摘要目的 探讨氯胺酮对大鼠海马神经元cAMP反应元件结合蛋白(CREB)磷酸化水平的影响.方法 原代培养1 d龄SD大鼠海马神经元5 d,随机分为对照组(C组)和氯胺酮组(K组),每组6孔.K组在终浓度为1 000 μmol/L氯胺酮的Neurobasel培养液中孵育3 h,C组不做任何处理.采用流式细胞仪Aanexin V-PI共染法检测凋亡神经元,计算神经元凋亡率;采用免疫组化法测定神经元Ser133位点磷酸化的CREB(pCREB)表达水平;采用RT-PCR法半定量测定CREB下游基因脑源性生长因子(BDNF)mRNA和抑凋亡基因Bcl-2 ndlNA的表达.结果 与C组相比,K组神经元凋亡率升高,pcREB、BDNFmRNA及Bcl-2 mRNA的表达均下调(P＜0.01).结论 氯胺酮可能通过抑制CREB磷酸化,下调BDNF及Bcl-2表达,诱导新生大鼠海马神经元凋亡.

abstractsObjective To investigate the effects of ketamine on cAMP response element binding protein phosphorylation (pCREB) in hippecampal neurons of rats.Methods Neurons were enzymatically isolated from hippocampi of newborn SD rats.After being cultured for 5 days the primary hippocampal neurons were randomly divided into 2 groups: control group and ketamine group.The neurons in ketamine group were incubated in culture medium containing ketamine 1 000 μmol/L for 3 h.Neuronal apoptosis was detected by flow cytometry after staining with Annexin V-PI.The expression of pCREB was measured by immuno-histochemistry and the expression of BDNF mRNA and Bcl-2 mRNA was detected by RT-PCR.Results The apoptosis index was significantly higher in ketsmine group (92.6%±2.8%) than in control group (26.1%±1.2%).The expression of pCREB,Bcl-2 mRNA and BDNF mRNA was down-regulated in ketamine group.Conclusion Ketaminc can induce apoptosis in the primary hippocumpal neurons by down-regulating the expression of pCREB,Bcl-2 and BDNF.