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目的:研究去甲肾上腺素引起大鼠C6神经胶质瘤细胞中钙离子浓度([Ca~(2+)])增加的机理.方法:以荧光染料fura-2为指示剂,采用双波长荧光比值成像 的方法测定细胞中钙离子浓度.结果:通过激活细胞上的α_1肾上腺素能受体,去甲肾上腺素剂量依赖地使C6细胞中钙离子浓度增加.这种反应不依赖细胞外钙,且不受百日咳毒素(PTX)处理的影响.将细胞与磷酯酶C(PLC)抑制剂U73122或内质网Ca~(2+)-ATP酶抑制剂thapsigargin预孵育,去甲肾上腺素引起的胞内钙反应则消失;蛋白激酶C激动剂佛波醉酯(PMA)预处理细胞可以使去甲肾上腺素引起的胞内钙离子浓度升高幅度降低,而佛波醇酯的效应能够被蛋白激酶C抑制剂Ro31-8220或GF-109203X完全阻断.但是,改变胞内蛋白激酶A活性的药物对去甲肾上腺素的作用没有影响.结论:通过激活胞内的磷酯酶C,去甲肾上腺素使C6细胞的胞内钙库释放钙离子.去甲肾上腺素引起的细胞内钙离子浓度增加受蛋白激酶C的负性调节.
Objective: To study the mechanism of norepinephrine-induced increase of calcium ion concentration ([Ca 2+]) in rat C6 glioma.Methods: Using fluorescent dye fura-2 as indicator, Ratio imaging.Results: Norepinephrine dose-dependently increased the concentration of calcium in C6 cells by activating the alpha 1 adrenergic receptors on the cells.This reaction was independent of extracellular calcium, And was not affected by pertussis toxin (PTX) treatment.The cells were preincubated with phospholipase C (PLC) inhibitor U73122 or the endoplasmic reticulum Ca (2 +) - ATPase inhibitor thapsigargin, and norepinephrine-induced Intracellular calcium response disappeared; protein kinase C agonist PMA pretreatment cells can make norepinephrine-induced increase in intracellular calcium concentration decreased, and the effect of phorbol esters can be protein Kinase C inhibitor Ro31-8220 or GF-109203X completely blocked.However, drugs that change the activity of intracellular protein kinase A have no effect on the effect of norepinephrine.Conclusion: By activating intracellular phospholipase C, Epinephrine liberates calcium ions from the intracellular calcium stores of C6 cells Epinephrine-induced increase in intracellular calcium concentration is negatively regulated by protein kinase C.