血管钠肽对大鼠肠系膜动脉平滑肌细胞Ca2+激活K+通道的作用

目的研究血管钠肽(VNP)对大鼠肠系膜动脉血管平滑肌细胞(VSMCs)Ca2+激活K+通道(Kca)的作用及其机制.方法采用全细胞膜片钳技术观察VNP对Kca的影响,以及HS-142-1、8-Br-cGMP和美蓝(MB)在这一过程中的作用.结果①VNP(10-6 mol/L)显著增强Kca(P＜0.05,n=5).②8-Br-CGMP(10-3mol/L)模拟VNP增强Kca的作用(P＜0.05,n=6).③HS-142-1(2×10-5mol/L)或MB(10-5mol/L)完全阻断VNP增加Kca电流密度的作用.结论VNP通过作用于VSMCs的钠尿肽GC耦联受体,升高细胞内的cGMP水平,激活Kca.     

血管钠肽对离体人乳内动脉的舒张作用

为了研究血管钠肽(VNP)对人乳内动脉(human intramammary artery,HIMA)的舒张作用及其机制,采用离体血管灌流的方法,观察VNP对内皮完整和去内皮HIMA的舒张作用,以及HS—142—1、TEA、8—Br—cGMP和镁蓝(MB)对这一过程的影响。实验中观察到,VNP(0．0001—1μmol/L)可引起剂量依赖性的舒张效应,且无内皮依赖性;8—Br—cGMP(0．1—1000μmol/L)也可引起剂量依赖性的血管舒张效应。钠尿肽鸟苷酸环化酶(guanylate cyclase,GC)受体的特异性阻断剂HS—142—1(20μmol/L)使VNP舒张HIMA的作用几乎完全消失。MB是GC的抑制剂,10μmol/L的MB不但使VNP舒张HIMA的作用完全消失,而且可增强HIMA对去甲肾上腺素(NE)产生的收缩反应。钙激活钾通道(KCa)的阻断剂TEA(1mmol/L)可减弱(但是不完全阻断)VNP的舒血管作用。上述结果表明,VNP对HIMA具有不依赖内皮的舒张作用;此作用是通过作用于平滑肌细胞的钠尿肽GC受体,引起细胞内的cGMP水平升高实现的,并且与Kca有关。     

腺苷抗豚鼠室性心律失常的电生理研究

实验用全细胞膜片钳技术在单个豚鼠心室肌细胞上研究了腺苷 (Ado)对正常及异丙肾上腺素 (Iso)致豚鼠心室肌细胞动作电位、迟后除极 (DAD)、L 型钙电流 (ICa.L)和短暂内向电流 (Iti)的作用。结果表明 :(1)Ado在2 0～ 10 0 μmol/L时对豚鼠心室肌细胞动作电位和ICa .L无明显直接作用 ,但却可明显降低Iso所致的动作电位时程(APD)延长和ICa .L峰值增大 ,Iso (10nmol/L)使细胞APD50 从 3 40± 2 1ms延长到 486± 2 8ms (P <0 0 1) ,APD90从 3 61± 17ms延长至 5 0 1± 2 9ms (P <0 0 1) ;ICa .L峰值从 - 6 5 3± 1 4pA/pF增大到 - 18 2 8± 2 4pA/pF (P <0 0 1) ,电流电压曲线明显左移和下移 ;Ado (5 0 μmol/L)使APD50 和APD90 降至 40 3± 19ms和 419± 2 6ms ,但并不影响动作电位其它参数 ,使ICa.L峰值降低至 - 10 2± 1 5pA/pF (P <0 0 1)。 (2 )Iso (3 0nmol/L)可诱发心室肌细胞产生DADs,其发生率为 10 0 % ;Ado (5 0 μmol/L)可完全抑制Iso引发DADs;细胞经 - 40～ +2 0mV、时程 2s的除极电压 ,Iso (3 0nmol/L)诱导出Iti,其发生率为 10 0 % ;Ado (5 0 μmol/L)可明显抑制Iso致Iti的发生 ,其发生率降为 14 3 %。研究结果提示 ,Ado对豚鼠心室肌细胞动作电位和ICa.L无明显直接作用 ,但却可显著降低Is     

去甲肾上腺素、乙酰胆碱对绵羊心肌浦肯野纤维瞬时性内向离子流的影响

用电压箝制术观察了去甲肾上腺素、乙酰胆碱对绵羊浦肯野纤维由乙酰毒毛旋花子甙元诱发的瞬时性内向离子流(I_Ti)的效应。当乙酰毒毛旋花子甙元浓度为4.5×10~(-8)mol/L 时,诱发出的I_(T1)稳定并能维持约1.5h。去甲肾上腺素1.5×10~(-6)mol/L,可使I_(Ti)的峰值由11.4±2.5nA 增加到14.5±4.1nA(n=11,P相似文献   

血管钠肽抑制低氧对大鼠心成纤维细胞内钙浓度升高的作用

为了观察血管钠肽(vasonatrinpeptide,VNP)对低氧作用时心成纤维细胞增殖的影响,将分离纯化乳鼠心成纤维细胞,随机分为4组对照组、低氧组(2%～3%)、VNP组(10－8～10－6mol/L)和VNP+低氧组。用MTT比色法、～3H-TdR掺入法观察细胞增殖情况,采用激光共聚焦方法研究VNP对细胞内钙浓度(［Ca     

Amiloride和Ni2+抑制缺血/复灌引起的大鼠心肌细胞内钙的增加

本文旨在研究Na+/H+交换以及Na+/Ca2 +交换对模拟缺血 /复灌引起的大鼠心肌细胞内游离钙水平变化的调节作用。分别利用模拟缺血液和正常台氏液对大鼠心肌细胞进行缺血 /复灌处理 ,在缺血期间分别应用Na+/H+交换抑制剂阿米洛利 (amiloride)、Na+/Ca2 +交换抑制剂NiCl2 以及无钙液 ,观察它们对细胞内游离Ca2 +浓度变化的影响。利用Zeiss LSM 5 10激光共聚焦显微镜检测、采集细胞内游离Ca2 +的指示剂Fluo 3 AM的荧光信号 ,计算出相对于正常(缺血前 )的相对荧光强度 ,以表示胞内游离Ca2 +浓度的变化。结果显示 ,模拟缺血引起大鼠心肌细胞内游离Ca2 +持续上升 ,缺血前的相对荧光强度值为 10 0 % ,模拟缺血 5min后为 140 3± 13 0 % (P <0 0 5 ) ,复灌 15min后为 142 8±15 5 % (P <0 0 5 )。经 10 0 μmol/Lamiloride、5mmol/LNiCl2 和无钙液分别预处理 ,模拟缺血 5min后的相对荧光强度分别为 10 1 4± 16 3 % (P <0 0 5 )、110 4± 11 1% (P <0 0 5 )和 10 7 1± 10 8(P <0 0 5 ) ;复灌 15min后则分别为 97 8±14 3 % (P <0 0 5 )、10 6 2± 14 5 % (P <0 0 5 )和 10 6 6± 15 7(P <0 0 5 )。另外 ,与对照组细胞相比 ,再灌注期间NiCl2和无钙液处理的细胞钙振荡的产生幅度明显减弱 ,amilorid     

一氧化氮供体对过氧化氢引起的心肌细胞损伤的保护作用

关于一氧化氮(NO)对心肌细胞是否具有保护作用目前尚存在争议,为探讨NO对过氧化氢(H2O2)引起的心肌细胞损伤是否具有保护作用及其可能的机制,实验将体外培养的新生大鼠心肌细胞分为3组(1)阴性对照组(Normal组);(2)H2O2组H2O2(0.1mmol/L)与心肌细胞共育4h;(3)S-亚硝基-N-乙酰青霉胺(SNAP)+H2O2组NO供体SNAP(0.5mmol/L)处理心肌细胞10min后,加入H2O2与心肌细胞共育4 h.用流式细胞术检测心肌细胞凋亡率,心肌细胞损伤程度以心肌细胞存活率和乳酸脱氢酶(lactate dehydrogenase,LDH)活性来表示,同时检测心肌细胞超氧化物歧化酶(superoxide dismutase,SOD)活性和丙二醛(MDA)含量.通过激光共聚焦显微术检测在不同处理条件下心肌细胞胞内钙的变化.结果表明,正常心肌细胞LDH活性和细胞存活率分别为631.4±75.6 U/L和93.1±6.2%,细胞凋亡率为0;H2O2处理细胞后可使细胞LDH活性显著增高(1580.5±186.7 U/L,P＜0.01),细胞存活率明显下降(58.3±7.6%,P＜0.01),流式细胞仪检测到大量心肌细胞凋亡,凋亡率为26.4±5.7%;SOD活性较正常细胞19.67±0.85 NU/ml显著下降,为14.73±1.68 NU/m(P＜0.01),MDA含量较正常细胞6.95±0.83μmol/L显著增高,为15.35±3.49μmol/L(P＜0.01).SNAP预处理细胞可显著提高心肌细胞存活率(79.7±9.3%,P＜0.01),降低LDH活性和细胞凋亡率(分别为957.8±110.9 U/L和9.1±3.3%,P＜0.01);并提高细胞抗氧化能力,表现为较H2O2处理组的SOD活性增高(21.36±3.11 NU/ml,P＜0.01),MDA含量下降(9.12±1.47 μmol/L,P＜0.01).激光共聚焦显微镜检测结果表明,H2O2可升高细胞内钙,而SNAP则可降低细胞内钙,SNAP预处理细胞后可取消H2O2升高细胞内钙的作用.上述结果提示,NO供体SNAP可对抗H2O2对心肌细胞的损伤,其机制与提高心肌细胞抗氧化损伤能力和对抗H2O2引起的细胞内钙超载有关.     

DMA增加正常大鼠心肌细胞钙瞬变和收缩

实验观察了钠氢交换或钠钙交换抑制剂 5 (N ,N 二甲基 )氨氯吡咪 (DMA)对正常和心肌肥厚大鼠分离心室肌细胞钙瞬变和细胞收缩的影响。通过负载荧光染料Fura 2 /Am ,应用离子影像分析系统 (IonImagingSystem)同步测定离体大鼠心肌细胞钙瞬变和细胞长度。结果表明 :DMA 10 μmol/L分别使钙瞬变和细胞缩短从对照组的 2 0 9.6 0± 5 4.96和 3.0 7± 0 .97μm增加到 2 38.5 0± 80 .41和 4.0 7± 1.0 2 μm (P <0 .0 5 ,n =7)。应用特异性反向钠钙交换阻断剂KB R7943可完全阻断DMA的激动作用。DMA还可使尼卡地平抑制L 型钙通道后的钙瞬变和细胞收缩增加。在肥厚心肌细胞 ,DMA表现出相同的药理作用 ,但对钙瞬变和细胞缩短的刺激作用更强。结果表明 :DMA可通过反向钠钙交换途径增加正常和肥厚大鼠心肌细胞钙瞬变和细胞收缩 ,且对肥厚心肌细胞的影响比对正常心肌细胞大。     

NO对兔窦房结自律性的影响

目的观察外源性NO供体硝普钠(SNP)和吗啉-斯德酮亚胺(SIN-1)对离体兔窦房结起搏细胞(SANC)自律性的影响,并初步探讨其作用机制.方法利用细胞内微电极技术记录SANC动作电位,分析APA(动作电位幅值),Vmax(0期最大除极速率),DD(舒张期除极速率),RPF(起搏细胞放电频率)的变化.结果SNP(10-5-10-2mol/L)增快SANC自发放电频率(RPF)和舒张期除极速率(VDD),并呈浓度依赖性.10-3 mol/LSNP使RPF(beats/min)由163±10.8增至195.0±13.1,DD(mV/s)由50.3±9.6增至70.2±12.1(P<0.01).SIN-1(10-3-10-2mol/L)亦使RPF和VDD增快(P<0.01).鸟苷酸环化酶抑制剂 10-4mol/L甲基美兰(MB)完全阻断了1 mmol/LSNP引起的RPF和VDD增长(P<0.01).If阻断剂2 mmol/L CsCl部分阻断了1 mmol/LSNP引起的正性变时作用和VDD增长(P<0.05).ICa-L阻断剂0.46 μmol/L NIF对于1 mmol/LSNP所致的RPF和VDD的增长无明显影响(P>0.05).结论外源性NO可增快离体兔SANC自律性,此效应通过NO-cGMP途径产生,至少部分与If增强有关,Ca-L对此无显著作用.     

白介素-2对心肌细胞[Ca~(2 )]_i的作用及其信号转导途径

为研究白介素 2 (interleukin 2 ,IL 2 )对心肌细胞内钙浓度 ([Ca2 ]i)的影响及其信号转导途径 ,实验采用酶解法分离成年大鼠心室肌细胞 ,以Fura 2 /AM为钙探针 ,用细胞内双波长钙荧光系统检测细胞 [Ca2 ]i 的变化。结果发现 :(1)IL 2 (0 5～ 2 0 0U/ml)浓度依赖性地降低单个心室肌细胞内钙瞬态 ,IL 2 (2 0 0U/ml)对咖啡因诱导的肌浆网内储钙的释放无影响 ;(2 )纳洛酮 (naloxone ,Nal) (10 -8mol/L)和nor binaltorphimine (nor BNI,10 -8mol/L)可阻断IL 2对心肌细胞钙瞬态的作用 ,而纳曲吲哚 (naltrindole ,NTI) (10 -6mol/L)不能阻断此作用 ;(3)κ阿片受体激动剂U5 0 488H (10 -6mol/L)降低心肌细胞钙瞬态 ,nor BNI (10 -8mol/L)可阻断此作用 ;(4 ) 5mg/L百日咳毒素 (PTX)预处理可取消IL 2降低心肌细胞钙瞬态的作用 ,而酪氨酸激酶抑制剂genistein (10 -4 mol/L)不能取消IL 2的作用 ;(5 )U7312 2预处理可阻断IL 2的作用。研究结果表明 ,IL 2降低心肌细胞钙瞬态的作用 ,是通过心肌细胞上κ阿片受体介导的 ,其下游途径包括PTX敏感的G蛋白和磷脂酶C。     

内皮素-1预处理对培养乳鼠心肌细胞低氧损伤的保护作用

实验观察了 0 0 1- 1nmol/L内皮素 1(ET 1)预处理对低氧孵育 ( 3 %O2 5 %CO2 ,12h)的培养乳鼠心肌细胞乳酸脱氢酶 (LDH)释放量、培养液上清超氧化物歧化酶 (SOD)活性以及丙二醛 (MDA)含量的影响。用Fluo 3 /AM负载培养的心肌细胞 ,在激光扫描共聚焦显微镜下监测急性低氧的心肌细胞 [Ca2 +]i 的变化和ET 1预处理对低氧所致 [Ca2 +]i 变化的影响。结果如下 :( 1)心肌细胞低氧孵育 12h后 ,培养液上清LDH活力和MDA含量较常氧对照组明显升高 ,分别为 43 3 3± 1 2 1U/Lvs 19 3 3± 1 0 3U/L和 1 71± 0 0 2nmol/Lvs 0 91± 0 0 3nmol/L (P<0 0 1) ,SOD活性为 16 93± 1 11U/ml明显低于常氧对照组的 3 3 48± 1 15U/ml (P <0 0 1) ;0 0 1- 1nmol/LET 1预处理呈浓度依赖性抑制低氧培养心肌细胞LDH释放 ,减少培养液上清MDA含量、提高SOD活性 (P <0 0 1)。 ( 2 )低氧灌流后 2 9± 1 5s (n =2 3 )心肌细胞自发性钙瞬变完全终止 ,[Ca2 +]i 升高了 10 7± 13 2 % (P <0 0 0 1) ;0 0 1- 1nmol/LET 1能明显加快心肌细胞钙瞬变的频率 (P <0 0 1) ;ET 1预处理后低氧所致钙瞬变终止的时间较单纯低氧组明显推迟 ,[Ca2 +]i过度升高被明显减轻 (P <0 0 1)。上述结果表明 ,0 0 1- 1nmol/LET 1预处理可减轻培     

胍丁胺对大鼠心室肌细胞内游离钙浓度的影响

本研究旨在观察胍丁胺 (agmatine ,Agm)对分离大鼠心室肌细胞内游离钙浓度 ( [Ca2 +]i)的影响。用酶解方法分离大鼠心室肌细胞 ,用Fluo 3 AM负载 ,然后用激光共聚焦法测定单个心室肌细胞 [Ca2 +]i 的荧光强度 (fluorescenceintensity ,FI) ,结果以FI或相对荧光强度 (F/F0 % )表示。实验结果表明 ,在正常台氏液 (含钙 1 0mmol/L)和无钙台氏液中 ,单个大鼠心室肌细胞的荧光密度分别为 12 8 8± 13 8和 119 6± 13 6,两者无差异。Agm 0 1、1和 10mmol/L浓度依赖性地显著降低细胞的钙浓度 ;在正常台氏液中加入EGTA 3mmol/L ,Agm同样降低细胞的钙浓度。KCl 60mmol/L ,PE 3 0 μmol/L ,和Bay K 864 410 μmol/L均升高心室肌细胞的[Ca2 +]i。Agm同样降低高浓度KCl、Bay K 864 4和PE诱发的心室肌细胞 [Ca2 +]i 升高。当细胞外液钙浓度由 1mmol/L增加到 10mmol/L时 ,诱发心室肌细胞钙超载 ,同时部分心室肌细胞产生可传播的钙波 (Ca2 +wave) ,Agm 1mmol/L降低钙波的传播速度和持续时间 ,最终阻断钙波。以上结果提示 ,Agm对心室肌细胞的胞浆[Ca2 +]i具有抑制作用 ,此作用通过阻断电压依赖性钙通道而实现 ;并可能与抑制大鼠心室肌细胞内钙释放有关     

白藜芦醇降低大鼠心室肌细胞内游离钙浓度

实验旨在研究白藜芦醇(resveratrol)对大鼠心室肌细胞内钙浓度(intracellular calcium concentratoin,[Ca2+]i)的影响.应用激光共聚焦显微镜技术记录心室肌细胞内的钙荧光强度.结果表明在正常台氏液和无钙台氏液中,白藜芦醇(15～60μmol/L)呈浓度依赖性地降低[Ca2+]i.蛋白酪氨酸磷酸酶抑制剂正钒酸钠(sodium orthovanadate,1.0 mmol/L)和L型Ca2+通道激动剂Bay K8644(10 μmol/L)可部分抑制正常台氏液中白藜芦醇的效应.但NO合酶阻断剂L-NAME(1.0 mmol/L)对白藜芦醇的作用无影响.白藜芦醇也能明显抑制无钙台氏液中由低浓度ryanodine(1.0 nmol/L)引起的[Ca2+]i增加.当细胞外液钙浓度由1 mmol/L增加到10 mmol/L而诱发心室肌细胞钙超载时,部分心室肌细胞产生可传播的钙波,白藜芦醇(60 μmol/L)可降低钙波的传播速度和持续时间,最终阻断钙波.结果提示,白藜芦醇能够降低心室肌细胞内游离钙浓度,此作用可能与其抑制电压依赖性Ca2+通道、酩氨酸激酶和肌浆网内钙释放有关.     

Effects of sprint training on contractility and [Ca(2+)](i) transients in adult rat myocytes.

The effects of 6-8 wk of high-intensity sprint training (HIST) on rat myocyte contractility and intracellular Ca(2+) concentration ([Ca(2+)](i)) transients were investigated. Compared with sedentary (Sed) myocytes, HIST induced a modest (5%) but significant (P < 0.0005) increase in cell length with no changes in cell width. In addition, the percentage of myosin heavy chain alpha-isoenzyme increased significantly (P < 0.02) from 0.566 +/- 0.077% in Sed rats to 0.871 +/- 0.006% in HIST rats. At all three (0.6, 1.8, and 5 mM) extracellular Ca(2+) concentrations ([Ca(2+)](o)) examined, maximal shortening amplitudes and maximal shortening velocities were significantly (P < 0.0001) lower and half-times of relaxation were significantly (P < 0.005) longer in HIST myocytes. HIST myocytes had significantly (P < 0.0001) higher diastolic [Ca(2+)](i) levels. Compared with Sed myocytes, systolic [Ca(2+)](i) levels in HIST myocytes were higher at 0.6 mM [Ca(2+)](o), similar at 1.8 mM [Ca(2+)](o), and lower at 5 mM [Ca(2+)](o). The amplitudes of [Ca(2+)](i) transients were significantly (P < 0.0001) lower in HIST myocytes. Half-times of [Ca(2+)](i) transient decline, an estimate of sarcoplasmic reticulum (SR) Ca(2+) uptake activity, were not different between Sed and HIST myocytes. Compared with Sed hearts, Western blots demonstrated a significant (P < 0.03) threefold decrease in Na(+)/Ca(2+) exchanger, but SR Ca(2+)-ATPase and calsequestrin protein levels were unchanged in HIST hearts. We conclude that HIST effected diminished myocyte contractile function and [Ca(2+)](i) transient amplitudes under the conditions studied. We speculate that downregulation of Na(+)/Ca(2+) exchanger may partly account for the decreased contractility in HIST myocytes.     

In situ SR function in postinfarction myocytes.

Previous studies have shown lower systolic intracellular Ca(2+) concentrations ([Ca(2+)](i)) and reduced sarcoplasmic reticulum (SR)-releasable Ca(2+) contents in myocytes isolated from rat hearts 3 wk after moderate myocardial infarction (MI). Ca(2+) entry via L-type Ca(2+) channels was normal, but that via reverse Na(+)/Ca(2+) exchange was depressed in 3-wk MI myocytes. To elucidate mechanisms of reduced SR Ca(2+) contents in MI myocytes, we measured SR Ca(2+) uptake and SR Ca(2+) leak in situ, i.e., in intact cardiac myocytes. For sham and MI myocytes, we first demonstrated that caffeine application to release SR Ca(2+) and inhibit SR Ca(2+) uptake resulted in a 10-fold prolongation of half-time (t(1/2)) of [Ca(2+)](i) transient decline compared with that measured during a normal twitch. These observations indicate that early decline of the [Ca(2+)](i) transient during a twitch in rat myocytes was primarily mediated by SR Ca(2+)-ATPase and that the t(1/2) of [Ca(2+)](i) decline is a measure of SR Ca(2+) uptake in situ. At 5.0 mM extracellular Ca(2+), systolic [Ca(2+)](i) was significantly (P 相似文献   

Overexpression of phospholemman alters contractility and [Ca(2+)](i) transients in adult rat myocytes

Previous studies showed increased phospholemman (PLM) mRNA after myocardial infarction (MI) in rats (Sehl PD, Tai JTN, Hillan KJ, Brown LA, Goddard A, Yang R, Jin H, and Lowe DG. Circulation 101: 1990-1999, 2000). We tested the hypothesis that, in normal adult rat cardiac myocytes, PLM overexpression alters contractile function and cytosolic Ca(2+) concentration ([Ca(2+)](i)) homeostasis in a manner similar to that observed in post-MI myocytes. Compared with myocytes infected by control adenovirus expressing green fluorescent protein (GFP) alone, Western blots indicated a 41% increase in PLM expression after 72 h (P < 0.001) but no changes in Na(+)/Ca(2+) exchanger, SERCA2, and calsequestrin levels in myocytes infected by adenovirus expressing GFP and PLM. At 5 mM extracellular [Ca(2+)] ([Ca(2+)](o)), maximal contraction amplitudes in PLM-overexpressed myocytes were 24% (P < 0.005) and [Ca(2+)](i) transient amplitudes were 18% (P < 0.05) lower than control myocytes. At 0.6 mM [Ca(2+)](o), however, contraction and [Ca(2+)](i) transient amplitudes were significantly (P < 0.05) higher in PLM-overexpressed than control myocytes (18% and 42%, respectively); at 1.8 mM [Ca(2+)](o), the differences in contraction and [Ca(2+)](i) transient amplitudes were narrowed. This pattern of contractile and [Ca(2+)](i) transient abnormalities in PLM-overexpressed myocytes mimics that observed in post-MI rat myocytes. We suggest that PLM overexpression observed in post-MI myocytes may partly account for contractile abnormalities by perturbing Ca(2+) fluxes during excitation-contraction.     

Sprint training normalizes Ca(2+) transients and SR function in postinfarction rat myocytes.

Previous studies have shown that myocytes isolated from sedentary (Sed) rat hearts 3 wk after myocardial infarction (MI) undergo hypertrophy, exhibit altered intracellular Ca(2+) concentration ([Ca(2+)](i)) dynamics and abnormal contraction, and impaired sarcoplasmic reticulum (SR) function manifested as prolonged half-time of [Ca(2+)](i) decline. Because exercise training elicits positive adaptations in cardiac contractile function and myocardial Ca(2+) regulation, the present study examined whether 6-8 wk of high-intensity sprint training (HIST) would restore [Ca(2+)](i) dynamics and SR function in MI myocytes toward normal. In MI rats, HIST ameliorated myocyte hypertrophy as indicated by significant (P 相似文献   

Calmodulin kinase II and protein kinase C mediate the effect of increased intracellular calcium to augment late sodium current in rabbit ventricular myocytes

An increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) augments late sodium current (I(Na.L)) in cardiomyocytes. This study tests the hypothesis that both Ca(2+)-calmodulin-dependent protein kinase II (CaMKII) and protein kinase C (PKC) mediate the effect of increased [Ca(2+)](i) to increase I(Na.L). Whole cell and open cell-attached patch clamp techniques were used to record I(Na.L) in rabbit ventricular myocytes dialyzed with solutions containing various concentrations of [Ca(2+)](i). Dialysis of cells with [Ca(2+)](i) from 0.1 to 0.3, 0.6, and 1.0 μM increased I(Na.L) in a concentration-dependent manner from 0.221 ± 0.038 to 0.554 ± 0.045 pA/pF (n = 10, P < 0.01) and was associated with an increase in mean Na(+) channel open probability and prolongation of channel mean open-time (n = 7, P < 0.01). In the presence of 0.6 μM [Ca(2+)](i), KN-93 (10 μM) and bisindolylmaleimide (BIM, 2 μM) decreased I(Na.L) by 45.2 and 54.8%, respectively. The effects of KN-93 and autocamtide-2-related inhibitory peptide II (2 μM) were not different. A combination of KN-93 and BIM completely reversed the increase in I(Na.L) as well as the Ca(2+)-induced changes in Na(+) channel mean open probability and mean open-time induced by 0.6 μM [Ca(2+)](i). Phorbol myristoyl acetate increased I(Na.L) in myocytes dialyzed with 0.1 μM [Ca(2+)](i); the effect was abolished by G?-6976. In summary, both CaMKII and PKC are involved in [Ca(2+)](i)-mediated augmentation of I(Na.L) in ventricular myocytes. Inhibition of CaMKII and/or PKC pathways may be a therapeutic target to reduce myocardial dysfunction and cardiac arrhythmias caused by calcium overload.