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目的研究豚鼠Ⅱ型前庭毛细胞乙酰胆碱(acetylcholine,ACh)敏感性钾电流对钙离子的依赖性。方法应用全细胞膜片钳技术研究新鲜单离的豚鼠Ⅱ型前庭毛细胞ACh-敏感性钾电流在细胞内外钙离子浓度改变时其电流幅值的变化。结果①细胞外ACh激活一缓慢持久的外向性钾电流,其反转电位为(-70±10)mV;②ACh-敏感性钾电流电流对细胞外四乙胺(10mmol/L)敏感,而对细胞外4-氨基吡啶(100μmol/L)不敏感;③ACh-敏感性钾电流的幅值大小依赖于细胞外的钙离子浓度,无钙外液中ACh激活一很小的电流,4mmol/L外钙溶液中ACh-敏感性钾电流的幅值达到最大值,而0·5mmol/L外钙溶液中ACh-敏感性钾电流的幅值抑制至(36·5±6·5)%;④细胞内三磷酸肌醇-钙离子释放过程不参与ACh-敏感性钾电流的激活,细胞内透析肝素8mg/ml后30min,ACh-敏感性钾电流的幅值没有明显改变;⑤ACh敏感性钾电流对钙通道阻断剂Cd2+敏感。结论豚鼠Ⅱ型前庭毛细胞ACh-敏感性钾电流的激活依赖于细胞外的钙离子浓度,ACh与豚鼠Ⅱ型前庭毛细胞胆碱能受体结合后,首先激活膜上钙通道引起细胞外钙离子内流,毛细胞内游离钙离子浓度的升高进一步激活钙依赖性钾电流。
Objective To study the calcium ion dependence of acetylcholine (ACh) sensitive potassium current in guinea pig type Ⅱ vestibular hair cells. Methods The whole cell patch clamp technique was used to study the change of current amplitude of ACh-sensitive K + currents in freshly isolated guinea pig type Ⅱ vestibular hair cells with changes of intracellular Ca 2+ concentration. RESULTS: (1) Extracellular ACh activated a slow and sustained outward potassium current (-70 ± 10) mV; (2) ACh-sensitive potassium currents were sensitive to extracellular tetraethylammonium (10mmol / L) (100μmol / L); (3) The amplitude of ACh-sensitive potassium currents depended on the concentration of extracellular calcium, and ACh-activated extracellular currents The amplitude of ACh-sensitive potassium current in calcium solution reached the maximum, while the amplitude of ACh-sensitive potassium current in 0.5 mmol / L Ca (superscript 2+) solution was inhibited to (36.5 ± 6.5)%. Inositol triphosphate - calcium release process is not involved in ACh-sensitive potassium current activation, intracellular hemodialysis 8mg / ml after 30min, ACh-sensitive potassium current amplitude did not change significantly; ⑤ ACh sensitive potassium current The calcium channel blocker Cd2 + is sensitive. Conclusion The activation of ACh-sensitive potassium current in type Ⅱ vestibular hair cells in guinea pigs depends on the extracellular calcium concentration. After ACh binds to the cholinergic receptor of type Ⅱ vestibular hair cells in guinea pigs, activation of extracellular calcium Ionic influx, intracellular free calcium concentration in the plasma further activation of calcium-dependent potassium current.