Puerarin facilitates Ca2+ -induced Ca2+ release triggered by KCl-depolarization in primary cultured rat hippocampal neurons
Puerarin facilitates Ca2+ -induced Ca2+ release triggered by KCl-depolarization in primary cultured rat hippocampal neurons
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DOI：
10.1016/j.ejphar.2007.05.023
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发表时间：
2007-09
影响因子：
5
通讯作者：
Fankai Lin;Yan Xin;Jiang-hua Wang;Liqun Ma;Jue Liu;Chao Liu;Li‐Hong Long;Fang Wang;You Jin;Jun Zhou;Jian-Guo Chen
中科院分区：
文献类型：
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作者：
Fankai Lin;Yan Xin;Jiang-hua Wang;Liqun Ma;Jue Liu;Chao Liu;Li‐Hong Long;Fang Wang;You Jin;Jun Zhou;Jian-Guo Chen
在动物研究中，葛根素对行为和大脑神经元活动的影响已经被描述过。然而，这些效应背后的分子机制却知之甚少。在这里，我们利用基于Fura-2的钙离子成像技术研究了葛根素对原代培养大鼠海马神经元钙信号的调节。应用葛根素对87%的记录神经元的基础细胞内钙离子浓度([Ca~(2+)]_i)无影响，但可增强KCl诱发的[Ca~(2+)]_i。兰尼定受体的阻断剂丹曲林或钌红可完全阻断葛根素诱导的这种增强作用。此外，在无钙溶液中，预先应用葛根素可显著增强咖啡因(3 MM)引起的[Ca~(2+)]升高，咖啡因是兰尼定受体的特异性激动剂。相反，硝苯地平不能阻止葛根素的增强作用。同样，在全细胞膜片钳记录实验中，葛根素对去极化脉冲产生的钙电流没有任何影响。这些结果表明，葛根素的增强作用是通过兰尼定受体促进钙离子诱导的钙释放(CICR)，而不是细胞外钙内流。使用雌激素受体拮抗剂ICI182780和三苯氧胺，我们进一步证明葛根素的增强作用是由雌激素受体介导的。此外，蛋白激酶A(PKA)的膜透性抑制剂H89完全抑制这种增强作用。而磷脂酶C的抑制剂U-73122则无此作用，提示cAMP/PKA信号转导通路参与了葛根素对CICR的激活作用。
The effects of puerarin on behaviour and brain neuronal activity in animal studies have been described previously. However, molecule mechanisms underlying these effects were poorly understood. Here, we examined the regulation of puerarin on the Ca2+signals in primary rat hippocampal neurons using Fura-2 based calcium imaging techniques. Application of puerarin had no effect on the basal intracellular calcium concentration ([Ca2+]i), but potentiated the KCl-evoked [Ca2+]itransient in 87% of recorded neurons. Dantrolene or ruthenium red, the inhibitors of ryanodine receptors, completely blocked this potentiation induced by puerarin. Moreover, in Ca2+-free solution, pre-application of puerarin significantly augmented the elevation of [Ca2+]ievoked by caffeine (3 mM), which is a specific agent to activate the ryanodine receptors. In contrast, nifedipine failed to prevent the potentiation induced by puerarin. Similarly, in the experiments of whole-cell patch-clamp recording, puerarin did not show any effect on calcium currents generated by depolarization pulses. These data demonstrated that the potentiation induced by puerarin was attributed to the facilitation of Ca2+-induced Ca2+release (CICR) via ryanodine receptors, rather than extracellular Ca2+influx. Using estrogen receptor antagonist ICI 182780 and tamoxifen, we further demonstrated that the potentiation induced by puerarin was mediated by the estrogen receptor. Furthermore, the membrane-permeant inhibitor of protein kinase A (PKA) H89 completely inhibited this potentiation. However, U-73122, the inhibitor of phospholipase C (PLC) had no effect, indicating that the cyclic AMP/PKA signaling pathway was involved in the activation of CICR by puerarin.