)=9.59，p＜0.001]，pH值较正常组子鼠下降[F(3,68)=18.85，p＜0.001]，因此GAD活性比正常组子鼠升高[F(7,136)=155.08,p＜0.001]，而GABA-T活性较正常组子鼠降低[F(7,136)=140.28,p＜0.001]，导致GABA较正常组子鼠增加[F(7,136)=52.64,p＜0.001]，这种改变在应激组雌性子鼠右侧海马明显，因此，应激组雌性子鼠在强迫游泳中表现出抑郁样行为；（4）酸中毒会引起Ca2+内流，血Ca2+下降[F(3,68)=23.8，p＜0.001]，负反馈抑制GluR[F(7,136)=60.82,p＜0.001]，使p-PKA减少[F(7,136)=61.76,p＜0.001]，引起p-CREB下降[F(7,136)=45.9,p＜0.001]，最终导致PKA下游的BDNF和TrkB表达下降。研究结果提示母体妊娠前慢性应激可能会增加子代在青春前期罹患急性应激所致行为异常的风险，且这种影响存在“半球偏侧化效应”和“性别差异”。本结果可拓展对急性应激所致儿童心理行为疾病的认识，有利于早期发现和早期干预该类疾病。