SNARE蛋白复合体在产前应激引起的子代抑郁样行为中的作用机制研究

Exposure to prenatal stress could significantly increase the incidence of depression in offspring, the pathogenesis is unclear and there are many defects in the existing treatments. Numerous reports and our studies showed that the glutamate (Glu) regulation barriers will lead to the occurrence of depression, and the SNARE complex play an important role in the release of neurotransmitter such as Glu. Whether the SNARE proteins and the mechanisms are involved in prenatally stressed offsprings are unclear, we preliminarily studied the effects of SNARE complex in prenatally stressed offspring found that the expression of mRNA and protein of SNARE core protein are remarkably increased. On these basises, we will observe the SNARE complex and their phosphorylation and the membrane fusion-associated proteins in the hippocampus and prefrontal cortex, analyze the interaction between core proteins of SNARE complex, investigate the variation of vesicular glutamate transporter and and ionotropic glutamate receptors，also decrease SNARE core protein by RNA interference in primary cultured hippocampal neurons in vitro to observe their impact on Glu release. The study of the mechanism of SNARE complex in Glu release in synaptic cleft clarifies in prenatally stressed offspring may have important implications for the mechanism of Glu regulation in pathogenesis of depression in prenatally stressed offspring, but also provide the theoretical and experimental evidence in the discovery of antidepressant drug target for clinical treatment.

产前应激（Prenatal Stress，PS）可显著增加子代抑郁的发生，但其发病机制不清，现有的治疗药物也存在诸多缺陷。已证实谷氨酸（Glu）调控障碍会导致抑郁的发生，而SNARE蛋白复合体在Glu等神经递质释放过程中至关重要。SNARE是否参与PS引起的子代抑郁及机制尚不明确，前期研究发现PS子代SNARE组成蛋白mRNA及蛋白表达显著上调。本项目拟在此基础上，探讨PS对子代海马和前额叶皮层中SNARE复合体的形成、磷酸化以及膜融合相关蛋白的影响，分析SNARE组成蛋白之间的相互作用，考察囊泡Glu转运体以及离子型Glu受体的变化，并且体外RNA干扰原代培养海马神经元中SNARE组成蛋白，观察神经元谷氨酸释放的改变，从而阐明SNARE蛋白复合体在PS子代Glu释放中的相关分子机制。这些研究对探讨PS引起子代抑郁的发病机制具有重要意义，也为临床发现抗抑郁治疗的药物靶点提供理论及实验依据。

产前应激（Prenatal stress, PS）可显著增加子代抑郁的发生，但其发病机制不清，现有的治疗药物也存在诸多缺陷。大量文献以及本课题组研究表明PS会导致子代表现出抑郁样行为并引起海马及前额叶皮层谷氨酸（Glu）浓度的升高，其调控障碍会导致精神疾病的发生。Glu的释放依赖突触小体的外排作用，即突触小体质膜与神经元突触前膜的相互融合，这个融合过程正是由SNARE (soluble NSF-attached protein receptor) 复合体的形成来介导的。并且，SNARE蛋白复合体的形成是由若干个关键的步骤组成，受到N-乙基马来酰亚胺敏感因子（NSF）、nSec1/Munc18（SM蛋白）等一系列蛋白的影响。此外，SNARE蛋白磷酸化在调控突触囊泡和神经元突触前膜分子的相互作用方面起了重要的作用。为了阐明PS引起子代抑郁的具体机制，课题组建立了产前束缚应激大鼠模型，采用药理学，动物行为学，细胞生物学，分子生物学等方法，考察了SNARE蛋白在PS引起的子代抑郁样行为中的作用及机制。研究结果表明，PS可引起子代发生抑郁样行为并导致HPA轴功能亢进；同时，显著上调子代海马及前额叶皮层中SNARE复合体核心蛋白SNAP-25、VAMP1、Syntaxin 1A及其复合体以及膜融合相关蛋白NSF、CSPα、α-synuclein、synaptotagmin、synaptophysin、complexin、nSec1/Munc18（SM蛋白）等的基因及蛋白表达；此外，还可显著上调子代大鼠海马及前额叶皮层中phospho-SNAP-25、phospho-syntaxin1A、phospho-VAMP2、PKA、PKC的蛋白表达以及囊泡谷氨酸转运体vGLUT1的基因及蛋白表达。课题组还考察了PS对子代海马及前额叶皮层中糖皮质激素受体GR的核转位及GR敏感蛋白FKBP5蛋白表达的影响，结果表明，PS可显著下调GR的核转位，上调GR敏感蛋白FKBP5的蛋白表达，从而降低PS对HPA轴的负反馈调节。通过以上的研究，本项目阐明了SNARE蛋白和相关膜融合蛋白的过度表达促进了突触间隙Glu释放的增加是PS引起子代发生抑郁样行为的重要机制。该研究为抑郁症的预防和临床干预PS所致的抑郁样行为提供理论依据，也为临床发现抗抑郁治疗的药物靶点提供理论及实验依据。