苄基四氢巴马汀对豚鼠心室肌细胞快激活延迟整流钾电流的作用

目的研究苄基四氢巴马汀(BTHP)对心室肌细胞快激活(I_kr)延迟整流钾电流的作用。方法 用全细胞膜片钳技术记录豚鼠心室肌细胞钾离子通道电流。结果BTHP在1～100 μmol·L^-1以浓度依赖性方式阻滞I_kr,其IC₅₀为13.5 μmol·L^-1(95%可信范围:11.2～15.8 μmol·L^-1)。30 μmol·L^-1 BTHP可使I_kr及I_kr,tail分别降低(31±4)%和(36±5)% (N=6,P<0.01)。与多数III类抗心律失常药物不同,BTHP可频率依赖性地抑制I_kr。该药主要改变I_kr的失活过程,可使I_kr的失活时间常数(τ)从(238±16) ms降至(196±14) ms,而对I_kr的激活动力学影响不大。结论BTHP对I_kr有明显的抑制作用,且其阻滞作用呈现频率依赖性特征。     

阿米洛利对豚鼠心肌细胞钾电流及钙电流的作用

目的研究阿米洛利（amiloride）对豚鼠心肌细胞钾电流及钙电流的作用。方法采用全细胞膜片钳技术记录豚鼠心室肌细胞钾通道及钙通道电流。结果阿米洛利在10~100 μmol·L^-1抑制L型及T型钙电流,不改变钙电流I-V曲线的形状，仅抑制这两型电流的幅度。当累积浓度达100 μmol·L^-1时，阿米洛利轻微抑制快激活延迟整流钾电流(I_Kr)，对慢激活延迟整流钾电流(I_Ks)无影响。阿米洛利在1~100 μmol·L^-1浓度依赖性地抑制内向整流钾电流(I_K1)。结论阿米洛利抑制电压依赖性的钾、钙电流，为其抗心律失常作用提供了离子基础。     

苄普地尔对豚鼠甲亢性肥厚心肌中延迟整流钾电流的抑制作用

目的　研究苄普地尔(bepridil)对肥厚心肌细胞延迟整流钾电流(I_K)中快激活成份(I_Kr)和慢激活成份(I_Ks)及内向整流钾电流(I_K1)的作用。方法　全细胞膜片钳技术。结果　在肥厚心肌细胞中,Bepridil 30 μmol·L- 1 对I_Kr及I_Ks有阻断作用,抑制率分别为20.9% (0 mV)和27.2 % (+50 mV)。“Envelopeoftail”显示bepridil对I_Ks的阻断作用大于I_Kr。Bepridil(1 - 100 μmol·L^-1 )浓度依赖性的阻断I_Ks及I_Kr,其IC₅₀ 分别为23.8μmol·L^-1 和46.7μmol·L^-1 。Bepridil 30 μmol·L^-1 也能阻断I_K1 ,抑制率为15.1% (- 100 mV) ,但不影响其反转电位。结论　Bepridil对甲亢性豚鼠肥厚心肌中I_Ks,I_Kr及I_K1有阻断作用     

苄基四氢巴马汀细胞内用药对豚鼠乳头状肌动作电位和心室肌细胞延迟整流钾电流的作用

目的　研究将苄基四氢巴马汀(BTHP)导入细胞内对豚鼠乳头状肌动作电位及单个心室肌细胞延迟整流钾电流的影响。方法　利用外加电压脉冲将药物导入乳头状肌细胞内，并用标准微电极方法测定动作电位；利用浓度差扩散方式使药物进入单个心室肌细胞内，采用全细胞膜片钳技术记录延迟整流钾电流（I_K）。结果　100 μmol.L^-1 BTHP使APD₂₀和APD₉₀分别延长13.5%和20.5%。30 μmol.L^-1 BTHP使I_K和I_K,tail分别从（14.1±2.2) pA.pF^-1和(4.0±0.6) pA.pF^-1降至(9.4±1.3) pA.pF^-1和(2.1±1.0) pA.pF^-1，下降率分别为33.2%和35.3%。 该药使I_K和I_K,tail的I-V曲线幅度降低，对曲线形状影响不明显。结论　BTHP入细胞内后可阻滞延迟整流钾电流和延长动作电位时程。     

用β-escin穿孔膜片技术研究粉防己碱对豚鼠心室肌钙电流的作用

目的用β-escin穿孔膜片(PPR)技术研究粉防己碱(tetrandrine,Tet)对I_Ca,L和I_Ca,T的作用。方法用PPR和全细胞记录(WCR)模式记录心室肌细胞I_Ca,L和I_Ca,T。结果25 μmol·L^-1 β-escin可在心室肌细胞形成稳定的PPR模式,用此模式记录的I_Ca,L衰减明显减慢。在PPR模式下1～300 μmol·L^-1 Tet浓度依赖性地减小I_Ca,L幅值。3,30和300 μmol·L^-1 Tet对I_Ca,T的抑制率分别为(16±5)%,(40±7)%和(75±11)%。结论用25 μmol·L^-1 β-escin在豚鼠心室肌细胞能得到较稳定的PPR模式,在此模式下Tet浓度依赖性地抑制I_Ca,L和I_Ca,T。     

4-氨基吡啶对豚鼠心室肌钙和钠电流的影响

目的研究4-氨基吡啶(4-AP)对心肌细胞L型钙通道和钠通道的影响。方法用全细胞膜片钳技术考察4-AP对豚鼠心室肌细胞L型钙电流和钠电流的作用。结果4-AP0.1,0.5,1.0mmol·L^-1浓度依赖性地抑制L型钙电流(I_Ca,L)和钠电流(I_Na),抑制率分别为(11.6±1.7)%,(37.5±8.3)%和(54.5±6.9)%以及(22.1±14.3)%,(39.4±8.8)%和(62.3±6.8)%。0.5mmol·L^-14-AP使I_Ca,L和I_NaI-V曲线均上移。结论4-AP可浓度依赖性地阻滞豚鼠心室肌细胞L型钙通道和钠通道。     

抗疟药青蒿素抗心律失常的作用机制

目的：探讨青蒿素抗心律失常的离子电流基础。方法：用全细胞膜片钳技术和双电极电压钳技术。结果：当细胞超极化到-100 mV时,青蒿素以浓度依赖方式明显抑制家兔心室肌细胞I_k1,50μmol.L^-1青蒿素可使家兔心室肌细胞I_k1从对照组的-2.36±0.39　nA减少到-1.43±0.31nA。给予非洲蛙卵母细胞注射K_{ir 2.1}　cRNA后,用不同浓度青蒿素灌注,可减低K_{ir 2.1}钾通道电流,此作用呈电压和浓度依赖性。青蒿素对K_{ir 2.1}钾通道的阻断作用呈可逆性。结论：青蒿素能有效抑制离体心肌细胞I_k1,其抗心律失常作用机理与其抑制心肌细胞I_k1及阻断K_{ir 2.1}通道电流有关。     

苦参碱、小檗胺与胺碘酮、RP58866抗心律失常作用的比较

目的阐明苦参碱和小檗胺抗心律失常作用弱于胺碘酮和RP58866的分子机制。方法采用冠脉结扎、电刺激和乌头碱诱导的心律失常模型观察药物的抗心律失常作用，采用全细胞膜片钳技术测定单个心室肌细胞的I_K1，I_Kr，I_Ks和 I_to。结果苦参碱和小檗胺对冠脉结扎、乌头碱诱发的大鼠心律失常有明显对抗作用，对家兔电刺激致颤阈（VFT）有明显提高作用，但与胺碘酮和RP58866相比，抗心律失常作用明显低于前者。电生理结果显示：苦参碱和小檗胺对家兔I_K1,I_Kr,I_Ks和犬I_to有抑制作用，但较胺碘酮和RP58866作用弱。结论苦参碱和小檗胺的抗心律失常作用及对I_K1,I_Kr,I_Ks和I_to的抑制作用弱于胺碘酮和RP58866。     

妥卡尼对豚鼠心室肌细胞钙电流和钾电流的抑制作用

利用酶分散的成年豚鼠心室肌细胞和全细胞电压钳技术,研究了妥卡尼(tocainide)对心室肌细胞钙电流(I_ca)、延迟整流钾电流(I_k)和ATP敏感性钾电流(Ik,ATP)的作用。结果表明,妥卡尼对I_ca和I_k均显示浓度相关的抑制作用,妥卡尼50umol·L^-1对I_ca和I_k的抑制率分别为16%和3%。这可能是妥卡尼有效抑制室上性心动过速和缩短心肌动作电位平台期的重要机制。     

III类抗心律失常药RP58866对豚鼠及犬内向整流及瞬时外向钾电流的作用

目的:研究III类抗心律失常药RP58866 对I_K1 ,瞬时外向钾电流(Ito) 的作用。方法:用豚鼠和犬离体心肌细胞及全细胞电压钳技术。结果:在- 100 m V 时,RP58866 以浓度依赖方式明显减少了豚鼠心室肌细胞I_K1,其IC₅₀为(3-4±0-8) μmol·L^-1。在犬心室肌细胞,RP58866 可明显抑制Ito( 在100 μmol·L^-1 时减少87% ±2-1% ),其IC₅₀为(2-3±0-5) μmol·L^-1 。结论:RP58866 对心肌细胞的IK1 和Ito 均有抑制作用,而不是一种特殊的IK1抑制剂。     

Inhibitory effect of gallic acid on voltage-gated Na+ channels in rat cardiomyocytes

Gallic acid (GA) has a protective effect on the cardiovascular system. To study its cardiac electrophysiological effects, voltage-gated Na⁺ channel currents (I_Na) were recorded in rat cardiomyocytes using whole-cell patch clamp techniques. Moreover, the effects of GA on aconitine-induced arrhythmias were assessed using electrocardiograms in vivo. We found that the current–voltage characteristic curve (I-V curve) of I_Na significantly shifted in the presence of 1, 3, and 10 μmol/L of GA. The peak sodium current density (I_Na-Peak) was reduced from −84.02 ± 5.68 pA/pF to −65.78 ± 3.96 pA/pF with 1 μmol/L, −54.45 ± 5.18 pA/pF with 3 μmol/L, and −44.20 ± 4.35 pA/pF with 10 μmol/L, respectively. GA shifted the steady-state activation curve of I_Na and recovery curve to the right and the steady-state inactivation curve to the left. The observed inhibitory effect was comparable to that of amiodarone. GA pre-treatment significantly prolonged the onset of fatal ventricular fibrillation. Our results indicated that GA inhibited I_Na in rat ventricular myocytes and aconitine-induced arrhythmias in vivo. These results suggest the potential of GA for development as a novel anti-arrhythmic therapeutic.     

Pharmacological evidence for altered src kinase regulation of <Emphasis Type="Italic">I</Emphasis><Subscript>Ca,L</Subscript> in patients with chronic atrial fibrillation

A reduction in l-type Ca²⁺ current (I _Ca,L) contributes to electrical remodeling in chronic atrial fibrillation (AF). Whether the decrease in I _Ca,L is solely due to a reduction in channel proteins remains controversial. Protein tyrosine kinases (PTK) have been described as potent modulators of I _Ca,L in cardiomyocytes. We studied α_1C l-type Ca²⁺ channel subunit expression and the regulation of I _Ca,L by PTK in chronic AF using PTK inhibitors: genistein, a nonselective inhibitor of PTK, and 4-amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo-3,4-d-pyrimidine (PP1), a selective inhibitor of src kinases. Furthermore, type-1 and type-2A protein phosphatase activity was measured with phosphorylase as substrate in whole-cell lysates derived from atrial tissue of AF patients. Right atrial appendages were obtained from patients undergoing open-heart surgery. Protein levels of α_1C l-type Ca²⁺ channel subunit were determined using Western blot analysis and normalized to the protein amounts of calsequestrin as internal control. The protein concentrations of α_1C did not differ between AF and sinus rhythm (SR; α_1C/calsequestrin: 1.0 ± 0.1 and 1.2 ± 0.2, respectively, n = 8 patients). In cardiomyocytes from patients in SR (n = 20 patients), genistein and PP1 both evoked similar increases in I _Ca,L from 3.0 ± 0.3 to 6.1 ± 0.8 pA/pF and from 2.8 ± 0.4 to 6.1 ± 0.6 pA/pF, respectively. In cells from AF patients (n = 10 patients), basal I _Ca,L was significantly lower. In this case, genistein lead to the same relative increase in I _Ca,L as in SR cells (from 1.46 ± 0.30 to 3.2 ± 1.0 pA/pF), whereas no increase was elicited by PP1 suggesting impaired regulation of I _Ca,L by src kinases in AF. Total and type 1 and type 2A-related phosphatase activities were higher in tissue from patients with chronic AF compared to SR (4.8 ± 0.4, 2.1 ± 0.2, and 2.7 ± 0.4 nmol/mg/min and 3.6 ± 0.4, 1.3 ± 0.2, and 2.4 ± 0.3 nmol/mg/min, respectively, n = 7 patients per group). Downregulation of I _Ca,L in AF is not due to a reduction in l-type Ca²⁺ channel protein expression. Indirect evidence for an impaired src kinase regulation of I _Ca,L together with an increased phosphatase activity suggests that a complex alteration in the kinase/phosphatase balance leads to I _Ca,L dysregulation in chronic AF.     

苄基四氢巴马汀对表达于非洲爪蟾卵母细胞及中华大蟾蜍卵母细胞的延迟整流钾电流的抑制作用

苄基四氢巴马汀对表达于非洲爪蟾卵母细胞及中华大蟾蜍卵母细胞的延迟整流钾电流的抑制作用童秋生，夏国瑾，姚伟星，江明性，白小川，包永德（同济医科大学基础医学院药理学教研室，武汉４３００３０；中国科学院上海生理学研究所，上海２０００３１）苄基四氢巴马汀（ｂ...     

Risperidone-induced action potential prolongation is attenuated by increased repolarization reserve due to concomitant block of <Emphasis Type="Italic">I</Emphasis><Subscript>Ca,L</Subscript>

The neuroleptic risperidone is an effective blocker of the rapidly activating component of the delayed rectifier current (I_Kr) and hence is expected to prolong cardiac action potential duration (APD). However, unlike with other typical I_Kr blockers we failed to demonstrate a marked prolongation of late repolarization with risperidone. It is hypothesized that the APD-prolonging effect of risperidone is masked by the high repolarization reserve due to the prominent delayed rectifier currents I_Kr and I_Ks in guinea pig papillary muscle. Action potentials and force of contraction were recorded in isolated guinea pig papillary muscles. L-type calcium current I_Ca,L and I_Kr were measured using the standard patch clamp technique in single ventricular cardiomyocytes. Reduction of the repolarization reserve by the blocking of I_Ks with chromanol 239B augmented the effect of the selective I_Kr blocker E-4031, but not of risperidone, although both drugs completely blocked I_Kr. In contrast to E-4031 risperidone markedly reduced the force of contraction due to the partial blocking of I_Ca,L in the same concentration range as required for block of I_Kr. Reduction of the repolarization reserve by the blocking of I_Ks cannot exacerbate the APD-prolonging effect of risperidone. However, even incomplete concomitant blocking of I_Ca,L attenuates the APD-prolonging effect of the complete blocking of I_Kr. This behaviour may explain the small APD-prolonging effect of risperidone despite the drugs robust blocking of I_Kr.T. Christ and E. Wettwer contributed equally to this work     

盐酸非洛普对豚鼠心室肌细胞延迟整流钾电流的抑制作用

目的　已知盐酸非洛普〔1 (2 ,6 二甲基苯氧基 ) 2 (3,4 二甲氧基苯乙氨基 )丙烷盐酸盐 ,DDPH〕对心肌钙电流和钠电流具有抑制作用 ,为全面了解其抗心律失常作用的离子机理 ,研究其对延迟整流钾电流的影响。方法　全细胞膜片钳技术记录豚鼠心室肌细胞快激活的延迟整流钾电流的尾电流(IKr tail)和慢激活的延迟整流钾电流 (IKs)及其尾电流 (IKs tail)。结果　DDPH(1～ 10 0 μmol·L- 1)浓度依赖性地抑制IKr tail,其IC50 为 7.0 (95 %可信限为4 .2 3～ 9.76 ) μmol·L- 1;DDPH 10 μmol·L- 1对IKr tail具有电压依赖性抑制作用。DDPH 10 ,30和 10 0 μmol·L- 1可浓度依赖性地抑制IKs及其IKs tail,使IKs从给药前的 (9.1± 0 .7)pA·pF- 1分别降至 (7.7± 1.7) ,(7.5± 1.8)和 (5 .6± 1.8)pA·pF- 1(P <0 .0 1) ;使IKs tail从给药前的 (1.4± 0 .2 )pA·pF- 1分别降至(1.1± 0 .2 ) ,(0 .9± 0 .2 )和 (0 .6± 0 .2 )pA·pF- 1(P <0 .0 5或P <0 .0 1) ;DDPH 30 μmol·L- 1对IKs tail具有电压依赖性抑制作用。结论　DDPH对豚鼠心室肌细胞延迟整钾电流具有抑制作用。     

大鼠肺内动脉平滑肌细胞钾通道的研究

目的　研究大鼠肺内动脉平滑肌细胞钾电流。方法　用急性酶解分离法得到单个大鼠肺内动脉平滑肌细胞,采用膜片钳全细胞记录方式研究钾通道特性。结果　将平滑肌细胞钳制在-70 mV,给予-70～50 mV的斜坡刺激,时间为600 ms。可引出一随电压逐渐增大的电流,在+50 mV时其值为359±31 pA。细胞内用CsCl取代KCl后,该电流几乎完全消失;细胞外用无钙台氏液灌流,电极内液用高浓度的EGTA时,电流可被抑制50%±1%;细胞外给予特异性钙激活钾通道阻断剂TEA和延迟整流钾通道阻断剂4-AP均可使该电流明显下降。结论　大鼠肺内动脉平滑肌细胞的钾电流主要由钙激活钾电流和延迟整流钾电流组成。     

Asynchronous activation of calcium and potassium currents by isoproterenol in canine ventricular myocytes

Adrenergic activation of L-type Ca²⁺ and various K⁺ currents is a crucial mechanism of cardiac adaptation; however, it may carry a substantial proarrhythmic risk as well. The aim of the present work was to study the timing of activation of Ca²⁺ and K⁺ currents in isolated canine ventricular cells in response to exposure to isoproterenol (ISO). Whole cell configuration of the patch-clamp technique in either conventional voltage clamp or action potential voltage clamp modes were used to monitor I _Ca, I _Ks, and I _Kr, while action potentials were recorded using sharp microelectrodes. ISO (10 nM) elevated the plateau potential and shortened action potential duration (APD) in subepicardial and mid-myocardial cells, which effects were associated with multifold enhancement of I _Ca and I _Ks and moderate stimulation of I _Kr. The ISO-induced plateau shift and I _Ca increase developed faster than the shortening of APD and stimulation of I _Ks and I _Kr. Blockade of β₁-adrenoceptors (using 300 nM CGP-20712A) converted the ISO-induced shortening of APD to lengthening, decreased its latency, and reduced the plateau shift. In contrast, blockade of β₂-adrenoceptors (by 50 nM ICI 118,551) augmented the APD-shortening effect and increased the latency of plateau shift without altering its magnitude. All effects of ISO were prevented by simultaneous blockade of both receptor types. Inhibition of phosphodiesterases decreased the differences observed in the turn on of the ISO-induced plateau shift and APD shortening. ISO-induced activation of I _Ca is turned on faster than the stimulation of I _Ks and I _Kr in canine ventricular cells due to the involvement of different adrenergic pathways and compartmentalization.     

Electrophysiological profile of propiverine – relationship to cardiac risk

Drugs that prolong the QT interval by blocking human ether-a-go-go (HERG) channels may enhance the risk of ventricular arrhythmia. The spasmolytic drug propiverine is widely used for the therapy of overactive bladder (OAB). Here, we have investigated the effects of propiverine on cardiac ion channels and action potentials as well as on contractile properties of cardiac tissue, in order to estimate its cardiac safety profile, because other drugs used in this indication had to be withdrawn due to safety reasons. Whole-cell patch clamp technique was used to record the following cardiac ion currents: rapidly and slowly activating delayed rectifier K⁺ current (I_Kr, I_Ks), ultra rapidly activating delayed rectifier K⁺ current (I_Kur), inwardly rectifying K⁺ current I_K1, transient outward K⁺ current (I_to), and L-type Ca²⁺ current (I_Ca,L). Action potentials in cardiac tissue biopsies were recorded with conventional microelectrodes. The torsade de pointes screening assay (TDPScreen^TM) was used for drug scoring. Propiverine blocked in a concentration-dependent manner HERG channels expressed in HEK293 cells, as well as native I_Kr current in ventricular myocytes of guinea pig (IC₅₀ values: 10 μM and 1.8 μM respectively). At high concentrations (100 μM), propiverine suppressed I_Ks. I_K1 and the transient outward current I_to and I_Kur were not affected. In guinea-pig ventricular and human atrial myocytes, propiverine also blocked I_Ca,L (IC₅₀ values: 34.7 μM and 41.7 μM, respectively) and reduced force of contraction. Despite block of I_Kr, action potential duration was not prolonged in guinea-pig and human ventricular tissue, but decreased progressively until excitation failed altogether. Similar effects were observed in dog Purkinje fibers. Propiverine obtained a low score in the TDPScreen^TM. In conclusion, in vitro and in vivo studies of propiverine do not provide evidence for an enhanced cardiovascular safety risk. We propose that lack of torsadogenic risk of propiverine is related to enhancement of repolarization reserve by block of I_Ca,L.