突触外及周围GABAA受体和神经甾体对雌性小鼠青春期抑郁行为的影响

The depression in pubertal females is characteristic with high incidence, suicide rate and severe symptoms, but its pathogenesis is still complicated and many factors are involved in it. Monoamine hypothesis has been considered as the pathogenesis of depression for a long time. But monoaminergic antidepressant drugs required chronic administration to evoke an antidepressant-like effect in humans for several weeks. Furthermore, in nearly 40% of patients the antidepressant therapy presently conducted is not effective, thus confirming the limited role of monoamines in depression. Recent studies have shown that it is related to GABA /glutamate imbalance in central nervous system. NMDA receptor antagonist ketamine injected intravenously can significantly alleviate the symptoms of depression in a few hours, especially for severe refractory depression as well as most of monoamine ineffective antidepressant drugs. Research results showed that mixed GABAmimetics can also alleviate the depression symptoms. The dysfunction of GABAA receptor subunits or subtypes, such as α2, α3, γ which locate inside synapses may lead to depression, but the role of peri- or extra-synaptic subtypes of GABAA receptors was unclear. We discovered earlier that the anxiety in pubertal females was induced by the high expression of GABAA α4βδ receptor in peri- or extra-synapse of the dendritic spines in hippocampal pyramidal cells in puberty, and 3α,5α(β) - allopregnanolone (THP), a neurosteroid and the strongest modulator of GABAA receptor changed the direction of GABA tonic current produced by GABAA α4βδ receptor. Anxiety is the early symptoms of depression, and both may have same mechanism of occurrence. In this study, we will replicate female depression model in pubertal mice with both wild and δ subunit knockout types, and will use the advanced techniques of two-photon imaging, two-photon shadow patch clamp, two-photon uncage of two-photon fluorescence microscopy, immuno-electron microscopy, Western blot, and behavioral observation methods, and observe the expression of GABAA receptors in peri- or extra-synapse of the dendritic spines in the cells of hippocampal pyramidal cells, GABA tonic current produced by the receptors and the impact of the GABAA receptors on excitatory currents and NMDA current and the density of dendritic spines and synapses，as well as severity of depression in those female pubertal mice. We also observe the effect of THP on the GABAA receptors in peri- or extra-synapse of the dendritic spines in the cells of hippocampal pyramidal cells and the change of the direction of GABA tonic current, excitatory currents, NMDA current, density of dendritic spines and synapses, and severity of depression in those mice. Thus, try to reveal the biological mechanisms of the pathogenesis of depression in adolescent females and provide a theoretical basis for the prevention and treatment of depression in pubertal females.

青春期女性抑郁症发病率高，预后严重，研究其发病机制具有重要学术价值和临床意义。 单胺假说不能完全解释抑郁症发病机制。近年来GABA/谷氨酸失衡假说受到重视。突触外及周围GABAA受体及神经甾体THP在青春期女性焦虑的发生中起重要作用，而焦虑是抑郁症的前期症状，项目科学问题：突触外及周围GABAA受体及THP是否导致青春期女性抑郁症的发生？ 研究内容：1）制备青春期雌性野生及敲基因抑郁症动物模型2）应用双光子成像、双光子阴影膜片钳和双光子解笼等先进技术，结合抑郁行为学观测、免疫电镜及Western Blot方法，观察THP对：①青春期抑郁海马锥体细胞树突棘突触外及周围GABAA受体亚型表达及其GABA tonic电流方向 ②谷氨酸及其NMDA电流 ③树突棘和突触密度 ④青春期动物抑郁行为特征的影响。 目的：期待在青春期女性抑郁症发病机制研究上取得突破，为青春期女性抑郁症的防治提供理论依据。

青春期女性抑郁症发病率急剧增加，症状更为严重，药物治疗效果差、易反复发作、自杀率较高。迄今其发病机制不情，生物学基础不明。我们前期研究表明，青春期GABAA 受体δ受体亚基表达变化是青春期女性焦虑症的发生生物学基础，青春期GABAA 受体δ受体亚基表达变化是否与青春期女性抑郁症发病率急剧增加有关？.为此我们应用了野生和GABAA 受体δ受体亚基敲基因小鼠，观察了慢性不可预知应急（CUMS）、神经甾体THP及δ受体亚基特异性激活剂Gaboxadol对小鼠抑郁行为、海马神经元膜上GABAA 受体δ受体亚基及AMPA受体GluA1和GluA2表达、膜片钳神经元电生理指标、双光子显微镜及Golgi染色标记神经元树突棘密度以及免疫组化荧光标记中间神经元δ受体亚基表达变化。.我们的研究结果表明：GABAA受体δ亚基在青春期抑郁的发生中起重要的作用，δ受体亚基敲基因小鼠导致对慢性不可预知应急敏感，易导致青春期抑郁状态；神经甾体THP引起青春期小鼠抑郁行为表现或加重青春期抑郁小鼠的抑郁状态，降低青春期小鼠海马神经元δ亚基表达及GABA tonic电流，降低海马中间神经元δ亚基荧光强度，减少海马神经元树突棘密度。GABAA受体δ亚基特异性激活剂Gaboxadol（即THIP）通过激活中间神经元突触外及周围的GABAA受体δ亚基，抑制中间神经元兴奋性，使其对椎体神经元的抑制作用消除，使椎体神经元AMPA受体trafficking上膜增加，改善青春期抑郁行为，有可能是治疗青春期抑郁症的一个重要途径。 .本项目研究成果揭示了青春期女性抑郁症发病的生物学的可能机制，明确了GABAA 受体δ亚基在青春期女性抑郁症发病中的作用以及神经甾体THP诱发及加重青春期女性抑郁症状所起作用，特别是发现GABAA 受体δ亚基特异性激活剂Gaboxadol能快速缓解改善青春期雌性小鼠抑郁状态，为抑郁症特别是青春期女性抑郁症的防治提供新途径，研究成果取得重大突破。