(-)-Epicatechin-3-gallate對第四型電壓閥鈉離子通道與初級培養大鼠胚胎神經細胞電流的影響

Abstract

飲用綠茶對健康有許多益處，並且可以提振我們的精神。在試管和活體內的實驗已經證實兒茶素具有對抗氧化傷害的能力，並且會影響和細胞存活與細胞死亡有關的訊息傳遞途徑。除此之外，也發現 (-)-epigallocatechin-3-gallate (ECGC)具有抗癌症發生、抗高血壓以及降低心血管疾病發生風險的功效。更進一步地，EGCG和 (-)-epicatechin-3-gallate (ECG)也被發現會藉由調控離子通道的活性來影響神經的活性，尤其是藉由電壓閥鈉離子通道(VGSC)來調控。為了驗證兒茶素如何調控電壓閥鈉離子通道，我們對初級培養大鼠胚胎神經細胞和表現第四型電壓閥鈉離子通道(Nav1.4)的HEK 293T細胞使用全細胞模式的膜片箝制技術，並在施加ECG (30 μM)前後分別記錄細胞的鈉離子電流。我們的結果顯示ECG會減慢神經細胞鈉離子電流的衰退，但並不會影響神經細胞中VGSC的電壓依賴性的開關機制。施加ECG會使神經細胞中VGSC的不活化曲線往較負的電壓方向位移，並且會讓從不活化狀態回復到關閉狀態的比例下降。另外，ECG會減慢Nav1.4的電流衰退，並且會使離子通道的活化曲線往更低的電壓平移。ECG也會使Nav1.4的不活化曲線往較正的電壓方向位移，並且減低在從不活化狀態回復到關閉狀態的比例。再來，為了研究ECG會對神經細胞的突觸傳導造成什麼影響，我們施用Fluo-2來做胞內鈣離子的指示劑，並用波長為405 nm的雷射光活化MNI-glutamate來刺激目標神經細胞。初步的結果顯示，ECG可能對皮質神經細胞的突觸傳導有抑制的效果。這些結果指出一些關於ECG的新進展，包括ECG對VGSC的調控以及ECG會對VGSC不同表現型的動力學有不同的影響。

Consuming green tea refreshes the mind and provides many health benefits. Catechins have the ability against the oxidative damages in vitro and in vivo, and their effects on signal transduction pathways are associated with cell death and cell survival. Besides, (-)-epigallocatechin-3-gallate (ECGC) has the effects as anti-carcinogenesis, anti-hypertension, and lowering cardiovascular disease risk. Furthermore, EGCG and (-)-epicatechin-3-gallate (ECG) have effects on neural activities by modulating the ion channel activities, especially the voltage-gated sodium channels (VGSCs). To verify how catechins modulate the VGSC activities, primary cultured embryonic cortical neurons and HEK 293T cells which expressed Na(v)1.4 were patch-clamped in whole-cell mode, and sodium currents were recorded before and after ECG (30 μM) treatment. Our results suggested ECG slows the slow decay of neuronal sodium currents, whereas ECG did not alter the voltage-dependence activation of neuronal VGSCs. ECG shifts the inactivation curve negatively and lengthens the recovery from inactivation. Besides, ECG slows the decay of Na(v)1.4 current, and shifts the activation curve towards negative voltage. ECG shifts the inactivation curve positively and reduces the steady-state recovery of Na(v)1.4. To investigate how ECG affects the synaptic transmission in neuron, we applied the Fluo-2, a calcium dye, and MNI-glutamate to stimulate the target neuron locally by 405-nm laser. The preliminary results suggested that ECG may have an inhibitory effect on synaptic transmission in cortical neurons. These findings reveal the new knowledge about the modulation of ECG on VGSC, and ECG has different effects on kinetics of different VGSC subtypes.