CAJ | 학술논문

目的研究豚鼠Ⅱ型前庭毛细胞乙酰胆碱(acetylcholine,ACh)敏感性大电导钙依赖性钾通道(big conductance,calcium-dependent potassium channel,BK)激活过程中的钙离子内流机制.方法健康杂色豚鼠52只,断头后取出前庭终器,经胶原酶IA消化后获取Ⅱ型前庭毛细胞.采用全细胞膜片钳技术检测新鲜单离的Ⅱ型前庭毛细胞ACh-敏感性BK电流对细胞外钙通道阻断剂和激动剂的敏感性.结果 ①细胞外ACh激活一缓慢持久的外向性电流,其反转电位为(-70.5±10.6)mV(-x±s,下同,n=10);-50 mV钳制电压下,100 μmol/L ACh激活电流的幅值为(267±106)pA(n=11).②ACh-敏感性钾电流对细胞外IBTX(iberiotoxin,200 nmol/L)敏感,而细胞外蜂毒明肽(apamin,1 μmol/L)对ACh-敏感性钾电流幅值无抑制作用.③ACh-敏感性BK对细胞外钙通道阻断剂NiCl2敏感,可被细胞外钙通道阻断剂CdCl2强力阻断.NiCl2和CdCl2对ACh-敏感性BK的半数抑制浓度分别为(135.5±18.5)μmol/L(n=7)和(23.4±2.6)μmol/L(n=7).④L型钙通道激动剂(-)-Bay-K 8644激活Ⅱ型前庭毛细胞产生一种与ACh-敏感性BK类似的外向性电流,并能被IBTX强力阻断.结论 Ⅱ型前庭毛细胞中ACh-敏感性BK与细胞膜L型钙通道共存,可能具有重要的生理学作用和意义.
목적 연구돈서Ⅱ형전정모세포을선담감(acetylcholine,ACh)민감성대전도개의뢰성갑통도(big conductance,calcium-dependent potassium channel,BK)격활과정중적개리자내류궤제.방법 건강잡색돈서52지,단두후취출전정종기,경효원매IA소화후획취Ⅱ형전정모세포.채용전세포막편겸기술검측신선단리적Ⅱ형전정모세포ACh-민감성BK전류대세포외개통도조단제화격동제적민감성.결과 ①세포외ACh격활일완만지구적외향성전류,기반전전위위(-70.5±10.6)mV(-x±s,하동,n=10);-50 mV겸제전압하,100 μmol/L ACh격활전류적폭치위(267±106)pA(n=11).②ACh-민감성갑전류대세포외IBTX(iberiotoxin,200 nmol/L)민감,이세포외봉독명태(apamin,1 μmol/L)대ACh-민감성갑전류폭치무억제작용.③ACh-민감성BK대세포외개통도조단제NiCl2민감,가피세포외개통도조단제CdCl2강력조단.NiCl2화CdCl2대ACh-민감성BK적반수억제농도분별위(135.5±18.5)μmol/L(n=7)화(23.4±2.6)μmol/L(n=7).④L형개통도격동제(-)-Bay-K 8644격활Ⅱ형전정모세포산생일충여ACh-민감성BK유사적외향성전류,병능피IBTX강력조단.결론 Ⅱ형전정모세포중ACh-민감성BK여세포막L형개통도공존,가능구유중요적생이학작용화의의.
Objective To explore the mechanisms of the influx of calcium ions during the activation of Ach-sensitive BK channel(big conductance,calcium-dependent potassium channel)in type Ⅱ vestibular hair cells of guinea pigs. Methods Type Ⅱ vestibular hair cells were isolated by collagenase type IA.Under the whole-cell patch mode,the sensitivity of Ach-sensitive BK current to the calcium channels blockers was investigated,the pharmacological property of L-type calcium channel activator-sensitive current and Ach-sensitive BK current was compared. Results Following application of Ach, type Ⅱ vestibular hair cells displayed a sustained outward potassium current,with a reversal potential of(-70.5±10.6)mV(-x±s,n=10). At the holding potential of -50 mV, the current amplitude of Ach-sensitive potassium current activated by 100 μmol/L Ach was(267±106) pA(n=11). Ach-sensitive potassium current was potently sensitive to the BK current blocker, IBTX(iberiotoxin, 200 nmol/L). Apamin,the well-known small conductance, calcium-dependent potassium current blocker, failed to inhibit the amplitude of Ach-sensitive potassium current at a dose of 1 μmol/L. Ach-sensitive BK current was sensitive to NiCl2 and potently inhibited by CdCl2. NiCl2 and CdCl2 showed a dose-dependent blocking effect with a half inhibitionmaximal response of(135.5±18.5)μmol/L(n=7) and (23.4±2.6) μmol/L(n=7). The L-type calcium channel activator,(-)-Bay-K 8644(10 μmol/L),mimicked the role of Ach and activated the IBTX-sensitive outward current. Conclusion Ach-sensitive BK and L-type calcium channels are co-located in type Ⅱ vestibular hair cells of guinea pigs.