HCN4与生物心脏起搏的基础研究现状

HCN4作为超极化激活环核苷酸门控阳离子通道(hyperpolarization-activated cyclic nucleotide-gated cation channel,HCN)基因家族亚型,被认为是心脏起搏细胞产生自动节律重要基础,其中超极化激活电流(funny current,If)是重要的起搏调控部分,HCN家族基因参与编码If电流,由于HCN4与心脏起搏的产生和调节关系密切,所以被称为起搏基因.     

窦房结细胞起搏基因HCN4的研究进展

HCN即超级化激活的环核苷酸门控阳离子通道,其激活后产生的If/Ih离子流是窦房结起搏细胞动作电位正常形成的分子基础。随着对窦房结细胞起搏机制和HCN基因家族研究的不断深入,人们对HCN亚型HCN4的结构、分布、特性已有了较深入的了解。近年来有较多研究表明,人窦房结起搏基因HCN4突变与病态窦房结综合征密切相关。现就窦房结细胞起搏基因HCN4的特性及其与窦房结功能之间的关系作进一步研究和探讨。     

甲状旁腺素增加心率的机制研究

目的研究甲状旁腺素(PTH)增加心率的机制。方法运用微电极、膜片箝技术,记录PTH对家兔自律性动作电位和兔起搏离子流的影响。结果2×10-7mol/Lnifedipine使窦房结细胞动作电位的幅值减小及自律性减慢(P<0.05)。PTH20nmol/L增加窦房结动作电位的自律性及动作电位幅值的作用可被nifedipine减弱(P<0.05)。PTH10nmol/L使酶解游离窦房结细胞起搏离子流(If)明显增加。结论甲状旁腺素通过增强窦房结细胞起搏离子流If及钙内流来增加窦房结的自律活动。     

甲状旁腺素增加心率的机制研究

目的研究甲状旁腺素(PTH)增加心率的机制.方法运用微电极、膜片箝技术,记录PTH对家兔自律性动作电位和兔起搏离子流的影响.结果2×10-7 mol/L nifedipine使窦房结细胞动作电位的幅值减小及自律性减慢(P＜0.05).PTH 20 nmol/L增加窦房结动作电位的自律性及动作电位幅值的作用可被nifedipine减弱(P＜0.05).PTH 10 nmol/L使酶解游离窦房结细胞起搏离子流(If)明显增加.结论甲状旁腺素通过增强窦房结细胞起搏离子流If及钙内流来增加窦房结的自律活动.     

超极化激活环核苷酸门控阳离子通道在心血管病中的研究进展

心脏的自律性是由窦房结(sinuatrial node,SAN)的起搏细胞所控制的,这种自律活动的电生理基础是窦房结细胞的4期自动除极.在动作电位结束,细胞去极化可使膜电位逐渐增高接近新动作电位启动的阈电位,因而产生周期性的电活动.     

HCN4基因修饰大鼠MSCs体外诱导获得的心脏起搏样细胞动作电位检测

目的 探讨超极化激活及环化核苷酸调控的阳离子通道亚型4(hyperpolarization-activated cyclic nucleotidegated cation channel 4,HCN4)基因修饰大鼠MSCs体外诱导分化的心脏起搏样细胞的动作电位特性.方法 以HCN4基因修饰大鼠MSCs体外诱导获得的具有自发搏动的心脏起搏样细胞为实验组,同期培养的原代乳鼠窦房结细胞为对照组,采用全细胞膜片钳技术对两组细胞的动作电位进行检测.结果 心脏起搏样细胞及原代培养乳鼠窦房结细胞均可记录到具有舒张期自动去极化的动作电位,心脏起搏样细胞的动作电位特性与原代培养的乳鼠窦房结细胞相比,其静息电位、动作电位幅度及动作电位周期均无显著性差异(P＞0.05),但阈电位较原代培养的乳窦鼠房结细胞显著降低(P＜0.01).结论 从电生理角度证实,心脏起搏样细胞的特性与原代培养的乳鼠窦房结细胞相似.     

腺病毒介导超极化激活环核苷酸门控阳离子通道基因亚型4过度表达对乳鼠心肌细胞电生理特性的影响

目的观察过度表达超极化激活环核苷酸门控阳离子通道基因亚型(HCN)4对乳鼠心肌细胞电生理特性的影响。方法酶解法分离20只乳鼠的原代心肌细胞并进行培养,用含HCN4基因的重组腺病毒AdHCN4感染心肌细胞,以未感染细胞作为对照组,通过RT-PCR及细胞免疫荧光方法检测心肌细胞中HCN4基因在mRNA和蛋白质水平的表达,用全细胞膜片钳方法测定转染HCN4基因的心肌细胞的动作电位和起搏电流If。结果AdHCN4感染组心肌细胞中HCN4mRNA和蛋白质的表达水平明显高于对照组细胞;感染组心肌细胞自发搏动的频率为(92·5±7·4)次/分,明显高于对照组心肌细胞的(68·9±6·2)次/分(P<0·05);动作电位出现明显的4相自动除极化,其最大舒张电位(-52·8±4·2)mV明显高于对照组细胞(-62·1±2·6)mV(P<0·05);动作电位复极50%和90%的时间两者比较差异无统计学意义;在感染组细胞中,起搏电流If的电流密度为(115·7±7·8)pA/pF,也明显高于对照组细胞的(7·2±0·6)pA/pF(P<0·05)。结论过度表达起搏通道基因HCN4能够明显增强乳鼠心肌细胞的自律性,从而增加心肌细胞自发搏动的频率,这提示HCN4基因有可能成为治疗缓慢性心律失常疾病的目的基因。     

窦房结P细胞起搏的离子流机制研究进展

窦房结是心脏正常起搏点,其自动电节律的机制是4期自动除极。因为参与4期除极的内向和外向离子流众多,加之实验动物种属、标本采集、细胞分离方法、灌流液成分和实验设备等多种因素的影响,研究结果尚不完全一致。目前公认的观点是:窦房结P细胞4期自动除极是多种离子流综合作用的结果:①窦房结起搏细胞高膜电阻;②衰减性钾电流(IK);③背景内向钠电流(Ib-Na);④钠-钙交换电流(INa-Ca);⑤电压门控钙电流(ICa);⑥超极化激活的内向离子流(If)。因为起搏细胞高膜电阻,在众多离子共同作用下引起4期除极速度达60～70 mV/s,成为心脏主导起搏细胞。     

超极化激活环核苷酸门控阳离子通道（HCN）与心脏生物起搏的研究进展

心脏兴奋刺激来源于窦房结（sinoatrial node,SAN）,并经结间束传导至房室结（atrioventrieular node,AVN）,通过房室结纤维传导至心室浦氏纤维。心脏兴奋起源、传导等障碍是形成心律失常的基础。研究发现在心脏起搏中,有一种在超极化时激活,受环核苷酸调控的离子通道,一般称作超极化激活环核苷酸门控阳离子通道（hyperpolarization—activated and cyclic nucleotide—gate cation channel,HCN）。超极化激活的阳离子电流命名为If（funny current）或Ih（hyperpolarization current）。If的最重要作用在于使心脏起搏细胞产生自动节律,保证心脏的节律性和规则性跳动,是保证心脏正常的生理功能的前提。     

超极化激活环核苷酸门控阳离子通道的研究进展

1976年Noma首次报道在家兔心脏窦房结组织记录到一种超极化激活的内向电流,称为拼。随后在感光细胞和海马CA1区的锥体细胞也记录到这种超极化激活的内向电流,称为Iq。在神经系统,这种超极化激活电流称为Ih。1997年发现介导Ih的离子通道HCN,此后2年间陆续克隆出多种编吗HCN通道的基因。If表示出与超级化激活的阳离子电流的分子组成关联,在包括心脏和大脑起搏活性的生理功能上起着至关重要的作用,它决定了静息膜电位、树突样整合和突触传导。[第一段]     

The automaticity of Mahaim fibre and its response to effective ablation

Background Typical accessory pathways (APs) of Wolf-Parkinson-White syndrome have been widely discussed in recent decades. However, the characteristics of the special AP, Mahaim fibre,are not so clear. It is known that these fibres have antegrade conduction only, long conduction time,decremental node-like conduction and automaticity properties. This study was to elucidate the automaticity of Mahaim fibre and its response to effective ablation.Methods Thirteen patients with Mahaim fibre (ten atrioventricular and three atriofascicular accessory pathways) were subjected to electrophysiological study and radiofrequency ablation via catheter. The incidence and characteristics of anautomatic rhythm originating from Mahaim fibre were observed during the whole procedure, especially during radiofrequency current delivery.Results Repetitive and short-run automatic rhythm (rate: 65 -72 beats per minute), with a QRS morphology similar to that of clinical pre-excited atrioventricular re-entrant tachycardia (AVRT),occurred in two patients during sinus rhythm. Conduction via Mahaim fibre was successfully eliminated by radiofrequency current. Fourteen applications of RF were associated with irregularly accelerated automatic tachycardia of Mahaim fibre ( with a sensitivity of 78% ), lasting for 1.2 - 14 seconds.However, such automatic tachycardia of Mahaim fibre did not occur during 54 failed applications of radiofrequency current.Conclusions Mahaim fibre has the function of automaticity. The accelerated automatic tachycardia of Mahaim fibre occur red during radiofrequency catheter ablation can be used as a predictor for successful procedure.     

参仙升脉口服液对病态窦房结综合征小鼠ROS表达的影响和对HCN4离子通道的调控作用

探讨参仙升脉口服液对病态窦房结综合征（SSS）小鼠窦房结线粒体活性氧（ROS）释放的影响及其对环化核苷酸调控阳离子通道蛋白亚型4（HCN4）离子通道的调控作用,探讨参仙升脉口服液改善SSS的作用机制。     

动态心电图结合窦房结功能检查诊断病窦综合征的价值

报告15例病态窦房结综合征的动态心电图监测与窦房结功能检查（经食道心房调搏）的比较，发现：动态心电图比较容易发现窦性停搏、阵发性室上性快速型心律失常，可以补充窦房结功能检查的不足；动态心电图监测最易发现植物神经对窦房结自律性的影响。并对上述发现的发生机理进行了讨论。     

微波辐射对大鼠窦房结组织HCN4基因表达的影响

目的观察微波辐射对受损大鼠窦房结(SAN)组织超极化激活环核苷酸门控阳离子通道4(HCN4)基因表达的影响,探讨微波辐射致窦房结损伤的可能机制。方法将60只Wistar雄性大鼠随机分为假辐射组和50 mW·cm-2组,每组30只,建立微波辐射致窦房结损伤大鼠模型,于辐射后1、7、14、28 d及3、6个月,采用原位杂交法检测窦房结HCN4 mRNA表达变化。结果与假辐射组比较,50 mW·cm-2组大鼠于辐射后1、7、14、28 d,窦房结细胞胞质内HCN4 mRNA表达显著增加;而于辐射后3、6个月显著减少,其差异均有统计学意义(P<0.05或P<0.01)。结论HCN4异常表达是微波辐射致SAN损伤的重要致伤分子之一。     

EFFECTS OF HIGH DOSE NARCOTICS AND CONTROLLED HYPOTENSORS ON THE HEART CONDUCTION SYSTEM

The effects of narcotics on the heart conduction system were studied in 51 rabbits. It was found that at equal efficacy dosages, the effects of high dose morphine, pethidine and fentanyl on the heart con duction system differ. Morphine 3 mg/kg injected iv did not influence the heart conduction system; fentanyl 30 ug/kg might improve both sinus node automaticity and sino-atrial conduction and pethidine 30 Vg/kg might seriously harm the heart conduction system. So fentanyl may be an ideal narcotic in patients with sick sinus syndrome. But when the dosage of fentanyl was increased t0 75 yg/kg, the sinus node was inhibited. The degree of pethidine effect upon the heart conduction system depends upon the dosage and rate of injection. The effects of controlled hypotensors on the heart conduction system were studied in 48 dogs. Trimethaphan (TMTP), sodium nitroprusside (SNP) nitro-glycerine (NG) and phentolamine (PA) were dripped iv respectively to lower the blood pressure by 30-40%. Five and 60 minutes after TMTP ad- ministration the atrio-ventricular junction was in inhibited and at 60 minutes the sino-atrial conduction was also inhibited. NG had no influence on the heart conduction system but SNP might inhibit the sinus node automaticity PA may also inhibit the sino atrial conduction, so this agent contraindicated in patients with sick sinus syndrome or sino-atrial block.     

Overexpression of connexin 45 in rat mesenchymal stem cells improves the function as cardiac biological pacemakers

Background Extensive research toward creating a biological pacemaker by enhancement of inward depolarizing current has been performed. However, studies have mainly focused on inducing spontaneous activity and have not adequately addressed ways to improve pacemaker function. In this study we attempted to improve pacemaker function by altering connexin expression in rat mesenchymal stem cells （MSCs） to a phenotype similar to native sinus node pacemaker cells. Methods To generate a biological pacemaker, MSCs were transduced with a cardiac pacemaker gene- hyperpolarization-activated cyclic nucleotide-gated channel 4 （HCN4）, via transfection with a lentiviral vector. Funny current （If） in HCN4~ MSCs was recorded by voltage-clamp. Overexpression of connexin 45 （gene Gja7） in MSCs was achieved by transfection with the plasmid pDsRED2-N1-Gja7-RFP. Double-immunolabelling with anti-connexin 43 and anti-connexin 45 antibodies were used to identify the gap junction channels. The effects of the genetically modified MSCs on cardiomyocyte excitability were determined in MSCs cocultured with neonatal rat ventricular myocytes. Spontaneous action potentials of neonatal rat ventricular myocytes were recorded by current-clamp. Results High level time- and voltage-dependent inward hyperpolarization current that was sensitive to 4 mmol/L Cs＋ was detected in HCN4＋ MSCs, confirming that HCN4 acted as Ir channels in MSCs. Connexin 43 and connexin 45 were simultaneously detected in CX45＋ MSCs. Beating frequency was （82±8） beats per minute （n=-5） in myocytes cocultured with non-transfected control MSCs, versus （129±11） beats per minute （n=-5） in myocytes cocultured with HCN4＋ MSCs. Myocytes cocultured with MSCs cotransfected with HCN4 and connexin 45 had the highest beating frequency at （147±9） beats per minute （n=5）. Conclusion These findings demonstrate that overexpression of connexin 45 and subsequent formation of heteromeric connexin 45/connexin 43 gap junction channels in HCN4 expressing MSCs can improve their function as cardiac biological pacemakers in vitro.     

莲心碱对正常大鼠的降压作用和对心电学的影响

本文观察了不同剂量的莲心碱对正常大鼠血压和心电学的影响。结果表明２￣１２ｍｇ／ｋｇ莲心碱静脉注射可产生不同程度的降压作用，其降压效应和有效作用时间呈剂量依赖性。莲心碱降低舒张压的作用大于收缩压，提示该药的主要作用部位在外周血管。此外，莲心碱还可延长ＰＲ、ＱＲＳ和ＱＴｃ间期，提示该药对房室结和心室肌的传导有抑制作用，并可延缓心室复极过程，表明莲心碱具有Ｉａ类抗心律失常药物的电生理特征。大剂量（１２ｍ     

内皮素-1通过一种不依赖于p38MAPK的信号转导机制刺激心肌细胞起搏通道If的表达

摘要：目的探讨血管活性肽内皮素-1（ET-1）对新生鼠心室肌细胞起搏通道If表达的影响及其细胞内信号机制。方法采用酶消
化法急性分离1~3 d龄新生大鼠心室肌细胞,用全细胞膜片钳技术记录If电流,定量RT-PCR技术检测介导If电流的、超极化激活
的环核苷酸门控通道亚型HCN2 和HCN4 的表达。结果ET-1 呈剂量和时间依赖性增强了HCN2 和HCN4 通道亚型的表达,
ET-1的这一作用可被其ETA受体阻断剂BQ-123阻断,而非ETB受体阻断剂BQ-788,而且特异性p38 MAPK抑制剂SB-203580
也不影响ET-1的作用。结论ET-1通过一种ETA受体介导的、不依赖于p38 MAPK的信号转导机制刺激了心肌细胞起搏通道If
的表达。这一作用与ET-1致心律失常机制有关。
     

克班宁对豚鼠心肌生理特性的影响

目的:研究克班宁(Cre)对豚鼠离体心房肌生理特性的影响。方法:常规离体器官实验法。结果:Cre能明显减少豚鼠右心房自律性活动,明显抑制肾上腺素诱发豚鼠左心房自律性和兴奋性,豚鼠的左心房肌的功能,不应期也能明显延长,但对收缩性无明显影响。结论:Cre的抗心律失常作用可能与其降低自律性和兴奋性有关。