磷脂酶C-三磷酸肌醇途径参与β-淀粉样肽(25-35)诱导的海马神经元[Ca^2＋]i的增加

Pathway of phospholipase C-inositol trisphosphate is involved in the increase of [Ca^(2+)]i induced by β-amyloid peptide (25-35) in hippocampal neurons

目的探讨内质网(ER)钙库在β-淀粉样肽(25-35)(Aβ25-35)诱导的细胞内钙超载中的作用。方法用钙高度特异性荧光探针Flou-3/AM负载海马神经元,进行荧光染色,利用激光扫描共焦显微镜观察在不同药物干预条件下海马神经元内游离钙离子荧光强度的变化。结果 2、10、20μmol/L各浓度组凝聚态Aβ25-35均增加了海马神经元胞内[Ca2+]i(n=5,P<0.05);三磷酸肌醇受体(IP3R)的特异性抑制剂2-APB明显地抑制了胞内[Ca2+]i的增加,蓝尼碱受体(RyR)的特异性抑制剂硝苯呋海因(dantrolene)却无法抑制。Aβ25-35作用1h后内质网钙容量即有明显下降,作用24h后降低更为明显。在无细胞外钙情况下,磷脂酶C(PLC)的抑制剂U73122部分抑制了Aβ25-35诱导的胞内[Ca2+]i的增加。结论凝聚态Aβ25-35可通过IP3途径产生,IP3作用于IP3R而引起内质网钙的释放,磷脂酶C的活化可能参与了上述过程。

Objective To investigate the effect of endoplasmic reticulum（ER） induced by β-amyloid peptide （25-35） （Aβ25-35） in hippocampal neurons. Methods calcium stores on calcium overload The hippocampal neurons were loaded with calcium-sensitive fluorescent indicator Fluo-3/AM. Intracellular calcium concentration （ [ Ca^2＋ ] i）changes were measured in different conditions of drug intervention by using laser scanning eonfoeal microscopy. Results Groups of 2, 10, 20μmol/L Aβ25-35 all increased [ Ca^2＋ ] i in hippoeampal neurons（ n = 5, P 〈 0.05）. 2-APB, an inhibitor of ER Ca^2＋ release through channels associated to inositol trisphosphate receptor（IP3R） , was shown to prevent the aggregated Aβ25.35^- induced rise of [ Ca^2＋ ] i, suggesting the involvement of Ca^2＋ released by ER. However, dantrolene, an inhibitor of ER Ca^2＋ release through channels associated to ryanodine receptor（RyR）, could not prevent the rise of [ Ca^2＋] i induced by Aβ25-35. Treatment with aggregated Aβ25-35 for 1 hour induced a significant decrease in the ER Ca^2＋ content （P 〈0. 01 ） , which was more pronounced 24 hours after the addition of aggregated Aβ25-35. The increase in [ Ca^2＋ ] i observed in aggregated Aβ25-35-treated cells was prevented by the phospholipase C （PLC） inhibitor U-73122 in the absence of extracellular Ca^2＋. Conclusion IP3 pathway is involved in the early release of Ca^2＋ from ER induced by Aβ25-35, the activation of PLC may involve in the process.