Ro20-1724对氯胺酮诱导乳鼠海马神经元凋亡的影响

Effect of Ro20-1724 on ketamine-induced apoptosis in hippocampal neurons of neonatal rats

目的 探讨Ro20-1724对氯胺酮诱导乳鼠海马神经元凋亡的影响.方法 采用随机数字表法将乳鼠原代海马神经元分为4组（n=24）：对照组（C组）、氯胺酮组（K组）、溶剂对照组（E组）和Ro20-1724组（R组）.C组采用常规培养液培养72 h;K组采用含150 μmol/L氯胺酮的培养液孵育72 h;E组和R组经含150 μmol/L氯胺酮培养液孵育30 min后,更换培养液,分别加入终浓度为0.01％的乙醇和1x10^-3 μmol/L的Ro20-1724孵育72 h.各组处理结束后,采用MTT法测定海马神经元存活率,流式细胞仪测定海马神经元凋亡率,RT-PCR法测定海马神经元Bcl-2 mRNA和Bax mRNA表达,Western blot法测定海马神经元突触素Ⅰ表达.结果 与C组比较,K组和E组海马神经元存活率降低,凋亡率升高,Bcl-2 mRNA和突触素Ⅰ表达下调,Bax mRNA表达上调（P＜0.05或0.01）;与K组比较,R组海马神经元存活率升高,凋亡率降低,Bcl-2 mRNA和突触素Ⅰ表达上调,Bax mRNA表达下调（P ＜0.05或0.01）.结论 Ro20-1724可抑制氯胺酮导致的乳鼠海马神经元凋亡,机制与纠正Bcl-2/Bax失衡状态有关.

Objective To valuate the effect of Ro20-1724 on ketamine-induced apoptosis in hippocampal neurons of neonatal rats.Methods Hippocampal neurons from newborn Sprague-Dawley rats were obtained and cultured in vitro.The primary hippocampal neurons were randomly divided into 4 groups （n =24 each） using a random number table：control group （group C）,ketamine group （group K）,solvent control group （group E）,and Ro20-1724 group （R group）.The neurons were incubated for 72 h in the normal culture medium in group C.The neurons were incubated for 72 h in the culture medium containing ketamine 150 μmol/L in group K.In E and R groups,after the neurons were incubated for 30 min in the culture medium containing ketamine 150 μmol/L,the culture medium was then replaced,0.01% ethanol （final concentration） and 1 ＆#215; 10-3 μmol/L Ro20-1724 （final concentration） were added to the culture medium,respectively,and the neurons were then incubated for 72 h.After 72 h incubation,the cell viability was detected by MTT assay,the cell apoptosis was detected by flow cytometry,the expression of Bcl-2 mRNA and Bax mRNA was determined by RT-PCR,and synaptophysin Ⅰ expression was detected by Western blot.The apoptosis rate was calculated.Results Compared with group C,the survival rate was significantly decreased,the apoptosis rate was increased,the expression of Bcl-2 mRNA and synaptophysin Ⅰ was down-regnlated,and Bax mRNA expression was up-regulated in K and E groups （P 〈 0.05 or 0.01）.Compared with group K,the survival rate was significantly increased,the apoptosis rate was decreased,the expression of Bcl-2 mRNA and synaptophysin Ⅰ was up-regulated,and Bax mRNA expression was downregulated in R group （P 〈 0.05 or 0.01）.Conclusion Ro20-1724 can inhibit ketamine-induced apoptosis in hippocampal neurons of neonatal rats and correction of Bcl-2/Bax imbalance is involved in the mechanism.