不同浓度二十二碳六烯酸对正常大鼠冠状动脉平滑肌细胞BK通道的激活作用及其机制

发布时间：2018-10-25 20:41

【摘要】：目的:探讨不同浓度二十二碳六烯酸(DHA)对正常大鼠冠状动脉平滑肌细胞大电导钙激活钾离子通道(BK通道)的激活作用及其机制。方法:酶消化法分离正常大鼠冠状动脉平滑肌细胞。采用全细胞膜片钳实验技术记录在未孵育与孵育细胞色素P450环氧化酶抑制剂SKF525A并灌流不同浓度DHA条件下BK通道电流密度变化;采用单通道膜片钳实验技术记录灌流不同浓度DHA时BK单通道开放概率的变化。结果:0.01~1.00μmol/L(定义为低浓度)DHA激活BK通道的半效浓度(EC50)为(0.24±0.05)μmol/L,但经SKF525A孵育后激活作用消失;3.00~10.00μmol/L(定义为高浓度)DHA激活BK通道的EC50为(2.38±0.22)μmol/L,经SKF525A孵育后,激活作用仍存在,且EC50无明显变化。在电极外液钙离子浓度为1μmol/L和刺激电位60 m V条件下,DHA浓度为0、0.01、0.03、0.10、0.30、1.00μmol/L时,BK通道开放概率分别为(0.095 2±0.009 5)、(0.093 9±0.012 6)、(0.098 6±0.016 9)、(0.099 5±0.014 7)、(0.097 5±0.010 4)和(0.102 3±0.020 6)(P0.05,n=5),提示低浓度DHA不能激活BK单通道;继续增加DHA浓度,当浓度为3、10μmol/L时,BK单通道开放概率分别为(0.700 3±0.013 2)和(0.892 7±0.052 3)(P0.05,n=5),提示高浓度DHA呈浓度依赖性激活BK单通道,EC50为(3.37±0.10)μmol/L。结论:不同浓度DHA激活BK通道的机制不同,低浓度DHA通过细胞色素P450环氧化酶途径激活BK通道,高浓度DHA可能通过与BK通道结合后直接激活。
[Abstract]:Aim: to investigate the effect and mechanism of 22 carbohexaenoic acid (DHA) on calcium activated potassium channel (BK channel) in normal rat coronary artery smooth muscle cells. Methods: normal rat coronary artery smooth muscle cells were isolated by enzyme digestion. Whole cell patch clamp technique was used to record the changes of current density of BK channel in unincubated and incubated cytochrome P450 cyclooxygenase inhibitor (SKF525A) and perfused with different concentrations of DHA. The single channel patch clamp technique was used to record the change of BK single channel open probability when perfused with different concentrations of DHA. Results: the half effect concentration (EC50) of DHA activated BK channel was (0. 24 卤0. 05) 渭 mol/L, (defined as low concentration) by 0. 01 渭 mol/L (defined as low concentration), but the activation effect disappeared after incubation with SKF525A (0. 24 卤0. 05) 渭 mol/L,. The EC50 of 3.00 渭 mol/L (defined as high concentration) DHA activated BK channel was (2.38 卤0.22) 渭 mol/L,. After incubation with SKF525A, the activation still existed, and EC50 did not change significantly. The opening probability of BK channel was (0.095 2 卤0.009 5), (0.093 9 卤0.012 6) when the concentration of calcium ion was 1 渭 mol/L and the stimulation potential was 60 MV, and the concentration of DHA was 0 0. 01 渭 mol/L, 0. 10 and 0. 30 渭 mol/L, respectively, when the concentration of Ca2 + was 1 渭 mol/L and the stimulation potential was 60 MV, the opening probability of BK channel was (0.095 2 卤0.009 5), (0.093 9 卤0.012 6). (0.098 6 卤0.016 9), (, 0.099 5 卤0.014 7), (, 0.097 5 卤0.010 4) and (0.102 3 卤0.020 6) (P 0.05 卤0.020 6), indicating that low concentration DHA could not activate BK single channel. The open probability of single channel of BK was (0.700 3 卤0.013 2) and (0.892 7 卤0.052 3) (P 0.05), respectively, when the concentration of BK was 3 渭 mol/L and 10 渭 mol/L, suggesting that high concentration of DHA activated BK single channel in a concentration-dependent manner. EC50 = (3.37 卤0.10) 渭 mol/L. Conclusion: the mechanism of activation of BK channel by different concentration of DHA is different. Low concentration of DHA activates BK channel through cytochrome P450 cyclooxygenase pathway, and high concentration of DHA may be activated directly by binding to BK channel.
【作者单位】：南京医科大学附属无锡市人民医院心内科;山东省医学科学院基础医学研究所心血管病研究室;Mayo
【基金】：国家自然科学基金(81370303,81500249,81500323) 江苏省自然科学基金(BK20151110) 江苏省人事厅“六大人才高峰”第七批高层次项目(006) 江苏省医学重点人才资助项目(RC201134)
【分类号】：R54

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