PLC‐dependent intracellular Ca2+ release was associated with C6‐ceramide‐induced inhibition of Na+ current in rat granule cells
PLC‐dependent intracellular Ca2+ release was associated with C6‐ceramide‐induced inhibition of Na+ current in rat granule cells
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DOI：
10.1111/j.1471-4159.2008.05562.x
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发表时间：
2008-09
影响因子：
4.7
通讯作者：
Zheng Liu;Xiao‐Wei Fei;Yan-Jia Fang;Wen-Jie Shi;Yu-Qiu Zhang;Y. Mei
中科院分区：
文献类型：
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作者：
Zheng Liu;Xiao‐Wei Fei;Yan-Jia Fang;Wen-Jie Shi;Yu-Qiu Zhang;Y. Mei
在本报告中，研究了C6-神经酰胺对培养的大鼠小脑颗粒细胞的电压门控内向Na+电流（INa）、两种类型的主要K+电流[外向整流延迟K+电流（IK）和外向瞬时K+电流（IA）]以及细胞死亡的影响。在0.01-100 μM浓度下，神经酰胺对INa产生剂量依赖性和可逆性抑制，而不改变稳态活化和失活特性。C2神经酰胺治疗对INa产生了类似的抑制作用。然而，二氢-C6-神经酰胺未能调节INa。C6-神经酰胺对INa的影响可通过细胞内输注Ca 2 +-螯合剂1，2-双（2-氨基苯氧基）乙烷-N，N，N9，N9-四乙酸消除，但可通过应用咖啡因模拟。用Ryanodine受体阻断剂阻断肌浆网Ca ~（2+）释放可使INa振幅逐渐增加，并消除神经酰胺对INa的影响。相比之下，肌醇1，4，5-三磷酸敏感性Ca 2+受体的阻断剂不影响C6-神经酰胺的作用。细胞内应用GTPγS也诱导INa振幅逐渐降低，而GDPβS消除了C6-神经酰胺对INa的影响。此外，C6-神经酰胺对INa的影响在应用磷脂酶C（PLC）阻断剂后被消除，并被钙调蛋白抑制剂大大降低。荧光染色显示C6-神经酰胺降低细胞活力，河豚毒素阻断INa不能模拟C6-神经酰胺的作用，丹曲林抑制细胞内Ca 2+释放不能减少C6-神经酰胺诱导的细胞死亡。因此，我们认为，通过兰尼碱敏感的Ca 2+受体增加PLC依赖性Ca 2+释放可能是C6神经酰胺诱导的INa抑制的原因，这似乎与C6神经酰胺诱导的颗粒神经元死亡无关。
In this report, the effects of C6‐ceramide on the voltage‐gated inward Na+ currents (INa), two types of main K+ current [outward rectifier delayed K+ current (IK) and outward transient K+ current (IA)], and cell death in cultured rat cerebellar granule cells were investigated. At concentrations of 0.01–100 μM, ceramide produced a dose‐dependent and reversible inhibition of INa without alteration of the steady‐state activation and inactivation properties. Treatment with C2‐ceramide caused a similar inhibitory effect on INa. However, dihydro‐C6‐ceramide failed to modulate INa. The effect of C6‐ceramide on INa was abolished by intracellular infusion of the Ca2+‐chelating agent, 1,2‐bis (2‐aminophenoxy) ethane‐N, N, N9, N9‐tetraacetic acid, but was mimicked by application of caffeine. Blocking the release of Ca2+ from the sarcoplasmic reticulum with ryanodine receptor blocker induced a gradual increase in INa amplitude and eliminated the effect of ceramide on INa. In contrast, the blocker of the inositol 1,4,5‐trisphosphate‐sensitive Ca2+ receptor did not affect the action of C6‐ceramide. Intracellular application of GTPγS also induced a gradual decrease in INa amplitude, while GDPβS eliminated the effect of C6‐ceramide on INa. Furthermore, the C6‐ceramide effect on INa was abolished after application of the phospholipase C (PLC) blockers and was greatly reduced by the calmodulin inhibitors. Fluorescence staining showed that C6‐ceramide decreased cell viability and blocking INa by tetrodotoxin did not mimic the effect of C6‐ceramide, and inhibiting intracellular Ca2+ release by dantrolene could not decrease the C6‐ceramide‐induced cell death. We therefore suggest that increased PLC‐dependent Ca2+ release through the ryanodine‐sensitive Ca2+ receptor may be responsible for the C6‐ceramide‐induced inhibition of INa, which does not seem to be associated with C6‐ceramide‐induced granule neuron death.