RESUMEN
AIMS: Various pathological conditions can alter cardiac electrophysiological properties not only by physiological responses but also by modifying the gene expression of ion channels (electrical remodelling). To investigate the underlying mechanisms of the latter, electrophysiological alterations would require a simultaneous and comprehensive analysis of the mRNA level of the ion channel genes. METHODS AND RESULTS: We designed 19 cardiac ion channel cDNA templates to analyse the corresponding mRNAs and classified them into three template sets. Those sets were a voltage-dependent K(+) channel series (rat erg, KvLQT1, Kv4.3, Kv4.2, Kv2.1, Kv1.5, Kv1.4, Kv1.2), an inwardly rectifying K(+) channel series (rat Kir6.2, SUR2A/B, Kir3.4, Kir3.1, Kir2.2, Kir2.1), and an inward cationic ion channel series (rat SCN5A, alpha1C, beta2, alpha2delta2 of cardiac L-type Ca(2+) channel and alpha1G). These cDNA templates were used to synthesize antisense digoxigenin-labelled RNA probes. An amount of the total RNA of 25 microg was adequate to analyse simultaneously the mRNA levels of the ion channel genes with the use of multi-probe RPA, and these three multi-probe template sets enabled us to evaluate the profile of the spatial and temporal transcripts of the cardiac ion channels. CONCLUSION: The newly developed ion channel multi-probe RPA templates provide an aid in the comprehensive analysis of the electrical remodelling of the heart.
Asunto(s)
Perfilación de la Expresión Génica/métodos , Canales Iónicos/genética , Canales Iónicos/metabolismo , Miocardio/metabolismo , ARN Mensajero/genética , Animales , Bioensayo/métodos , Técnicas In Vitro , Canales Iónicos/análisis , Técnicas de Sonda Molecular , ARN Mensajero/análisis , Ratas , Ratas Sprague-Dawley , RibonucleasasRESUMEN
BACKGROUND: Many pathological conditions induce electrical remodeling, possibly through intracellular Ca2+ overload, but the currently available L-type Ca2+ channel blockers may be detrimental because of their global negative inotropic effects. METHODS AND RESULTS: To determine whether the L-type Ca2+ channel is identical throughout the heart, the distribution of the mRNAs and proteins comprising the L-type Ca2+ channel and its electrophysiological properties were analyzed in rat atria and ventricles. The mRNA of alpha2delta-2 (Cacna2d2) was more abundantly expressed in the atrium (approximately 5-fold) than in the ventricle. In contrast, alpha1C (Cacna1c) (Cav1.2) mRNA was significantly less abundant in the atrium. The level of the alpha1C (Cacna1c) (Cav1.2) protein was decreased (approximately 0.5-fold) and that of alpha2 delta-1 (Cacna2d1) was increased (approximately 2-fold) in the atrium compared with the ventricle. Although the peak ICa,L density showed no significant differences, voltage dependence of inactivation and activation of the current showed a more depolarized shift in the atrium than in the ventricle. CONCLUSION: These results indicate that in the rat heart the L-type Ca2+ channel differs between the atrium and ventricle with regard to gene expression and electrophysiological properties.
Asunto(s)
Función Atrial , Canales de Calcio Tipo L/análisis , Atrios Cardíacos/química , Ventrículos Cardíacos/química , Función Ventricular , Animales , Canales de Calcio/análisis , Canales de Calcio/genética , Canales de Calcio Tipo L/genética , Electrofisiología , Femenino , Regulación de la Expresión Génica/fisiología , Técnicas de Placa-Clamp , ARN Mensajero/análisis , Ratas , Ratas Sprague-DawleyRESUMEN
Antiarrhythmic drugs exert their effects by inhibiting the ion channels of cardiomyocytes. However, these effects could also modify the ionic environment around them, and thereby affect the expression of ion channels, leading to biochemical enhancement or attenuation of the antiarrhythmic effects. To test this hypothesis, the physiological and biochemical effects of cibenzoline were evaluated in a rapid atrial pacing model in rats. In rats with rapid atrial pacing, pretreatment with cibenzoline significantly inhibited the increases in Kv1.5 mRNA at 2 hours and immunoreactive protein at 4 hours by 35 +/- 15% and 30 +/- 10%, respectively. These effects were observed only in the rapid atrial pacing group, not in the sham-operated group. With cibenzoline pretreatment, 4-hour rapid atrial pacing resulted in significant prolongation of the atrial refractory period compared to the untreated group even after removal of cibenzoline. In contrast, the sham and rapid atrial pacing model with and without cibenzoline pretreatment showed similar acute physiological responses to cibenzoline. In conclusion, in addition to the acute physiological effects, pretreatment with cibenzoline exerted pleiotropic effects of inhibition of Kv1.5 channel upregulation by rapid pacing, implying differences in the cibenzoline effects when administered before and after onset of paroxysmal atrial fibrillation.
Asunto(s)
Antiarrítmicos/farmacología , Fibrilación Atrial/fisiopatología , Imidazoles/farmacología , Canales de Potasio con Entrada de Voltaje/genética , Animales , Fibrilación Atrial/genética , Fibrilación Atrial/metabolismo , Electrofisiología , Atrios Cardíacos/fisiopatología , Sistema de Conducción Cardíaco/fisiopatología , Canal de Potasio Kv1.5 , Marcapaso Artificial , Canales de Potasio con Entrada de Voltaje/biosíntesis , ARN Mensajero/biosíntesis , ARN Mensajero/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Periodo Refractario Electrofisiológico , Regulación hacia Arriba/efectos de los fármacosRESUMEN
BACKGROUND: Atrial fibrillation (AF) is well known as one of the cardiogenic causes for thromboembolism. Although decreased flow and hypercoagulable state of the blood in the fibrillating atrium have been emphasized as the underlying mechanisms, endocardial dysfunction in maintaining the local coagulation balance could also contribute to the thrombogenesis in AF. METHODS AND RESULTS: The paroxysmal AF model was created by rapid atrial pacing in anesthetized rats. To test the hypothesis that AF induces local coagulation imbalance by disturbing the atrial endocardial function, the gene expression of intrinsic anticoagulant factors, thrombomodulin (TM) and tissue factor pathway inhibitor (TFPI), were determined by means of ribonuclease protection assay, Western blotting, and immunohistochemistry. Rapid atrial pacing for 8 hours significantly decreased TM and TFPI mRNA levels in the left atrium but not in the ventricle, leading to the downregulation of their immunoreactive proteins. Immunohistochemical analysis revealed that TM and TFPI were expressed predominantly in the endocardial cells of the normal atrium, presumably preventing local blood coagulation, and that rapid atrial pacing induced the loss of TM and TFPI expression in the endocardium, leading to deficiency in anticoagulant barriers between the atria and the blood. CONCLUSIONS: Rapid atrial pacing acutely downregulated the gene expression of TM and TFPI in the atrial endocardium, thereby inducing local coagulation imbalance on the internal surface of the atrial cavity. These results would support the validity of supplement of anticoagulant molecules deficient in AF.
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Fibrilación Atrial/fisiopatología , Estimulación Cardíaca Artificial , Endocardio/metabolismo , Lipoproteínas/metabolismo , Trombomodulina/metabolismo , Animales , Fibrilación Atrial/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo , Atrios Cardíacos/metabolismo , Atrios Cardíacos/fisiopatología , Ratas , Ratas Sprague-Dawley , TaquicardiaRESUMEN
BACKGROUND: Many cardiac arrhythmias have their own characteristic circadian variations. Because the expression of many genes, including clock genes, is regulated variably during a day, circadian variations of ion channel gene expression, if any, could contribute to the fluctuating alterations of cardiac electrophysiological characteristics and subsequent arrhythmogenesis. METHODS AND RESULTS: To examine whether cardiac K+ channel gene expression shows a circadian rhythm, we analyzed the mRNA levels of 8 Kv and 6 Kir channels in rat hearts every 3 hours throughout 1 day. Among these channels, Kv1.5 and Kv4.2 genes showed significant circadian variations in their transcripts: approximately 2-fold increase of Kv1.5 mRNA from trough at Zeitgeber time (ZT) 6 to peak at ZT18 and a completely reverse pattern in Kv4.2 mRNA ( approximately 2-fold increase from trough at ZT18 to peak at ZT6). Actually, along with the variations in the immunoreactive proteins, the density of the transient outward and steady-state currents in isolated myocytes and the responses of atrial and ventricular refractoriness to 4-aminopyridine in isolated-perfused hearts showed differences between ZT6 and ZT18, a circadian pattern comparable to that of Kv1.5 and Kv4.2 gene expression. Reversal of light stimulation almost inverted these circadian rhythms, although pharmacological autonomic blockade only partially attenuated the rhythm of Kv1.5 but not of Kv4.2 transcripts. CONCLUSIONS: Among all the cardiac K+ channels, Kv1.5 and 4.2 channels are unique in showing characteristic circadian patterns in their gene expression, with Kv1.5 increase during the dark period partially dependent on beta-adrenergic activities and Kv4.2 increase during the light period independent of the autonomic nervous function.
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Ritmo Circadiano , Regulación de la Expresión Génica , Miocardio/metabolismo , Canales de Potasio con Entrada de Voltaje , Canales de Potasio/genética , Antagonistas Adrenérgicos beta/farmacología , Animales , Células Cultivadas , Conductividad Eléctrica , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/efectos de la radiación , Corazón/fisiología , Canal de Potasio Kv1.5 , Luz , Técnicas de Cultivo de Órganos , Técnicas de Placa-Clamp , Canales de Potasio/biosíntesis , Propranolol/farmacología , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Canales de Potasio ShalRESUMEN
In a pilot study, we determined the target ventricular rate of patients with atrial fibrillation by evaluating their ventriculoarterial coupling. Eleven patients with atrial fibrillation were studied. We recorded M-mode echocardiograms and radial artery blood pressure simultaneously. The left ventricular end-systolic pressure-volume ratio (Emax) and effective arterial elastance (Ea) were calculated for each beat, and the relationship of the preceding R-R interval (pRR) to Emax and Ea was evaluated. There was a significant positive correlation between pRR and Emax, and a significant negative correlation between pRR and Ea in all patients. The pRR that produced maximal stroke work was determined at the point of Emax=Ea, and the pRR that achieved maximal mechanical efficiency was determined at the point of 2Ea=Emax. By evaluating ventriculoarterial coupling in these patients who had atrial fibrillation, we were able to determine that the range between the 2 pRR intervals was the range of the optimal ventricular rate. A narrower range of the 2 pRR intervals was observed in patients with dilated cardiomyopathy than in the patients with no underlying cardiac disease. We conclude that it may be possible to determine the optimal ventricular rate in patients with atrial fibrillation by evaluating ventriculoarterial coupling.