The Study On Repolarization Potassium Current Of Ventricular Myocardium In Aged Rat

Age is a important factor of influencing cardiovascular function. As aging, electrophysiological remodeling occurs accompanying with anatomy remodeling, which lead to morphologic change and dysfunction in the aged cardiovascular system. Phynotypes of histomorphological show heart weight gain, progressing myocardium fibrosis, increasing intercellular collagen contents, thickening left ventricular wall, diminishing left ventricular cavum, hypertrophia ventricular myocytes,greater membrane capacitance. In clinical practice, the morbidity of myocardial ischemia, myocardial hypertrophy, heart failure, arrhythmia and SCD based on organic heart disease in the older are prevailing. The characteristic of cardiovascular disease in aged population differs from that in any other age group. In addition, the incidence of SCD is high as well in the aged group without identified heart disease evidence. The potential electrophysiologic mechanism, especially,the relationship with heart physiological senescence is still indefinite. Conclusions of associated electrophysiologic studies vary. The eletro-remodeling basis on anatomy remodeling induces electrophysiological characteristics changes in aged ventricular myocyte, which is the electrophysiologic mechanism of repolarization heterogeneity augmentation, arrhythmia based on trigger or reentry and myocardium dysfunction. The main changes in ventricular myocyte as aging display the APD and the amplitude, currentdensity, channel kinetics, gating mechanism of several important currents influencing ventricular repolarization dispertion in repolarization phase, which can lead to ventricular electro-steady alteration, thus,malignant arrhythmia and cardiac function failure will emerge. This study aim to bring new systematic and simple separation method to apply to the isolating aged ventricular myocyte by Langendorff perfusion and enzymolysis in rat, to study the repolarization possiumcurrents I_to, I_K1, I_K in different age groups in rats by patch-clamp, so to observe the electrophysiologyical characteristics change in ventricular myocyte in older rats and approach the possible electrophysiologic mechanism of arrhythmia in aged people. This study will offer one of the electrophysiologic mechanisms to treat geriatric cardiovascular disease and to develop a new selected channel blocker as an anti arrhythmic in aged population. This study include three parts.1. The cardiac index and the separation method to ventricular myocyte in a ged ratObjective: To bring a systematic and simple enzymolysis separation method to apply to aged ventricular myocyte in rat. To observe the cardiac index , procedure characteristic and influence factor when separating ventricular myocytes, cell morphology and membrane capacitance in aged rat myocardium. Methods: To select healthy aged rats(22～24 Mo.) and adult(13～15 Mo.), young groups(3～5 Mo.) as control. A isolated heart was flybackly perfused through aorta by Langendorff. The single ventricular myocyte was obtained by two steps enzymatic dissociation method in vitro in 3 different age groups rats. Results: Compared with the young group, ventricular myocytes dissociation needed a more collagenase quantity and longer perfusion time, but the rate of living cells was lower and cellular survival time was shorter in aged rat. Body weight, heart weight and left ventricular weight of aged rat all gained(P<0.01), the length and width were bigger(P<0.01), left ventricular cycle length decreased(P< 0.01), left ventricular wall thickened(P<0.05), myocytes hypertrophied(P<0.01), the ratio of length/width was smaller, cell membrane capacitance was bigger(P<0.01) compared with other age groups rats. Conclusion: By this method acquired myocytes possess good electrophysiological activity. The cardiac index and myocyte electrophysiological characteristics in aged rat differ from those in other age groups.2. The potassium current characteristics of aged ventricular myocyte There is three chapters in this part.Chapter one The characteristic of transient out potassium current (I_to) in myocardium myocyte of aged ratObjective: To observe whether the amplitude , density and channel gating kinetics oftransient out potassium current (I_to) in older rat differ from those in other age group rat.Methods: To separate the rat ventricular myocytes by Langendorff perfusion and enzymatic dissociation method in vitro, and whole cell patch-clamp technique was usedto record I_to in ventricular myocytes of 3 different age groups rats (3～5Mo., 13～15Mo., 22²4Mo.). Results: I_to of aged rat showed voltage-dependency and the currentamplitude of I_to was bigger, including peak current increas as well the steady-state current in ventricular myocytes of aged rat. In aged rat, peak current density changed insignificantly, but the steady-state one increased significantly, so the ratio of steady-state current/peak current increase. The aged rat showed a frequencyuse-dependency at 2.0 Hz. I_to display a smallest use-dependency in peak current, but noone in steady-state curent. The I_to current fast active, inactive slowly and significantly at high depolarization potential and recovery fast. Conclusion: The evidentaging-associated changes of I_to may be one of the important electrophysiologic mechanism of arrhythmia in aged people.Chapter two The characteristic of the inward rectifier potassium current (I_K1 ) in myocardium myocyte of aged ratObjective: To observe whether the inward rectifier potassium current (I_K1) in myocardium myocyte of aged rat display an aged-associated change. Methods: To separate the rat ventricular myocytes by Langendorff perfusion and enzymatic dissociationmethod in vitro, and whole cell patch-clamp technique was used to record I_K1 in ventricular myocytes of 3 different age groups rats (3～5Mo., 13～15Mo., 22～24Mo.).Results: The density and I-V curve of I_K1 didn't differ between 3 groups rats.Conclusion: There's no aging-associated changes of current density and voltage-dependency of I_K1 in aged rat.Chapter three The characteristic of the delayed rectifier potassium currentin myocardium myocyte of aged ratObjective: To observe whether the delayed rectifier potassium current (I_K) in myocardium myocyte of aged rat exist an aging-associated change. Methods: To obtain the rat ventricular myocytes by Langendorff perfusion and enzymatic dissociation method in vitro, and whole cell patch-clamp technique was used to record I_K in ventricular myocytes of 3 different age groups rats (3～5Mo., 13～15Mo., 22～24Mo.). Results: The density of I_ks.tail was bigger and a significant voltage-dependency in aged rat. The density and voltage-dependency of both time dependence current of I_Ks and I_kr had no age-associated changes in aged rat. The activation of I_Ks changed insignificantly, but slightly slow inactived. The I_Ks might contribute more in aged rat than other aged groups.Conclusion: The aging-associated changes of I_K in aged rat may be one of electrophysiological mechanism of arrhythmia and susceptibility to arrhythmogenic effect from antiarrhythmic drugs in the old.3. The change of I_to caused by 4-AP and ISO in ventricular myocyte of aged ratObjective: To observe the change of I_to caused by 4-AP and ISO in ventricular myocyte in aged rat. Methods: To separate the rat ventricular myocytes by Langendorff perfusion and enzymatic dissociation method in vitro, and whole cell patch-clamptechnique was used to respectively record I_to in ventricular myocytes of 2 different age groups rats (3～5Mo., 22～24Mo.). A accumulated dose regimen of 4-AP and ISO wasrespectively administered to every ventricular myocyte. The I_to was recorded before and after administration administration. Results: Compared with the young group, 4-APdecreased significantly the density of I_to, showing a concentration dependent, no change on voltage-dependency of I_to and I_to inhibition by 4-AP was partly reversible in aged group. 4-AP made I_to steady-state activation slower and inactivation faster in aged rat, but significantly prolonged the recovery after inactivation of I_to in young rat. ISOincreased significantly the density of I_to and no change on the voltage-dependency of I_toin aged rat. By ISO titration, the activation of I_to was faster and inactivation was slower in young group than the aged one. ISO didn't significantly affect steady-stateinactivaton of I_to in aged rat. Conclusion: The different reaction to administration bydrugs of I_to shows an aging-associated changes in aged group.