a Laboratoire de Pharmacologie, INSERM E 00.01, Faculté de Médecine Paris Sud, Paris, France
b Fédération de Cardiologie, Hôpital Henri Mondor, Créteil, France
* Correspondence to: Pr. Alain Berdeaux, Laboratoire de Pharmacologie, INSERM E 00.01, Faculté de Médecine Paris-Sud, 63, rue Gabriel Péri, 94270 Le Kremlin-Bicêtre, France. Tel.: +33-1-495967021; fax: +33-1-49600031
E-mail address: alain.berdeaux{at}kb.u-psud.fr
Received 20 March 2003; revised 18 September 2003; accepted 16 October 2003 See page 537 for the editorial comment on this article2
Abstract
Aims The respective contributions of reduction in heart rate and inotropism in the beneficial effects of beta-blockade in ischaemic heart disease remains debated. The effects of selective heart rate reduction by ivabradine (Ifinhibitor) were compared to those of atenolol on exercise-induced ischaemia and stunning.
Methods and results In seven instrumented dogs, coronary stenosis was calibrated to suppress increase in coronary blood flow during a 10-min treadmill exercise. When administered before exercise, atenolol and ivabradine similarly reduced heart rate versus saline at rest and during exercise (154±2 and 155±9 vs 217±13beats/min, respectively). During exercise, left ventricular wall thickening (LVWth) was reduced to 2±1% from 23±4% under saline but ivabradine limited this effect (10±3%) and reduced the subsequent myocardial stunning vs saline. Atenolol also limited LVWth decrease during exercise (17±4%) but had no effect during recovery. When administered after exercise, ivabradine attenuated stunning and this effect disappeared when heart rate reduction was corrected by atrial pacing. Atenolol administered after exercise severely depressed LVWth vs saline.
Conclusion Selective heart rate reduction not only provides an anti-ischaemic effect but also per se improves contractility of the stunned myocardium. Additional negative inotropism is protective against ischaemia but deleterious during stunning.
Key Words: Chronotropic agent Heart rate Ischaemia Myocardial stunning Ventricular function
Introduction
Heart rate is a major and independent predictor of cardiovascular morbidity and mortality in ischaemic heart disease.1 Among drugs which reduce heart rate, ß-blockers also decrease myocardial contractility and oxygen consumption, and improve diastolic perfusion time.2,3 However their negative inotropic effect might be deleterious when left ventricular dysfunction occurs, e.g. during myocardial stunning,4 although conflicting results have been reported on this issue.5 Furthermore, the anti-ischaemic benefit of negative inotropism is not well established.6,7 In this setting, agents which selectively reduce heart rate have been developed as an alternative approach. They provide a potent anti-ischaemic effect and strongly protect the myocardium against stunning in ischaemic exercising dogs.8,9
Accordingly, the aim of this study was to compare on the ischaemic and stunned myocardium the effects of the widely used ß-blocker, atenolol, to those of ivabradine, a novel selective inhibitor of the cardiac pacemaker hyperpolarization-activated If channel in the cardiac sino-atrial node.1012 By increasing the duration of spontaneous depolarization, it induces a selective heart rate reduction10 and its administration is devoid of any effect on myocardial contractility and coronary vasomotion.13 For this purpose, we used an experimental model in which ischaemia resulted from the combination of a treadmill exercise and a partial coronary artery stenosis in conscious dogs.9,14 The effects of ivabradine and atenolol were investigated when their administration was started either before exercise-induced ischaemia or during the recovery period, i.e., during myocardial stunning. To specifically investigate the contribution of heart rate reduction, the experiments were performed both at spontaneous heart rate and under atrial pacing.
Methods
The animal instrumentation and the ensuing experiments were conducted in accordance with the recommendations of the French Ministry of Agriculture.
Surgical preparation
As previously described,9 a left thoracotomy was performed in seven dogs (2229kg). Filled fluid catheters were implanted in the descending thoracic aorta and the left atrium for measurement of blood pressure and microspheres injections, respectively. A Silastic catheter was introduced into the pulmonary artery for drug administration. A solid-state micromanometer (Konigsberg Instruments, Pasadena CA, USA) was introduced into the left ventricle (LV). A Transonic flow probe and a pneumatic occluder were implanted on the circumflex coronary artery. Two pairs of ultrasonic crystals were placed within the distribution of the circumflex coronary artery (ischaemic zone) and of the left anterior descending coronary artery (non-ischaemic zone) for LV wall thickening measurement. Electrodes were fixed on the left atrial appendage for pacing. All catheters and wires were exteriorized between the scapulae. Cefazolin (1g, i.v.) and gentamycin (40mg, i.v.) were administered before and during the first week after surgery. Post-operative analgesia was provided with morphine.
Haemodynamic measurements
Data were recorded and analysed using the data acquisition software Notocord-HEM 3.3 (Notocord System, Croissy-sur-Seine, France). Aortic and left atrial pressures were measured with a Statham P23ID strain gauge transducer (Gould-Nicolet, Courtaboeuf, France). Because it was measured by a hydraulic technique, aortic pressure could not be accurately recorded during exercise. LV pressure was measured using the Konigsberg gauge and LV dP/dt was computed from the LV pressure signal. Circumflex coronary artery blood flow was measured with a transit-time flowmeter (Transonic T206, Transonic Systems, Ithaca, NY, USA). Diastolic perfusion time was measured from negative LV dP/dt to theinitiation of the upstroke of LV pressure tracing.
Measurements of regional contractility
According to the method used in our laboratory,15 wall thicknesses were obtained using sonomicrometry, i.e., with an ultrasonic transit-time dimension gauge. The sonomicrometer (Module 201, System 6, Triton Technology Inc., San Diego, CA, USA) measures the distance between the endocardial and epicardial crystals by measuring the transit time of ultrasound between this pair of crystals and converting this time to an equivalent distance, the ultrasonic impulses travelling at the velocity of 1.58µm/s. End-diastolic wall thickness was measured at the initiation of the upstroke of LV pressure tracing, and the end-systolic wall thickness was measured 20ms before negative LV dP/dt. Percent wall thickening was defined as end-systolic minus end-diastolic thicknesses times 100 and divided by end-diastolic thickness.
Measurements of regional myocardial blood flows
As previously reported,14 regional myocardial blood flows (RMBFs) were measured using the fluorescent microspheres technique. Microspheres labelled with fluorescent dyes (FluoSpheres, Triton System, San Diego, CA, USA) were injected via the left atrial catheter. Arterial blood reference samples were withdrawn (7.5ml/min during 120s). At termination of the study, the heart was excised and the left ventricle was cut into 34 slices and further divided into endocardium, mid-myocardium and epicardium in the non-ischaemic and ischaemic zones. Samples were then processed and blood flows (expressed as ml/min/g of myocardium) were calculated.
Experimental protocol
Three weeks after surgery, dogs were installed on a treadmill and baseline parameters were recorded (Base 1). A second set of measurements (Base 2) was initiated 20min later. A partial stenosis of the left circumflex coronary artery was then performed using the pneumatic occluder without altering LV posterior wall thickening at rest. A treadmill exercise (10min duration, 10km/h, 13% slope) was then started. The stenosis was maintained during exercise in order to keep mean coronary blood flow at its corresponding Base 2 value. The occluder was deflated at the end of exercise. All parameters were continuously recorded at baseline, during exercise and at selected intervals during the first 6h of the recovery period. Regional myocardial blood flows were measured between the 6th and the 8th min of exercise.
In the first part of the protocol (sequence A), saline, atenolol or ivabradine (Laboratoires Servier, Neuilly-sur-Seine, France) were administered immediately after Base 1. This sequence was performed to evaluate the potential anti-ischaemic effects of atenolol and ivabradine during exercise and their consequences on subsequent myocardial stunning during the recovery period. Ivabradine was administered as an i.v. bolus (1mg/kg over 5min) followed by a continuous i.v. infusion (0.5mg/kg/h) during 6h using an automatic programmable pump which was fixed on the back of the animal. We previously demonstrated that this regimen of administration of the drug induces a significant heart rate reduction which remains stable during the infusion period.9 Atenolol was administered as an i.v. bolus (1mg/kg over 5min). As previously reported,16 the regimen of ivabradine and atenolol provide similar heart rate reduction at rest and during exercise. In the second part of the protocol (sequence B), administration of atenolol or ivabradine was started immediately after the end of exercise. Sequence B was set up to investigate the direct effects of heart rate reduction on the already stunned myocardium, thus independently from the potential anti-ischaemic properties of the drugs.
Each recording made at rest before exercise and during the recovery period was performed both at spontaneous heart rate and during a 1-min episode of atrial pacing at 150beats/min in order to individualize the effects of heart rate reduction per se. Each animal performed the five experimental sessions (sequence A with saline, ivabradine, atenolol and sequence B with ivabradine and atenolol) in random order with at least a 5-day interval.
Statistical analysis
Data are reported as mean±SEM. The experiments were conducted as an incomplete design in which each dog received all five treatments in a randomized order. Data during stunning were analysed using two-way ANOVA for repeated measures (repeated times nested in treatments) and by checking for interactions. One way ANOVA were performed to analyse baseline values. The FisherSnedecor test was used to test the significance of analysis of variance. When needed, pairwise comparisons between (a) saline and atenolol, (b) saline and ivabradine and (c) ivabradine and atenolol were performed using a paired Student t-test with the Bonferroni correction. Analyses were performed separately for experimental designs under sequence A and those under sequence B. Significance was accepted at P<0.05.
Results
Heart rate
Values of heart rate at Base 1 were not significantly different and no further interaction was detected using ANOVA.
As illustrated in Fig. 1, heart rate increased from 109±7 to 217±13beats/min during exercise performed under saline in sequence A. Heart rate was similarly reduced by atenolol and ivabradine at rest (86±2 and 86±7beats/min, respectively) and during exercise (154±2 and 155±9beats/min, respectively). Throughout the recovery period, heart rate under atenolol and ivabradine remained constant and significantly reduced ascompared to saline (e.g. at 1h, 100±4 and 103±5 vs 126±7beats/min, respectively). As shown in Table 1, heart rate values were similar between saline, atenolol and ivabradine during the 1min of atrial pacing performed for each data recording at rest and during recovery.
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Haemodynamics
Values of mean arterial pressure, left ventricular pressure and end-diastolic left ventricular pressure measured at Base 1 were not significantly different and no further interaction was detected using ANOVA.
As heart rate reduction can per se modify LV wall thickening,9 it represents a confounding factor in the interpretation of the results and accordingly in sequence A, only values measured under atrial pacing (150beats/min) at baseline and during the recovery period were analysed.
In sequence A during atrial pacing (Table 1), mean arterial pressure was significantly reduced at rest, during exercise and the recovery period with administration of atenolol but not ivabradine. LV end-diastolic pressure and LV pressure were not significantly different among saline, atenolol and ivabradine.
In sequence B, administration of atenolol and ivabradine at exercise completion produced similar effects to those described in sequence A.
LV wall thickening in the ischaemic zone
Values of LV wall thickening and end-diastolic wall thickness measured at Base 1 were not significantly different. However, significant interactions treatment x time were detected using ANOVA for LV wall thickening during the stunning period in both sequences A and B.
In sequence A, at Base 2 during atrial pacing under atenolol but not ivabradine, LV wall thickening was significantly reduced as compared to saline (Table 2, Fig. 2). These effects were similar at spontaneous heart rate except that LV wall thickening was significantly increased in the session with ivabradine (data not shown). During exercise under saline, LV wall thickening decreased dramatically to 2±1% from 23±4% but this decrease was significantly reduced in the session with ivabradine (LV wall thickening: 10±3%) and to a significantly greater extent in the session with atenolol (LV wall thickening: 17±4%). During the recovery period, LV wall thickening measured during atrial pacing remained depressed for several hours under saline, indicating myocardial stunning. Values under atenolol were similar to those measured under saline. In contrast, LV wall thickening was significantly improved under ivabradine throughout the recovery period. Similar effects were observed at spontaneous heart rate (data not shown).
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In sequence A, at Base 2 during atrial pacing, LV wall thickening was significantly reduced after administration of atenolol as compared to saline (Table 2). This parameter was not altered under ivabradine. These effects were similar at spontaneous heart rate except that LV wall thickening was significantly increased during the administration of ivabradine (data not shown). During exercise under saline, LV wall thickening increased up to 52±9% from 28±2% in sequence A. In the session with ivabradine, similar effects were observed but LV wall thickening was significantly limited to 34±6% under atenolol (P<0.05 vs saline and ivabradine). During the recovery period, LV wall thickenings measured during atrial pacing under saline and ivabradine were similar but depressed after administration of atenolol. At spontaneous heart rate, LV wall thickening was significantly greater under ivabradine as compared to saline whereas it was smaller with atenolol (e.g. at 1h recovery, 28±3 and 40±5% for atenolol and ivabradine, respectively vs 35±5% for saline).
In sequence B, LV wall thickening was similar at Base 2 and during exercise in the three experimental sessions. During recovery, LV wall thickening was significantly greater under ivabradine than saline and depressed after administration of atenolol at spontaneous heart rate (e.g. at 1h recovery, 28±5 and 40±4 for atenolol and ivabradine, respectively vs 35±5% for saline). Under atrial pacing, LV wall thickening was similar between saline and ivabradine but still significantly reduced during the session with atenolol (Table 2).
Diastolic perfusion time
At rest, the diastolic perfusion time tended to be increased during sessions with atenolol and ivabradine as compared to saline (458±39ms, P=0.16 and 478±23ms, P=0.06 vs 379±40ms). This trend was reinforced during exercise under atenolol (199±5ms vs saline: 135±13ms, P<0.05) and to a greater extent for ivabradine (241±18ms, P<0.05 vs saline and atenolol). During the recovery period, such significant differences remained (e.g. at 2h: 388±24ms and 411±27ms for atenolol and ivabradine, respectively vs 298±25ms for saline). The latter effect was abolished by atrial pacing for ivabradine.
LV regional myocardial blood flows
For technical reasons, measurements of RMBFs were performed in five dogs only (defective blood withdrawal in two dogs). During exercise performed under saline, transmural RMBFs increased markedly in the non-ischaemic zone, but remained unchanged in the ischaemic zone due to the coronary stenosis (Table 3). Simultaneously, the endo/epi ratio fell significantly in the ischaemic zone whereas it remained unchanged in the non-ischaemic zone.
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Discussion
Decrease in heart rate is one of the goals to achieve for the treatment of ischaemic heart disease. In this setting, If inhibitors form a novel class of heart rate reducing agents which may be useful to achieve this goal. The present results confirm our previous results, i.e., that heart rate reduction induced by an If inhibitor is of major importance during exercise induced myocardial ischaemia and subsequent stunning.9 This study demonstrates that the negative inotropic effect of a ß-blocker such as atenolol modulates the cardioprotection afforded by heart rate reduction, either positively or negatively, depending on the time of administration. Indeed, heart rate reduction provides a powerful anti-ischaemic effect and additional negative inotropism afforded by atenolol tends to enhance this effect. Conversely, administered after exercise-induced ischaemia, i.e., when the myocardium is stunned, regional contractility is improved by selective heart rate reduction with ivabradine but is dramatically deteriorated by the negative inotropism of atenolol.
The experimental model used in this study enabled to investigate the anti-ischaemic effects associated with the reductions respectively in heart rate and in LV inotropism as these parameters were strongly and physiologically enhanced during treadmill exercise. The combination of an exercise and a coronary artery stenosis induced a severe regional imbalance between myocardial metabolic demand and oxygen supply.9,14,17 This resulted in a strong decrease in regional contractility in the ischaemic zone and a subsequent myocardial stunning. Importantly, in this model, repetition of exercises does not induce any late preconditioning-like effect as all experiments were performed at least 5 days apart, a delay after which any potential late preconditioning has vanished.18 Furthermore, late preconditioning against stunning does not occur after exercise-induced ischaemia in conscious dogs.14
In this study, selective heart rate reduction during the ischaemic insult induced by exercise clearly reduced regional myocardial contractile dysfunction in agreement with previous studies.8,9,19 Recently, we demonstrated that the beneficial effect of ivabradine was due to its negative chronotropic property as atrial pacing abolished its anti-ischaemic effects.9 This cardioprotection appears likely due to an enhanced diastolic perfusion time, an increased subendocardial perfusion and as previously reported,16,20 a decreased myocardial oxygen demand. In animal models using ameroïds, atenolol provided also a strong anti-ischaemic effect as previously reported.2,7 In our experimental conditions, in addition to the reduction in heart rate, the negative inotropic effect of atenolol tends to provide some anti-ischaemic effect during exercise. This could be due to a more favourable balance between oxygen demand and supply and we previously reported that atenolol decreases markedly more myocardial oxygen consumption than ivabradine during exercise in the normal heart.16 In addition, with ß-blockade, the auto-regulated increase in vascular resistance in the distal bed improves subendocardial perfusion.6 Finally, as transmural myocardial blood flows were similar among exercises, differences in blood supply levels due to different coronary stenosis levels cannot explain these differences between drugs.
One of the main results of this study is that opposite effects were observed depending on the time of administration of ivabradine and atenolol. As heart rate reduction can per se modify LV wall thickening,9 we thought that this factor should be taken into account for the interpretation of the cardioprotective effect of ivabradine and atenolol. Accordingly in sequence A, only values measured under atrial pacing (150beats/min) at baseline and during the recovery period were analyzed. In these conditions, administration of ivabradine prior to ischaemia dramatically reduced the severity of myocardial stunning during recovery. In contrast, LV wall thickening measured under atenolol was altered at baseline and remained depressed throughout the 6h of the recovery period and similar to that under saline. This means that the negative inotropic effect of atenolol generated dual but opposite effects, i.e., a cardioprotective one during ischaemia and a deleterious one during myocardial stunning. In fact, the negative inotropism of atenolol overrides its anti-ischaemic effect and further abrogates the cardioprotection afforded by heart rate reduction on regional myocardial contractility. In contrast, selective heart rate reduction with ivabradine attenuated myocardial dysfunction during ischaemia and further extended this effect to the protection against myocardial stunning.
This crucial role of the time of administration is also illustrated in sequence B when drugs were given at exercise completion, i.e., under conditions independent from any previous anti-ischaemic effect. As we previously demonstrated,9 selective heart rate reduction induced after ischaemia by ivabradine resulted in a significant enhancement of regional wall thickening as compared to saline. As this effect was abolished by atrial pacing, it appears solely due to heart rate reduction and not related to any direct action on the cardiomyocyte of the ischaemic zone, i.e., not related to a positive inotropic effect. It can be hypothesized that heart rate reduction improves the diastolic time for LV filling,16 increases the end-diastolic LV volume and finally enhances the LV regional contractility through a FrankStarling mechanism. Indeed, the enhancement of myocardial contractility was also observed in the non-ischaemic zone and ivabradine increased LV end-diastolic diameter in the normal heart (data not shown). In contrast, atenolol clearly deteriorated LV wall thickening of the stunned myocardium as a consequence of its negative inotropic effect.
Some limitations of this study should be addressed. Only one dose of atenolol was used in this study and therefore it is possible that different results would be observed with other beta-blockers eliciting other pharmacological profile or with smaller doses of atenolol. These findings should also not be extended to other situations with chronic LV dysfunction and likely different pathophysiological mechanisms, as beneficial effects with small doses of ß-blockers are well recognized in the setting of chronic heart failure.21 Finally from a statistical point of view, since the cross-over design is unbalanced in this study, the treatment effects cannot be unambiguously attributed to the treatment per se and could also include period and/or wash-out effects. In conclusion, this study highlights the fundamental role of reducing heart rate in the protection of the ischaemic and stunned myocardium induced by exercise associated with a coronary stenosis. In the case of atenolol, despite its negative inotropic effect which tends to enhance the anti-ischaemic effect afforded by heart rate reduction during ischaemia, it aggravates the LV systolic dysfunction of the stunned myocardium. In contrast, selective heart rate reduction with ivabradine not only affords cardioprotection during exercise-induced ischaemia but also strongly enhances regional contractility of the stunned myocardium. Although the recovery from myocardial ischaemia depends from many more factors including LV function at baseline, this might have important clinical implications during LV systolic dysfunctions, especially in the setting of chronic and repeated myocardial stunning as ivabradine has been demonstrated to be a potent anti-ischaemic agent in humans.22
Acknowledgments
Dr Patrice Colin was supported by the Société Française de Pharmacologie. We thank Dr F. Mahlberg, Dr J. P. Vilaine and Dr G. Lerebourg from Laboratoires Servier for fruitful discussions during the elaboration of this manuscript. The authors are greatly indebted to Dr J. X. Mazoit for statistical advises. We wish also to thank Alain Bizé and Stéphane Bloquet for their excellent technical assistance.
Footnotes
1 X. Monnet and P. Colin both contributed equally to this work
2 Doi:10.1016/j.ehj.2003.11.005.
References
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