突触发生期酒精介导CaMKII/GSK3β信号失衡促进脊髓背角GABA能神经元凋亡

突触发生期酒精暴露会导致引发中枢神经元凋亡，进而引发一连串的中枢神经系统结构和功能障碍。本课题组之前的研究表明，突触发生期小鼠酒精暴露，选择性诱导脊髓背角GABA能神经元凋亡，具有明显的时间易感性和区域细胞特异性，但目前尚不知道其发生机制。我们的预实验发现突触发生期酒精暴露激活脊髓GSK3β信号，但抑制CaMKII磷酸化。基于此，我们推测，突触发生期酒精暴露引起CaMKII/GSK3β信号失调，导致选择性脊髓GABA能神经元凋亡。本项目拟建立酒精损伤脊髓细胞、组织和动物模型，通过小分子阻断剂或激动剂、质粒和病毒，干预CaMKII/GSK3β信号，运用免疫组织化学染色、免疫荧光染色、蛋白印迹、蛋白沉淀和定量PCR等技术方法，研究CaMKII/GSK3β信号失衡在酒精诱导的脊髓GABA能神经元凋亡中的分子调控机制，期望为干预酒精及其类似的中枢毒性物质损害提供靶点。

The developing nervous system is sensitive to ethanol exposure. During the period of synaptogenesis it is particularly vulnerable to ethanol-induced neurodegeneration, which in turn triggers a series of structural and functional disorders of the central nervous system. We have previously shown that ethanol exposure during the synaptogenesis period (early postnatal days) (PD4 and PD7) produced robust apoptosis and neurodegeneration of GABAergic neurons in the developing mouse spinal cord, without significant effects on glutamatergic neurons, which has a temporal and regional susceptibility. However, the mechanism of GABAergic neuron apoptosis is not yet known. Studies revealed that activated CaMKII can directly phosphorylate GSK-3β in Ser9, resulting in its inactivation. Moreover, our pre-experiment results show that ethanol exposure during the synaptogenesis period activates GSK3β signaling in mouse spinal cord, but inhibits CaMKII phosphorylation. Based on above, this project intends to establish the alcohol-damaged model of spinal cord at the cellular, tissue and animal level. Small molecule compound, as well as CaMKII/GSK3β plasmids and viruses, is used to intervene with the CaMKII/GSK3β signal in the ethanol-damaged model, and immunohistochemical staining, fluorescence staining, western blotting, protein precipitation and quantitative PCR are used to study the molecular regulation mechanism of CaMKII/GSK3β signal imbalance in ethanol-induced spinal GABAergic neuron apoptosis. It is expected to provide the target for intervention in the harm of alcohol and similar central toxic substances.