兔心肌梗死1周及2月后心室肌细胞瞬间外向钾电流的变化

目的:研究心肌梗死后心室肌细胞瞬间外向钾电流(I_to)的变化。方法:采用结扎兔冠状动脉左前降支的方法复制心肌梗死动物模型,酶解法分离单个心室肌细胞,应用膜片钳全细胞记录方法,观察心肌梗死后1周及2月心外膜梗死区及非梗死区心室肌细胞I_to的变化。结果:①非梗死区2月组心肌细胞膜电容明显大于对照组,P＜0.05。②梗死区I_to电流密度(+60mV时)的对比显示:对照组为(17.4±5.2)pA/pF(n=16),梗死区1周组为(7.5±2.4)pA/pF(n=12),明显低于对照组(P＜0.01)。梗死区2月组为(10.6±4.1)pA/pF(n=18),明显低于对照组(P＜0.01),但高于梗死区1周组(P＜0.05)。③非梗死区2月组I_to电流密度为(13.2±4.1)pA/pF(n=23),明显低于对照组(P＜0.05),但高于梗死区2月组(P＜0.05)。结论:心肌梗死可引起心室肌细胞I_to电流密度下降,而且不同区域之间(梗死区及远离梗死区)也存在电生理的异质性,可能是导致心肌梗死后出现折返性室性心律失常的原因。心肌梗死后2月梗死区I_to的下降有恢复倾向。     

兔心肌梗死后心室肌Ito变化及雷米普利的作用

目的:探讨血管紧张素转化酶抑制剂雷米普利对兔心肌梗死后室性心律失常的影响及其可能机制。方法:24只家兔随机分为假手术组、心肌梗死组和雷米普利组。三组均在无菌条件下开胸,其中心肌梗死组和雷米普利组分别结扎左冠状动脉前降支。雷米普利组术后第二天给予雷米普利(1mg/kg/天)灌胃,三组均喂养12周。三组家兔分别在心肌梗死前、心肌梗死后12周记录程序刺激诱发的室性心动过速/室颤(VT/VF)发生次数,并采用全细胞膜片钳技术记录短暂外向钾电流(Ito)的变化。结果:心肌梗死后12周,雷米普利组较心肌梗死组VT/VF的发生次数明显减少(3.00±0.60vs 12.70±1.50,P0.05);假手术组、心肌梗死组和雷米普利组Ito电流密度分别为8.69±0.57、4.71±0.43和7.32±0.68pA/pF,心肌梗死组显著低于假手术组(P0.05);雷米普利组明显高于心肌梗死组,且与假手术组差异无统计学差异(P0.05)。结论:长期服用雷米普利可明显减少家兔心肌梗死后室性心动过速/室颤的发生,其机制可能与抑制家兔心肌梗死后Ito有关。     

丹皮酚对家兔心室肌细胞瞬时外向钾电流的抑制作用

目的 探讨丹皮酚(Paeonol,Pae)对家兔心室肌细胞瞬时外向钾电流(transient outward potassium current,Ito)及其动力学的影响.方法 用酶解法分离单个兔心室肌细胞.全细胞膜片钳技术记录Pae对兔心室肌细胞Ito的影响前后变化.结果 Pae 100 μg/ml作用心肌细胞5 min,可使Ito明显减小.指令电位为+50 mV时,可使Ito从(41.45±5.89)pA/pF减少到(8.97±2.78)pA/pF,两者之间有显著性差异(n=6,P＜0.01).冲洗15 min后,Ito部分恢复至(39.74±0.82)pA/pF(n=6,P＜0.01,与给药后比较),接近给药前的峰值,表明该药对Ito的抑制作用是可逆的.在50～400 μg/ml范围内,Pae对Ito的抑制作用表现为浓度依赖性,IC50的平均值为101.55 μg/ml.Pae 100 μg/ml使各膜电位水平下Ito减小,I-V曲线下移.Pae不改变Ito稳态激活动力学曲线.但使失活动力学曲线显著左移,即向超极化方向移动,50% Ito失活曲线点分别为(-45.3±3.5)mV和(-81.19±2.9)mV(n=8,P＜0.01),与对照组相比,差异有显著性.同时可使Ito失活后恢复时间延长,用药前后50% Ito恢复时间分别为(20.1±2.5)ms和(45.1±2.2)ms(n=8,P＜0.01),与对照组相比,有显著性差异.结论 Pae对家兔心室肌细胞Ito具有显著的抑制作用,这可能是Pae发挥抗心律失常作用的另一电生理基础.     

丹皮酚对家兔心室肌细胞时外向钾电流的抑制作用

目的 探讨丹皮酚(Paeonol,Pae)对家兔心室肌细胞瞬时外向钾电流(transient outward potassium current,Ito)及其动力学的影响.方法 用酶解法分离单个兔心室肌细胞.全细胞膜片钳技术记录Pae对兔心室肌细胞Ito的影响前后变化.结果 Pae 100 μg/ml作用心肌细胞5 min,可使Ito明显减小.指令电位为+50 mV时,可使Ito从(41.45±5.89)pA/pF减少到(8.97±2.78)pA/pF,两者之间有显著性差异(n=6,P＜0.01).冲洗15 min后,Ito部分恢复至(39.74±0.82)pA/pF(n=6,P＜0.01,与给药后比较),接近给药前的峰值,表明该药对Ito的抑制作用是可逆的.在50～400 μg/ml范围内,Pae对Ito的抑制作用表现为浓度依赖性,IC50的平均值为101.55 μg/ml.Pae 100 μg/ml使各膜电位水平下Ito减小,I-V曲线下移.Pae不改变Ito稳态激活动力学曲线.但使失活动力学曲线显著左移,即向超极化方向移动,50% Ito失活曲线点分别为(-45.3±3.5)mV和(-81.19±2.9)mV(n=8,P＜0.01),与对照组相比,差异有显著性.同时可使Ito失活后恢复时间延长,用药前后50% Ito恢复时间分别为(20.1±2.5)ms和(45.1±2.2)ms(n=8,P＜0.01),与对照组相比,有显著性差异.结论 Pae对家兔心室肌细胞Ito具有显著的抑制作用,这可能是Pae发挥抗心律失常作用的另一电生理基础.     

炙甘草汤含药血清对家兔心室肌细胞瞬时外向钾电流的抑制作用

目的:探讨炙甘草汤含药血清对家兔心室肌细胞瞬时外向钾电流(Ito)及其动力学的影响。方法:用酶解法分离单个兔心室肌细胞。血清药理学法确定血清中炙甘草汤含药浓度。全细胞膜片钳技术记录炙甘草汤含药血清对兔心室肌细胞Ito的影响前后变化。结果:20%炙甘草汤含药血清作用心肌细胞5min,可使Ito明显减小。指令电位为+50mV时,可使Ito从(43.3±5.89)pA/pF减少到(7.98±2.78)pA/pF(n=6,P<0.01)。该含药血清对Ito的抑制作用可逆。在5%~40%范围内,该含药血清对Ito的抑制作用为浓度依赖性,IC50的平均值为12.18%。20%该含药血清使各膜电位水平下Ito减小,I-V曲线下移。不改变Ito稳态激活动力学曲线,但使失活动力学曲线显著左移。50%Ito失活曲线点分别为(-41.4±3.5)mV和(-79.9±2.9)mV(n=8,P<0.05)。同时使Ito失活后恢复时间延长,用药前后50%Ito恢复时间分别为(18.2±2.5)ms和(44.1±2.2)ms(n=8,P<0.01)。结论:炙甘草汤含药血清对家兔心室肌细胞Ito具有显著的抑制作用,这可能是炙甘草汤抗心律失常的可能电生理基础之一。     

兔左室跨壁心肌细胞快钠电流和瞬间外向钾电流的异质性

目的进一步了解心室跨壁电生理异质性的细胞学机制。方法以酶解法分离兔左心室心外膜(Epi)、中层(M)和心内膜层(Endo)心肌细胞,应用膜片钳技术研究左心室钠电流(INa)和瞬间外向钾电流(Ito)的跨壁异质性。结果①三层细胞的INa均呈电压依赖性,当电压相同时M细胞的电流密度较大;M细胞的INa失活最快,与Epi、Endo相比P<0.05;三层细胞的INa灭活后再恢复曲线无明显差异。②三层细胞的Ito均呈电压依赖性激活过程,当电压相同时Epi细胞的电流密度较大;Epi的失活较快,M细胞的Ito灭活后再恢复未见明显变化。结论兔左室游离壁三层心肌细胞INa和Ito的分布及动力学特性存在异质性,这可能是动作电位形态差异及折返性心律失常发生的离子基础。     

心肌肽素抑制大鼠心室肌细胞瞬时外向钾电流

目的研究新药心肌肽素对大鼠心室肌细胞瞬时外向钾电流(Ito)的影响及其在离子通道水平的作用机制。方法用急性酶解法分离大鼠心室肌细胞,用标准的全细胞膜片钳技术,观察不同浓度的心肌肽素对Ito的影响。结果心肌肽素呈浓度依赖性抑制大鼠心室肌细胞Ito,心肌肽素浓度(mg/L)为10、50、100、250和500时分别使Ito降低(%)4、13、22、32和38。心肌肽素50 mg/L使Ito电流密度-电压曲线下移,但不改变曲线的形状,也不改变Ito稳态激活曲线。结论心肌肽素抑制大鼠心室肌细胞Ito,可能是其抗心律失常作用的机制之一。     

Molecular components of transient outward potassium current in cultured neonatal rat ventricular myocytes

We have previously reported that K(v1.4), K(v4.2), and K(v4.3) mRNAs are present in adult and neonatal rat ventricular myocytes, and that transient outward potassium current (I(to)) recovers from inactivation with a slow (I(to,s)) and a fast (I(to,f)) time course. This study was designed to determine the molecular correlates of I(to,s) and I(to,f) in cultured neonatal rat ventricular myocytes (NRVM) employing dominant-negative adenoviral infections to manipulate the function of endogenous I(to)-encoding K+ channels. Western blot data from cultured NRVM showed that K(v1.4), K(v4.2), and K(v4.3) channel proteins are present in these myocytes. The biphasic recovery from inactivation of I(to) in control GFP-infected myocytes demonstrated equal contribution of I(to,s) and I(to,f) in NRVM. Infection of cultured NRVM with adenoviruses expressing full-length K(v1.4) or K(v4.2) genes generated currents with recovery from inactivation kinetics similar to native I(to,s) and I(to,f) in GFP-infected myocytes, respectively. Overexpression of dominant-negative truncated K(v1.4) transgene (K(v1.4)N) caused a 51% reduction in I(to), selectively removing the slowly recovering I(to,s). Overexpression of dominant-negative K(v4.2)N reduced I(to) by 53% and eliminated the fast-recovering I(to,f). Our results establish that, in neonatal rat ventricular myocytes, the shaker K(v1) family (probably K(v1.4) and/or K(v1.7)) underlies I(to,s), and that the shal K(v4) family (probably K(v4.2) and K(v4.3)) is responsible for I(to,f).     

Reduction of calcium-independent transient outward potassium current density in DOCA salt hypertrophied rat ventricular myocytes

Saline-drinking, left-nephrectomized rats made hypertensive by deoxycorticosterone acetate (DOCA) pellet implantation at the time of surgery develop a cardiac hypertrophy, which becomes maximal after 6–7 weeks. The hypertrophy results in a marked increase in the amplitude and duration of both the early and the late component of the ventricular action potential plateau recorded in the isolated perfused rat heart. The 4-aminopyridine(4-AP)-sensitive calcium-independent transient outward potassium current was markedly depressed in hypertrophied ventricular myocytes resulting in a highly significant decrease in current density (from 19.9±3.5 to 6.4±3.1 pA/pF at +60 mV). Activation/ voltage and steady-state inactivation/voltage relationships were moderately although non-significantly shifted towards negative potentials. The steady-state outward current measured at the end of 1-s depolarizing pulses was not significantly changed in hypertrophied myocytes. 4-AP induced a smaller increase in plateau amplitude and duration in hypertrophied rather than in control hearts, a point that is well explained by the depression of the transient outward current resulting from hypertrophy. We also demonstrated that a complete recovery of both cell capacitance and transient outward current amplitude occurs in myocytes from saline-drinking rats studied 13 weeks after DOCA pellet implantation, showing that hypertrophy regresses as a result of pellet elimination. Several mechanisms can be involved in the observed phenomena, including the possibility that the expression of potassium channels responsible for the transient outward current is not enhanced by hypertrophy in contrast with what occurs in the case of calcium channels. We conclude that the depression of the calcium-independent transient outward potassium current appears responsible for the major part of the hypertrophy-induced action potential lengthening in rat ventricular myocytes.     

Effect of trimetazidine treatment on the transient outward potassium current of the left ventricular myocytes of rats with streptozotocin-induced type 1 diabetes mellitus

Cardiovascular complications are a leading cause of mortality in patients with diabetes mellitus (DM). The present study was designed to investigate the effects of trimetazidine (TMZ), an anti-angina drug, on transient outward potassium current (I_to) remodeling in ventricular myocytes and the plasma contents of free fatty acid (FFA) and glucose in DM. Sprague-Dawley rats, 8 weeks old and weighing 200-250 g, were randomly divided into three groups of 20 animals each. The control group was injected with vehicle (1 mM citrate buffer), the DM group was injected with 65 mg/kg streptozotocin (STZ) for induction of type 1 DM, and the DM+TMZ group was injected with the same dose of STZ followed by a 4-week treatment with TMZ (60 mg·kg⁻¹·day⁻¹). All animals were then euthanized and their hearts excised and subjected to electrophysiological measurements or gene expression analyses. TMZ exposure significantly reversed the increased plasma FFA level in diabetic rats, but failed to change the plasma glucose level. The amplitude of I_to was significantly decreased in left ventricular myocytes from diabetic rats relative to control animals (6.25 ± 1.45 vs 20.72 ± 2.93 pA/pF at +40 mV). The DM-associated I_to reduction was attenuated by TMZ. Moreover, TMZ treatment reversed the increased expression of the channel-forming alpha subunit Kv1.4 and the decreased expression of Kv4.2 and Kv4.3 in diabetic rat hearts. These data demonstrate that TMZ can normalize, or partially normalize, the increased plasma FFA content, the reduced I_to of ventricular myocytes, and the altered expression Kv1.4, Kv4.2, and Kv4.3 in type 1 DM.     

辛伐他汀对血脂正常兔心肌缺血再灌注后瞬间外向钾通道电流的影响

目的： 观察辛伐他汀预处理对兔心肌缺血再灌注后瞬间外向钾通道电流（Ito）的影响,探讨他汀类药物抗心律失常的细胞学离子机制。方法: 45只新西兰大耳白兔随机分为3组：缺血再灌注动物模型组（I－R组,结扎冠脉左前降支30 min后再开放120 min）;辛伐他汀治疗组（他汀组,手术前给予辛伐他汀 5 mg·kg-1·d-1）;假手术对照组（只开胸不结扎血管）。采用酶解的方法分离缺血部位心室肌外膜单个心室肌细胞,应用全细胞膜片钳技术记录Ito,同时检测各组血脂水平。结果: 各组动物血脂水平无显著差异。Ito电流密度峰值（＋60 mV）显示：对照组为（17.41± 3.13）pA/pF(n=15),I－R组为（9.49 ±1.91） pA/pF(n=11),低于对照组（P＜0.01）。他汀组为（15.24 ± 2.41） pA/pF (n=11),显著高于I－R组（P＜0.01）,但仍明显小于对照组（P＜0.05）。另外,I－R组Ito失活曲线左移,失活后恢复时间延长,他汀组这些异常也明显恢复。结论: 本研究显示缺血再灌注可导致梗死区心肌细胞Ito明显下降,形成电重构,可引起心肌细胞动作电位延长,为再灌注心律失常的形成机制之一。辛伐他汀预处理可减轻Ito的异常变化,逆转电重构,而不依赖于降血脂效应,可能为他汀类药物降低心律失常发生率的细胞学离子机制。     

Calcitonin gene-related peptide suppresses the transient outward current in rat ventricular myocytes

The effects of calcitonin gene-related peptide (CGRP) on the transient outward current (Ito) and the L-type calcium current (ICa,L) were investigated in isolated rat ventricular cardiomyocytes using the whole-cell, patch-clamp technique. CGRP influenced neither the amplitude nor the time course of ICa,L. On the other hand, at all membrane potentials at which a significant Ito was elicited, CGRP decreased its amplitude. The effect on Ito was completely reversible and independent of membrane potential. The steady-state activation and inactivation curves of Ito were not influenced by CGRP. The time course of Ito inactivation was satisfactorily fitted by two exponentials. Both the fast and the slow time constants of inactivation were voltage independent and were not influenced by CGRP. The effect of CGRP on Ito was concentration dependent with half-maximum inhibition at 99 nM. Chelerythrine, a selective inhibitor of protein kinase C, prevented the effect of CGRP on Ito. The data indicate that CGRP suppresses Ito in a concentration-dependent, but membrane potential-independent manner and that the effect is probably mediated via a protein kinase C-dependent pathway.     

糖尿病兔心肌梗死后钾离子通道蛋白基因表达的变化

目的: 探讨糖尿病合并心肌梗死后兔心室肌钾离子通道蛋白基因表达的变化。方法: 选择新西兰兔作为研究对象,通过高脂高糖喂养2个月后由耳缘静脉注射四氧嘧啶(80 mg/kg)制作糖尿病模型, 成模后改为普通饲料继续单笼喂养4个月,并随机分为糖尿病+心肌梗死组、糖尿病假手术组和非糖尿病假手术组,通过结扎冠状动脉前降支制作心肌梗死模型。手术后所有入选成活兔均常规喂养2个月后处死,取左心室梗死周围区心室肌采用实时荧光定量PCR方法观察心肌钾离子通道蛋白Kv4.2、Kv4.3和Kv1.4 mRNA表达变化。结果: 糖尿病+心肌梗死组和糖尿病假手术组兔心室肌Kv4.2和Kv4.3 mRNA表达均显著低于非糖尿病假手术组(P<0.05);而Kv1.4显著高于非糖尿病假手术组(P<0.05)。糖尿病+心肌梗死组兔心室肌Kv4.2和Kv4.3 mRNA表达显著低于糖尿病假手术组(P<0.05);而Kv1.4并无显著升高(P>0.05)。结论: 糖尿病兔心肌梗死后瞬时外向钾离子通道蛋白基因表达发生了改变即电重构,这可能是心肌梗死后室性心律失常易感性增加的机制之一。     

A sodium-activated potassium current in intact ventricular myocytes isolated from the guinea-pig heart

A current generated by the Na-activated K channel has been identified in whole cell currents recorded from isolated guinea-pig ventricular myocytes. A partial activation of this current can be achieved near to the physiological range of intracellular sodium concentration [( Na+]i) when it contributes significantly to the global outward current. The decline of the Na-activated K current, the lengthening of the action potential duration and the recovery of [Na+]i occur with a similar time course during recovery from Na loading.