豚鼠Ⅱ型前庭毛细胞乙酰胆碱敏感性大电导钙依赖性钾通道与L型钙通道共存

Co-location of Ach-sensitive BK channels and L-type calcium channels in type II vestibular hair cells of guinea pig

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LA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）；-50mV钳制电压下，100μmol／LACh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，200nmol／L）敏感，而细胞外蜂毒明肽（apamin，1μmol／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-K8644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。

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 100μmol/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. Condusinn ACh-sensitive BK and L-type calcium channels are co-located in type Ⅱ vestibular hair cells of guinea pigs.