钙调素拮抗剂E6对大鼠神经细胞内钙的影响

Effects of E6, a calmodulin antagonist, on intracellular free Ca^(2+) level in neurons

目的 观察钙调素拮抗剂E6对神经细胞静息[Ca2 +]i、KCl (5 0mmol·L-1)和谷氨酸 (glutamicacid ,Glu ,0 1mmol·L-1)引起的神经细胞 [Ca2 +]i 升高的影响。方法 用Ca2 +敏感荧光指示剂Fura 2 /AM负载大鼠脑细胞 ,测定神经细胞内游离Ca2 +浓度 ([Ca2 +]i)。结果 E6可升高神经细胞静息 [Ca2 +]i,EC50 为 6 6 8μmol·L-1;无外Ca2 +条件下 ,也升高 [Ca2 +]i,EC50 为 1 30 μmol·L-1,说明E6主要是通过促进细胞内贮存Ca2 +释放引起内Ca2 +升高。E6抑制KCl升高细胞 [Ca2 +]i 的IC50 为 6 0 μmol·L-1;抑制Glu激发的细胞 [Ca2 +]i 升高的IC50 为 0 15 μmol·L-1。结论 E6可能是通过与细胞内钙调素 (Calmodulin ,CaM)结合后 ,影响兴奋性氨基酸受体的开放和电压依赖性钙通道的活性状态 ,表现出对由去极化或受体激动剂引起的内Ca2

AIM To study effects of E6, a calmodulin antagonist, on the intracellular free Ca 2+ concentration([Ca 2+ ] i ) in brain cells isolated from new born rats. METHODS Using Fura 2/AM, a Ca 2+ sensitive fluorescent indicator, to load the brain cells isolated from new born rats, the intracellular free Ca 2+ concentration ([Ca 2+ ] i ) was measured with fluorescnce spectrophotometer. RESULTS E6 elevated resting [Ca 2+ ] i both in presence of 1 3 mmol·L -1 Ca 2+ and in absence of Ca 2+ in Hanks solution. EC 50 values were 6 68 μmol·L -1 and 1 30 μmol·L -1 respectively. E6 inhibited high K + induced [Ca 2+ ] i elevation with IC 50 value of 6 0 μmol·L -1 , and the glutamate induced [Ca 2+ ] i elevation with IC 50 value of 0 15 μmol·L -1 . CONCLUSION E6 increases [Ca 2+ ] i in resting status mainly by promoting the release of stored Ca 2+ in neurons. After addition of calmodulin to the neurons,E6 shows inhibition of [Ca 2+ ] i elevation induced by KCl and glutamate through affecting the opening state of channels.