心肌内向整流钾通道激动剂抑制异丙肾上腺素所致大鼠心室重构

目的:观察和探讨内向整流钾通道(Ⅰ_(K1))激动剂盐酸扎考必利(zacopride,Zac)对异丙肾上腺素(isoproterenol,Iso)所致心室重构的影响及作用机制。方法:SD大鼠随机分为正常对照组、Iso模型组、Zac干预组、Zac+氯喹干预组和卡托普利阳性对照组。腹腔注射异丙肾上腺素3 mg/kg,每天1次,连续给药10 d,观测各组全心质量/体质量比和左心室质量/体质量比。用全细胞膜片钳技术检测大鼠心室肌细胞电压门控钙电流(Ⅰ_(Ca-L))、静息膜电位(RMP)及动作电位时程(APD)的变化。选用新生1~3 d的SD乳鼠,用0.08%胰蛋白酶和0.04%Ⅱ型胶原酶消化心脏组织,经差速贴壁法和5-溴脱氧尿嘧啶核苷纯化心肌细胞后随机分成正常对照组、Iso模型组、Zac干预组、Zac+BaCl_2干预组和Zac+氯喹干预组,培养24 h后用激光共聚焦显微镜检测心肌细胞内游离钙离子浓度。结果:Iso模型组与正常对照组比较,全心肥厚指数、左心室肥厚指数明显增加,膜片钳结果提示RMP减小APD明显延长;Zac干预组明显抑制心肌肥大,并增大RMP,缩短APD。同时应用低剂量Ⅰ_(K1)抑制剂氯喹可明显抑制Zac的抗心室重构作用,并逆转Zac对RMP和APD影响。在乳鼠心肌细胞,Iso可使细胞表面积增大,细胞内Ca~(2+)]_i增高;Zac干预后细胞形态恢复至正常或接近正常水平,并显著减轻钙超载。Ⅰ_(K1)阻断剂BaCl_2和氯喹可阻断Zac的效应。结论:Ⅰ_(K1)选择性激动剂Zac明显抑制异丙肾上腺素所致的心室重构,其机制可能为增强Ⅰ_(K1),进而增大RMP,缩短APD,从而阻断心肌细胞内钙超载依赖的信号通路。

Cardiac IK1 agonist inhibits isoproterenol-induced ventricular remodeling in rats

AIM: To investigate the effect of inward rectifier potassium channels (I_K1) agonist zacopride on isoproterenol (Iso)-induced ventricular remodeling and the involved mechanisms.METHODS: SD rats were randomly divided into 5 groups:control, Iso model, Iso+zacopride, Iso+zacopride+chloroquine and Iso+captopril groups. The model of cardiac hypertrophy was developed by intraperitoneal injection of Iso (3 mg·kg^-1·d^-1 for 10 d) and verified by determination of heart-to-body weight ratio and left ventricle-to-body weight ratio. The changes of voltage-gated calcium current (I_Ca-L), resting membrane potential (RMP) and action potential duration (APD) of the rat ventricular myocytes were detected by the technique of whole-cell patch clamp. Neonatal rat cardiomyocytes were isolated by 0.08% trypsin and 0.04% type II collagenase, and purified using differential adherence method and 5-bromodeoxyuridine. Cultured neonatal rat cardiomyocytes were randomly divided into 5 groups:control, Iso, Iso+zacopride, Iso+zacopride+BaCl₂ and Iso+zacopride+chloroquine groups. After harvested for 24 h, the Ca²⁺]_i of the cardiomyocytes was recorded using a laser confocal scanning microscope.RESULTS: Compared with control group, the whole heart hypertrophic index and left ventricular hypertrophic index in Iso group were increased significantly (P<0.01). Patch clamp data suggested that RMP was reduced, and APD was obviously prolonged. Zacopride treatment obviously inhibited myocardial hypertrophy, increased the RMP and shortened APD (P<0.01). At the same time, application of low dose of I_K1 atagonist chloroquine reversed the effect of zacopride. In cultured neonatal rat cardiomyocytes, Iso increased the cell area and Ca²⁺]_i. Zacopride treatment restored the hypertrophic morphology of the cells to normal or nearly normal levels, and significantly attenuated calcium overload. I_K1 blocker, BaCl₂ or chloroquine, reversed the effect of zacopride.CONCLUSION: I_K1 agonist zacopride significantly inhibits left ventricular remodeling caused by isoproterenol. Via enhancing I_K1, thereafter increasing RMP and shor-tening APD, zacopride might decrease Ca²⁺ influx and inhibit intracellular calcium overload-dependent signaling pathway.