Naloxone improves functional recovery of myocardial stunning in conscious dogs through its action on the central nervous system
T. P. Weber1,
J. Stypmann2,
A. Meißner1,
M. Große Hartlage1,
H. Van Aken1 and
N. Rolf1
1Klinik und Poliklinik für Anästhesiologie und operative Intensivmedizin, Westfälische Wilhelms-Universität, Albert-Schweitzer-Strasse 33, D-48145 Münster, Germany. 2Klinik und Poliklinik für Kardiologie und Angiologie, Innere Medizin C, Westfälische Wilhelms-Universität, Albert-Schweitzer-Strasse 33, D-48145 Münster, Germany
Accepted for publication: October 4, 2000
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Abstract
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This study tests the hypothesis that naloxone, but not its quarternary salt, naloxone methiodide (which does not enter the central nervous system), improves recovery from myocardial stunning in conscious dogs. Twenty dogs were chronically instrumented for measurement of heart rate, left atrial, aortic and left ventricular pressure (LVP), LV dPdtmax1 and myocardial wall thickening fraction (WTF). Regional myocardial blood flow was determined with coloured microspheres. Occluder around the left anterior descending artery (LAD) allowed induction of reversible LAD ischaemia. Each of the 20 dogs underwent two LAD ischaemic challenges. Experiments (performed on separate days, in crossover fashion) were: (i) 10 min of LAD occlusion after application of naloxone 63 µg kg1 or naloxone methiodide 63 µg kg1 and (ii) occlusion without naloxone or naloxone methiodide. WTF was measured at baseline and until complete recovery occurred. LAD ischaemia significantly reduced LAD WTF with (mean (SD) 54 (15)% lower than baseline) and without naloxone (55 (16)% lower than baseline), without significant haemodynamic differences. Between 1 to 30 min of reperfusion, WTF was significantly higher with naloxone (P<0.05). There was no difference in WTF with or without naloxone methiodide. We conclude that naloxone improved recovery from myocardial stunning in conscious dogs, and that this was centrally mediated.
Br J Anaesth 2001; 86: 5459
Keywords: heart, contractility; blood, flow, regional; blood, reperfusion; heart, ventricular function; antagonists opioid
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Introduction
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Endogenous opioid peptides (EOPs) have important roles in cardiovascular regulation and are released upon myocardial ischaemia.1 There are three major families of EOPs: dynorphin (derived from pre-proenkephalin B), endorphins (derived from pre-pro-opiomelanocortin) and enkephalins (derived from pre-proenkephalin A). The effects of EOPs are probably mainly mediated by opioid receptors.2 Pharmacological studies have led to the proposal of five classes of opioid receptor (µ,
,
,
and
), but the pharmacological effects of each receptor type are not completely defined and this classification has been questioned. It remains unclear if EOPs contribute to myocardial stunning, a clinically relevant phenomenon in patients with coronary artery disease or those undergoing cardiac surgery. Myocardial stunning is a general term for the mechanical dysfunction that persists after reperfusion even in the absence of irreversible damage and despite the return of normal, near-normal or supranormal perfusion.35
The present investigation tests the hypothesis that naloxone, a non-selective opioid receptor antagonist that enters the central nervous system,6 reduces the severity and duration of myocardial stunning in an experimental model of reversible myocardial ischaemia. Control studies were performed with naloxone methiodide, which does not cross the bloodbrain barrier.6
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Methods
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This investigation conforms to the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication no. 85-23, revised 1996) and was approved by the District Government of Münster. After overnight fasting, 20 mongrel dogs (10 male and 10 female, weight 2428 kg) received intramuscular premedication with piritramide 1 mg kg1 and ketamine 5 mg kg1. The animals were anaesthetized intravenously with propofol 5 mg kg1. After tracheal intubation, anaesthesia was maintained with isoflurane in a mixture of oxygen (3540%) in air. Perioperative antibiotic prophylaxis was achieved with cefamandole 30 mg kg1. Details of the instrumentation methods have been published before.7 Briefly, a left thoracotomy was performed in the fifth intercostal space under aseptic conditions. Eighteen-gauge catheters were inserted into the descending aorta and the left atrium for pressure measurement, injection of microspheres, naloxone and naloxone methiodide, and withdrawal of blood. A pressure microtransducer (Janssen Pharmaceutica, Beerse, Belgium) was inserted into the left ventricle through an apical stab wound to measure left ventricular pressure (LVP). Pulsed Doppler blood flow velocity probes (20 MHz; Baylor College of Medicine, Houston, TX, USA) were fitted around the left anterior descending coronary artery (LAD). To measure the regional myocardial wall thickening fraction (WTF), 10 MHz pulsed Doppler crystals were sutured to the myocardium in the LAD-perfused areas. Proximal to the Doppler flow probe, a pneumatic occluder was positioned around the LAD (proximal to the first main diagonal branch) to induce reversible brief ischaemic episodes in the LAD-perfused myocardium. After closure of the thorax, all leads were tunnelled subcutaneously and exteriorized between the scapulae. After instrumentation, the animals were trained daily to accustom them to the experimental environment and to ensure that they could lie quietly in the cage when connected to the data acquisition system. Aortic and left atrial pressures were measured using disposable pressure transducers. Pressure, flow velocity and wall thickening signals were processed using a six-channel pulsed Doppler system (Baylor College of Medicine). The left ventricular micromanometer was calibrated to pressures measured in the aorta and left atrium. The LVP signal was differentiated electronically (Gould Inc., Cleveland, OH, USA) and all signals were digitally recorded. Experiments were not conducted until the animals had recovered completely from instrumentation and until blood gas and haemodynamic variables were in the normal range. This took 710 days after surgery.
Each of the 20 dogs was given two ischaemic challenges, one with no pretreatment and the other with either naloxone (Curamed Pharma Karlsruhe, Germany) or naloxone methiodide (Sigma-Aldrich Chemie, Deisenhofen, Germany)8 63 µg kg1 i.v. 5 min before ischaemia. Ten animals received their first coronary artery occlusion without pretreatment. Baseline values were measured in the awake state, 10 min of LAD ischaemia was induced and WTF was monitored until complete recovery occurred. The other 10 animals received naloxone or naloxone methiodide before their first ischaemia to elucidate the site (central or peripheral) of action. In this group, baseline values were measured in the awake state, naloxone 63 µg kg1 or naloxone methiodide 63 µg kg1 was given before induction of 10 min of LAD ischaemia and WTF was monitored until complete recovery occurred.
A second ischaemic episode was only induced when there was complete recovery of regional myocardial function in the LAD-perfused area; the minimum time interval between the two experiments was 72 h. Regional myocardial blood flow was measured using coloured microspheres (Triton Technology, San Diego, CA, USA). For each measurement, 9x106 microspheres suspended in a volume of 3 ml were injected into the left atrium. The reference blood sample was withdrawn from the aortic catheter at a rate of 10 ml min1. Animals were killed after complete recovery of WTF, after the last ischaemic episode. The heart was dissected and three tissue samples were obtained from the LAD-perfused left ventricle in each dog. LAD samples were taken from the immediate vicinity of the wall thickening probes. Only samples from animals with severe ischaemic dysfunction, as determined by the wall thickening probe, were included. Samples were further dissected into the subendocardial, subepicardial and mid-myocardial layers. Measurement of microspheres in the tissue samples was performed as described previously.9 Measurement of regional myocardial blood flow to the regions described was carried out four times during the experiment, as follows: (i) without naloxone/naloxone methiodide and without ischaemia (control); (ii) without naloxone/naloxone methiodide and with ischaemia; (iii) with naloxone/naloxone methiodide and without ischaemia; and (iv) with naloxone/naloxone methiodide and with ischaemia.
Experiments were conducted in chronically instrumented conscious dogs to avoid the effects of acute surgical trauma, anaesthesia, volume and ionic imbalances, and temperature on recovery from stunning. Since multiple stun manoeuvres may induce extensive development of coronary collaterals, precluding the induction of postischaemic dysfunction, the number of ischaemic episodes was restricted to two in each animal.
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Statistical analysis
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Data were analysed using repeated-measures two-way analysis of variance followed by Bonferroni-corrected Students t-test whenever appropriate; P<0.05 was regarded as significant. Data are presented as mean (SD).
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Results
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None of the animals had to be excluded from the analysis due to insufficient dysfunction. The maximum degree of regional ischaemic dysfunction was similar during the first and the second ischaemic episodes in each dog.
Blood pressure, heart rate, left atrial pressure and LV dP dt1
There were no significant changes in arterial blood pressure in either group during or after ischaemia (Tables 1 and 2). Left atrial pressure and heart rate during LAD ischaemia increased significantly in both groups, but there was no significant difference in these variables between the two groups during or after ischaemia. Induction of regional ischaemia did not produce a significant change in LV dP dtmax1 (Tables 1 and 2). After naloxone administration, LV dP dtmax1 was significantly higher than in the controls during ischaemia and until 30 min of reperfusion (Table 1). Results with naloxone methiodide were the same as those with controls (Table 2).
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Table 1 Arterial blood pressure (ABP), left atrial pressure (LAP), rate of increase in left ventricular pressure (LV dP dtmax1), heart rate (HR) and blood flow velocity in the left anterior descending coronary artery (BFV LAD) for the control and naloxone groups at baseline, during ischaemia and at predetermined time points during reperfusion. Data are presented as mean (SD).*Significantly different from baseline. #Significantly different from control
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Table 2 Arterial blood pressure (ABP), left atrial pressure (LAP), rate of increase in left ventricular pressure (LV dP dtmax1), heart rate (HR) and blood flow velocity in the left anterior descending coronary artery (BFV LAD) for the control and naloxone methiodide groups at baseline, during ischaemia and at predetermined time points during reperfusion. Data are presented as mean (SD). *Significantly different from baseline. #Significantly different from control
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Blood flow velocity in the LAD
Blood flow velocity in the LAD-perfused area in the experimental groups was not significantly different from baseline values. During LAD occlusion, LAD flow velocity decreased to zero and increased to values significantly higher than baseline for the first 10 min during reperfusion. There were no significant differences between the experimental groups in flow velocities in the LAD during reperfusion (Tables 1 and 2).
Regional myocardial wall thickening
The WTF in the LAD-perfused area is shown in Figure 1A and B for the naloxone and naloxone methiodide group, respectively. In all animals, severe myocardial dysfunction occurred during LAD ischaemia. Induction of regional ischaemia led to a significant reduction in WTF to negative values (wall thinning) in both experimental conditions; in the presence (mean (SD) 54 (15)% lower than baseline) or absence of naloxone (55 (16)% lower than baseline), and in the presence (57 (25)% lower than baseline) or absence of naloxone methiodide (56 (19)% lower than baseline). After reperfusion, WTF recovered significantly more quickly only in the group receiving naloxone at time points between 1 min and 30 min of reperfusion (92 (9)% vs 25 (46) at 1 min; 100 (5)% vs 37 (54)% at 5 min; 97 (5)% vs 42 (47) at 15 min; 95 (10)% vs 49 (34)% at 20 min; and 90 (7)% vs 53 (33)% at 30 min). WTF had returned to baseline after 48 h of reperfusion in the presence or absence of naloxone.

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Fig 1 Wall-thickening fraction (WTF) of the area perfused by the left anterior descending coronary artery as a percentage of control values during the reperfusion period for naloxone (A) and naloxone methiodide (B). Data are presented as mean±SD and P values refer to between-group differences. BL=baseline values. N=naloxone. NS=not significant. Isch=ischaemia (10 min). *P<0.05.
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Regional myocardial blood flow
During LAD occlusion, subendocardial blood flow decreased significantly, from 0.71 (0.22) to 0.09 (0.01) ml g1 min1 in the absence of naloxone, from 1.03 (0.25) to 0.11 (0.03) ml g1 min1 in the presence of naloxone (P<0.05), from 0.69 (0.16) to 0.07 (0.01) ml g1 min1 in the absence of naloxone methiodide, and from 0.72 (0.28) to 0.10 (0.01) ml g1 min1 in the presence of naloxone methiodide. Only administration of naloxone, and not administration of naloxone methiodide, in the absence of ischaemia significantly increased subendocardial blood flow of the LAD-perfused areas (0.71 (0.22) vs 1.03 (0.25) ml g1 min1). Without ischaemia, only naloxone significantly increased blood flow ratio between the subendocardial and the subepicardial layers (endocardial/epicardial) in the LAD-perfused area (1.13 (0.66) vs 2.06 (1.17) ml g1 min1). The blood flow ratio to the subendocardial layers between the occluded zone (LAD-perfused area) (occluded/normal) during ischaemia was not different with and without naloxone, or with and without naloxone methiodide.
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Discussion
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In this model, naloxone improved the functional recovery from myocardial stunning and improved ischaemic and postischaemic systolic left ventricular performance. Doseresponse studies of naloxone in dogs are not available, but naloxone has been used for several scientific purposes in dogs. The dose used in our study (63 µg kg1) antagonizes the µ-agonist effects in dogs.8 EOPs are found in central areas near the cardiovascular centers of the brain, in the hypothalamus, and peripherally in the myocardium.10 Naloxone exerts central and peripheral effects on heart tissue itself.11 However, the above-described protective effects of naloxone are probably mainly mediated by antagonistic action against central opioid receptors.2 The opioid receptor antagonist naloxone methylbromide, which unlike naloxone does not cross the bloodbrain barrier, did not produce an increase in regional blood flows in dogs with right-sided congestive heart failure.12 These data are in agreement with our findings. Central effects of naloxone are therefore predominant and probably occur mainly in the hypothalamus.13 The central sites at which naloxone exerts this action have not been determined. EOPs, released upon myocardial ischaemia,1 activate central opioid receptors, leading to sympathoinhibition and contributing to negative inotropic effects on the myocardium. Experimental data suggest that the increased endogenous opioids during heart failure act mainly at
-receptors14 and
-receptors.15 Activation of these receptors decreases myocardial mechanical performance and alters regional blood flow distribution. It has been suggested that the effects of sigma receptor ligands are mediated via two receptor subtypes, of which one has positive and the other negative inotropic activity.16 It is not clear which effect is predominant. Pharmacological studies led to the proposal that myocardial
-opioid receptors do not play an important role, at least in cardiac rhythm.17 In contrast, antiarrhythmic effects mediated by
-,18 µ-,
-receptor agonists and
-receptor-mediated proarrhythmic effects have been described.17 Interestingly, µ-receptors are only expressed at early periods in heart ontogeny. Also, there is a period in which the density of µ- (but not
- and
-) receptors decreases dramatically.19 This might explain, at least in part, the dose-related variability and species differences of opioid receptor-mediated effects.20 Selective opioid receptor blockade might provide a starting point for effective treatment of the negative inotropic effects of myocardial stunning. However, further investigations in this area will be needed.
It should be borne in mind that the results obtained here are restricted to dogs; there may be species differences in the severity and duration of the ischaemic response and its modulation by naloxone. Also, the lack of a doseresponse precludes extrapolation of the data to other doses of naloxone.
In conclusion, naloxone improves ischaemic and postischaemic systolic impairment and reduces the severity of myocardial stunning in chronically instrumented dogs. This study is the first to suggest that central opioid receptors are involved in recovery from myocardial stunning.
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