u-50488 and Myocardial-Infarction

The role of opioid kappa1 and kappa2 receptors in reperfusion cardiac injury was studied. Male Wistar rats were subjected to a 45-min coronary artery occlusion followed by a 120-min reperfusion. Opioid kappa receptor agonists were administered intravenously 5 min before the onset of reperfusion, while opioid receptor antagonists were given 10 min before reperfusion. The average value of the infarct size/area at risk (IS/AAR) ratio was 43 - 48% in untreated rats. Administration of the opioid kappa1 receptor agonist (-)-U-50,488 (1 mg/kg) limited the IS/AAR ratio by 42%. Administration of the opioid kappa receptor agonist ICI 199,441 (0.1 mg/kg) limited the IS/AAR ratio by 41%. The non-selective opioid kappa receptor agonist (+)-U-50,488 (1 mg/kg) with low affinity for opioid kappa receptor, the peripherally acting opioid kappa2 receptor agonist ICI 204,448 (4 mg/kg) and the selective opioid ?2 receptor agonist GR89696 (0.1 mg/kg) had no effect on the IS/AAR ratio. Pretreatment with naltrexone, the peripherally acting opioid receptor antagonist naloxone methiodide, or the selective opioid kappa2 receptor antagonist nor-binaltorphimine completely abolished the infarct-reducing effect of (-)-U-50,488 and ICI 199,441. Pretreatment with the selective opioid ? receptor antagonist TIPP[psi] and the selective opioid µ receptor antagonist CTAP did not alter the infarct reducing effect of (-)-U-50,488 and ICI 199,441. Our study is the first to demonstrate the following: (a) the activation of opioid kappa2 receptor has no effect on cardiac tolerance to reperfusion; (b) peripheral opioid kappa1 receptor stimulation prevents reperfusion cardiac injury; (c) ICI 199,441 administration resulted in an infarct-reducing effect at reperfusion; (e) bradycardia induced by opioid kappa receptor antagonists is not dependent on the occupancy of opioid kappa receptor.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Administration, Intravenous; Analgesics, Opioid; Animals; Arrhythmias, Cardiac; Disease Models, Animal; Heart Rate; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Myocytes, Cardiac; Narcotic Antagonists; Piperazines; Pyrrolidines; Rats, Wistar; Receptors, Opioid, kappa; Signal Transduction

Previous studies have shown that κ-opioid receptor activation possesses cardioprotection against myocardial ischemia and reperfusion (MI/R) injury. The current study was designed to investigate whether mitochondrial dysfunction after MI/R is regulated by the κ-opioid receptor and to further explore the underlying mechanisms involved. MI/R rat model was established in vivo, and a hypoxia and reoxygenation cardiomyocytes model was used in vitro. Mitochondrial morphology and function as well as myocardial apoptosis were determined. Our data indicated that treatment with U50,488H (a selective κ-opioid receptor agonist) not only reduced apoptosis but also significantly improved mitochondrial morphology and function. These effects were blocked by nor-binaltorphimine (nor-BNI, a selective κ-opioid receptor antagonist), Compound C (an AMPK inhibitor), and AR-A014418 (a GSK3β inhibitor). Moreover, in cardiomyocytes, treatment with U50,488H significantly increased the expression in phosphorylation of AMPK and the phosphorylation of GSK3β. Treatment of cardiomyocytes with AMPKα siRNA decreased the phosphorylation of AMPK and GSK3β. Moreover, AMPK activation resulted in the phosphorylation of GSK3β. Our findings suggested that U50,488H exerted cardioprotective effects by improving mitochondrial morphology and function against MI/R injury through activation of the κ-opioid receptor-mediated AMPK/GSK3β pathway.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; AMP-Activated Protein Kinases; Animals; Apoptosis; Cell Line; Disease Models, Animal; Glycogen Synthase Kinase 3 beta; Male; Mitochondria, Heart; Myocardial Infarction; Myocardial Reperfusion Injury; Myocytes, Cardiac; Phosphorylation; Rats, Sprague-Dawley; Receptors, Opioid, kappa; Signal Transduction

To keep U50488H from going through the blood-brain barrier, U50488H, a selective κ-opioid receptor agonist, was structurally transformed into its quaternary ammonium salt (Q-U50488H). The effect of Q-U50488H on ischemic/reperfused myocardium and its underlying mechanisms were also investigated. U50488H was transformed into Q-U50488H, which was identified with mass spectrometry. The existences of U50488H and Q-U50488H in serum and brain tissue fluid were determined by high performance liquid chromatography (HPLC). SD rats' hearts were subjected to 30min of ischemia followed by 120min of reperfusion in vivo. After reperfusion, myocardial enzymes and free radicals in serum, area of myocardial infarction, cardiomyocyte apoptosis and the expression of Kir6.2 in rats' myocardium were determined. Molecular weight and solubility of Q-U50488H were higher than those of U50488H. Result of HPLC showed that Q-U50488H existed in serum but not in brain tissue after Q-U50488H intravenous injection. However, U50488H was detected in both serum and brain tissue. Compared with the I/R group, treatment with Q-U50488H significantly attenuated the activity of LDH, CK, AST/GOT and content of MDA in serum, upregulated the activity of SOD, and increased the expression of myocardial Kir6.2. It also reduced myocardial infarct size and cardiomyocyte apoptosis induced by I/R. Moreover, pretreatment with Nor-BNI (a selective κ-opioid receptor antagonist), 5-HD and glibenclamide (KATP antagonists) abolished the effects of Q-U50488H. It is suggested Q-U50488H, a new compound of κ-opioid receptor agonist, which is not able to pass the blood-brain barrier, elicits a protective effect against myocardial ischemia/reperfusion injury. The cardioprotective effect of Q-U50488H is associated with the opening of KATP.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Apoptosis; Brain; Cardiotonic Agents; Gene Expression Regulation; Heart; Male; Malondialdehyde; Myocardial Infarction; Myocardial Reperfusion Injury; Myocardium; Potassium Channels, Inwardly Rectifying; Quaternary Ammonium Compounds; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa

To observe the effects of κ-opioid receptor agonist U50, 488H on myocardial ischemia and reperfusion injury and related mechanism.. Rats were randomly divided into sham operation, myocardial ischemia and reperfusion(I/R, 30 min ischemia followed by 120 min reperfusion), and MI/R+U50, 488H (1.5 mg/kg) and I/R+U50, 488H+ selective κ-opioid receptor antagonist Nor-BNI (2 mg/kg, n = 8 each). The infarction size and the incidence of ventricular arrhythmias were observed.Real-time PCR and DAB staining were used to define the myocardium Toll-like receptor 4(TLR4) expression. Myeloperoxidase level, TNF-α induction and the expression of NF-κB were also examined in rats.. After I/R, the expressions of myocardial TLR4 and NF-κB increased significantly both in ischemia area and area at risk. Compared with I/R, κ-opioid receptor stimulation with U50, 488H significantly attenuated the expressions of TLR4 and NF-κB and reduced myeloperoxidase (MPO) levels, myocardial TNF-α production, myocardial infarct sizes and the incidence of ventricular arrhythmias and arrhythmia score (2.9 ± 0.7 vs. 4.4 ± 0.9, P < 0.05) , above effects of U50, 488H were partly abolished by co-treatment with Nor-BNI.. These data provide evidence for the first time that κ-opioid receptor stimulation could attenuate myocardial I/R injury via downregulating TLR4/NF-κB signaling in rats.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Arrhythmias, Cardiac; Brugada Syndrome; Cardiac Conduction System Disease; Coronary Artery Disease; Down-Regulation; Heart Conduction System; Myocardial Infarction; Myocardial Ischemia; Myocardium; Naltrexone; NF-kappa B; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa; Reperfusion Injury; Signal Transduction; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

It is well documented that the Toll-like receptor 4 (TLR4)/NF-κB signaling mediates early inflammation during myocardial ischemia and reperfusion. Our previous study has demonstrated that κ-opioid receptor stimulation with U50,488H produces cardioprotective and anti-inflammatory effects. The aim of the present study was to investigate whether κ-opioid receptor stimulation could modulate the TLR4/NF-κB signaling and reduce neutrophil accumulation and TNF-α induction in an ischemia-reperfusion injured rat heart model. Rats were randomly exposed to sham operation, myocardial ischemia and reperfusion (MI/R), and MI/R+U50,488H in the absence or presence of Nor-BNI, a selective κ-opioid receptor antagonist. The results demonstrated that after MI/R, the expressions of myocardial TLR4 and NF-κB increased significantly both in ischemia area and risking area. Compared with MI/R, κ-opioid receptor stimulation with U50,488H significantly attenuated the expressions of TLR4 and NF-κB. At the mean time, it also reduced myeloperoxidase (MPO) levels, both serum and myocardial TNF-α production, myocardial infarct sizes (INF/AAR%) and myocardial apoptosis induced by MI/R, all the effects of U50,488H were abolished by Nor-BNI. These data provide evidence for the first time that κ-opioid receptor stimulation inhibits TLR4/NF-κB signaling in the rat heart subjected to MI/R.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Apoptosis; Down-Regulation; Gene Expression Regulation; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Myocardium; Neutrophil Infiltration; NF-kappa B; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa; Signal Transduction; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

Kappa-opioid receptors (κ-OR) and mechanoelectric feedback seem to have common pathways that influence electrophysiological changes resulting from acute myocardial infarction (MI). This study aims to determine the effects of the κ-OR on stretch-induced electrophysiological changes after acute MI.. Male Sprague-Dawley rats were randomly divided into 4 groups: sham operated, MI, U-50488H (a selective κ-OR agonist) -treated MI (MI+U-50488H) and nor-BNI (a selective κ-OR antagonist) -treated MI (MI+nor-BNI). After Langendorff perfusion to maintain stabilization, a transient stretch (5 seconds) was delivered early in diastole. Electrophysiological changes were recorded for 1 minute before and after stretch. Similarly, the 20%, 50% and 90% monophasic action potential duration (MAPD20, MAPD50 and MAPD90, respectively) and stretch-induced arrhythmias were recorded.. MAPD90 significantly increased in all 4 groups. MAPD90 in the MI and MI+nor-BNI groups increased significantly before stretch (P < 0.05) and after stretch (P < 0.01) but was reversed in the MI+U-50488H group (P > 0.05). MAPD90 in the MI group was increased compared with that of the MI+U-50488H group but decreased compared with that of the MI+ nor-BNI group after stretch (P < 0.01). The arrhythmia score in the MI and MI+nor-BNI groups was higher than that of the sham-operated group (P < 0.01), and the arrhythmia score in the MI+nor-BNI group was higher than that in MI group after stretch (P < 0.01). The arrhythmia score of the MI+U-50488H group was lower than that of MI group after stretch (P < 0.01).. The κ-OR could influence the stretch-induced electrophysiological changes and play an antiarrhythmic role in stretch-induced arrhythmias after acute MI.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Action Potentials; Animals; Arrhythmias, Cardiac; Electrophysiological Phenomena; Male; Myocardial Infarction; Naltrexone; Pressoreceptors; Random Allocation; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa

It remains unclear whether U50488H (a selective kappa-opioid receptor agonist) produces anti-apoptotic effect during ischemia and reperfusion (I/R). Therefore, the effect of U50488H on myocardial apoptosis was investigated in the present study.. Rats were subjected to 45min coronary artery occlusion and 180min of reperfusion. U50488H (1.5mg/kg IV) was given prior to occlusion. Nor-Binaltorphimine (nor-BNI) (2mg/kg IV), a selective kappa-opioid receptor antagonist, was given 10min prior to U50488H. Cardiac apoptosis was evaluated by terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL) assay and in situ identification of nuclear DNA fragmentation.. The ultrastructure injury of myocardium, myocardial infarct size, and plasma CK and LDH were reduced significantly with administration of U50488H before I/R, whereas the effects of U50488H were abolished by nor-BNI. DNA fragments were visualized by agarose electrophoresis, and clear DNA ladder formation was observed in myocardial tissue from hearts subjected to I/R. Administration of U50488H before ischemia exerted a significant anti-apoptotic effect as evidenced by markedly weaker DNA ladder formation. TUNEL staining showed U50488H treatment before I/R significantly reduced the percentage of apoptotic cells, which was blocked by 5-HD, a mitochondrial k(ATP) channel blocker. In accordance, U50488H treatment significantly inhibited I/R-induced elevated activities of caspase-3 and caspase-9. U50488H also produced an increase in Bcl-2 and a decrease in Bax protein expression in the I/R heart, and the anti-apoptotic effects of U50488H were all blocked by nor-BNI.. U50488H reduces myocardial necrosis and apoptosis after I/R and activation of kappa-opioid receptor may mediate a role in U50488H-induced myocardial protection.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Antihypertensive Agents; Apoptosis; bcl-2-Associated X Protein; Caspase 3; Caspase 9; DNA Fragmentation; Male; Microscopy, Electron, Transmission; Myocardial Infarction; Myocardium; Naltrexone; Narcotic Antagonists; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa; Reperfusion Injury

To determine whether U50488H, a selective agonist of kappa-opioid receptor, could induce biphasic (early and late) cardioprotection against myocardial ischemia/reperfusion injury and to explore the underlying mechanisms.. Isolated perfused rat hearts were subjected to 30 min of ischemia followed by 120 min reperfusion and the cardiac function was evaluated.. Left ventricular end-diastolic pressure (LVEDP), left ventricular developed pressure (LVDP) and maximal velocity of contraction and relaxation (+/-dP/dtmax) were improved when U50488H was administered 1 or 24 h before ischemia (P<0.05). Myocardial infarct size, activities of creatine kinase (CK) and lactate dehydrogenase (LDH) in the coronary effluent were lower in the U50488H pretreatment group than those in the control group. Administration of a selective cyclooxygenase-2 (COX-2) inhibitor, celecoxib abolished the late phase of cardioprotection produced by administration of U50488H 24 h before ischemia. Activities of CK and LDH in the coronary effluent were higher in U50488H and celecoxib co-pretreatment group than those in U50488H group. However, administration of celecoxib did not block the early phase of cardioprotection by 1 h treatment of U50488H before ischemia.. The late (but not the early) phase of cardioprotection induced by kappa-opioid receptor agonist might be mediated by COX-2.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Cardiotonic Agents; Creatine Kinase; Cyclooxygenase 2; In Vitro Techniques; Ischemic Preconditioning, Myocardial; L-Lactate Dehydrogenase; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa

Ischemic preconditioning fails to confer immediate cardioprotection in the absence of testosterone, indicating that the hormone is required for the process. Here we set out to determine whether testosterone is also necessary for delayed cardioprotection and, if so, how it acts. Male Sprague Dawley rats (7-8 wk) underwent sham operation or gonadectomy without (G) or with testosterone replacement (GT) for 8 wk. Isolated ventricular myocytes were preconditioned either by metabolic inhibition or with U50,488H, a kappa-opioid receptor agonist. In intact rats, U50,488H was administered systemically and 24 h later the hearts were removed. Ventricular myocytes were then subjected to metabolic inhibition and anoxia and isolated hearts to regional ischemia, followed by reperfusion to induce injury. Both types of preconditioning significantly increased the viability and decreased the lactate dehydrogenase release in ventricular myocytes from sham rats. They also activated heat shock transcription factor-1 and increased heat shock protein 70 expression. In contrast, all these effects were absent in myocytes from G rats and were restored by testosterone replacement. Parallel results were found in isolated hearts. In addition, preconditioning improved contractile functions impaired by ischemic insults in sham and rats gonadectomized with testosterone replacement but not G rats. The effects of testosterone replacement in ventricular myocytes were abolished by androgen receptor blockade. In conclusion, preconditioning requires testosterone to increase heat shock protein 70 synthesis, which mediates delayed cardioprotection in the male. These effects of testosterone are mediated by the androgen receptor.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Androgen Receptor Antagonists; Animals; Blotting, Western; Body Weight; Cell Hypoxia; Cell Survival; DNA-Binding Proteins; Heart Function Tests; Heat Shock Transcription Factors; HSP70 Heat-Shock Proteins; In Vitro Techniques; Ischemic Preconditioning, Myocardial; L-Lactate Dehydrogenase; Male; Myocardial Infarction; Myocardium; Myocytes, Cardiac; Orchiectomy; Organ Size; Rats; Rats, Sprague-Dawley; Testosterone; Transcription Factors

The involvement of opioid receptor activation during ischemia-reperfusion is somewhat controversial. While it is generally accepted that activation of the delta-opioid receptor (DOR) is cardioprotective, and may indeed be an important mediator of ischemic preconditioning, the role of the kappa-opioid receptor (KOR) is less well understood. To this end, we examined three different KOR agonists and their effects upon infarct size and arrhythmia development. Male Sprague-Dawley rats were subjected to 30 minutes of occlusion followed by 90 minutes of reperfusion. Opioid receptor agonists were administered 10 minutes before the onset of ischemia, while the opioid antagonists were given 20 minutes before occlusion. Untreated rats exhibited an infarct size (IS/AAR%) of 52.4 +/- 2.7%. Pretreatment with the DOR agonist, BW373U86, limited infarct development to 37.2 +/- 1.8%, which was reversed by the selective DOR antagonist, BNTX. All three KOR agonists studied, U50,488, ICI 204,448, and BRL 52537 significantly reduced infarct size to levels comparable to that of BW373U86. The infarct-sparing effects of U50,488 and ICI 204,448 were abolished by the selective KOR antagonist, nor-BNI. Nor-BNI failed to inhibit the cardioprotective effects of BRL 52537. Furthermore, U50,488 and BRL 52537, but not ICI 204,448, significantly reduced the incidence of arrhythmias. These effects were not blocked by nor-BNI. These data demonstrate that KOR activation provides a similar degree of infarct size reduction as DOR activation. KOR agonists also reduced arrhythmogenesis; however, these responses appear to be independent of KOR activation.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Antihypertensive Agents; Arrhythmias, Cardiac; Blood Pressure; Heart Rate; Male; Myocardial Infarction; Myocardial Reperfusion; Pyrrolidines; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa

For the first time the involvement of C-Reactive protein (CRP) in early (acute) and delayed ischemic (IPC) and pharmacological (chemical) preconditioning (CPC) in an in vivo model of rat myocardial infarction was presented. Acute IPC was produced by three 5 minute occlusion (ischemia) periods interspersed with 5 minute reperfusion, followed by 30 minute occlusion of the left coronary artery and 2 hour reperfusion injury. Acute CPC was produced by a k-opioid receptor agonist U50488H (5 mg/kg) applied i.v. 15 minutes before 30 minute ischemia/ 2 hour reperfusion. Delayed preconditioning was produced by 30 minute ischemia/ 2 hour reperfusion, induced 24 hour after either ischemic or pharmacological preconditioning. The myocardial ischemia/reperfusion injury was evaluated on the basis of total and cardiac creatine kinase isoenzyme activity, functional recovery of the heart (ECG), infarct size (% IS/RA) and mortality at the end of the experiments. The results obtained showed that: k-opioid receptor agonist U50488H mimics both the acute and delayed IPC in the above experimental protocol; Both acute IPC and most probably CPC act by opening of K(ATP) channels (the effects were blocked by nonspecific ATP-sensitive K channel blocker glybenclamide), and via activation of protein kinase C (a selective protein kinase C inhibitor chelerythrine blocked the efects); C-reactive protein (CRP) was significantly elevated by 54% in non-preconditioned acute ischemia/reperfusion injury. The elevation was more pronounced (82% increase) 24 hour after non-preconditioned ischemia/reperfusion injury. It reflected very well the increase in cardiac isoenzymes, infarct size and mortality of the rats, and can be used as a marker of the severity of myocardial injury in this model; The increase of CRP was prevented by both IPC and CPC in early, and especially in late preconditioning. This confirms the involvement of CRP as a marker in cardiac ischemic/reperfusion injury. It was concluded that in addition to the established involvement of adenosine, bradykinin, opioid and other receptors, a suppression of myocardial CRP/complement production might be involved in the biological mechanism of preconditioning. This could be a promising perspective in clinical interventions against ischemia/reperfusion injuries of the heart.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; C-Reactive Protein; Disease Models, Animal; Ischemic Preconditioning, Myocardial; Male; Myocardial Infarction; Potassium Channels; Rats; Rats, Wistar; Reperfusion Injury

The effect of preconditioning with U50488 H (UP), a selective kappa-opioid receptor (kappa-OR) agonist, on infarct size and intracellular Ca2+ ([Ca2+]i) in the heart subjected to ischaemic insults were studied and evaluated. U50488 H administered intravenously reduced the infarct size 18-48 h after administration in isolated hearts subjected to regional ischaemia/reperfusion (I/R). The effect was dose dependent. A peak effect was reached at 10 mg x kg-1 U50488 H and at 24 h after administration. The effect of 10 mg x kg-1 U50488 H at 24 h after administration was abolished by nor-binaltorphimine (nor-BNI), a selective kappa-OR antagonist, indicating the effect was kappa-OR mediated. The infarct reducing effect of U50488 H was attenuated when a selective blocker of mitochondrial (5-hydroxydecanoic acid, 5-HD) or sarcolemmal (HRM-1098) ATP-sensitive potassium channel (KATP) was coadministered with U50488 H 24 h before ischaemia or when 5-HD was administered just before ischaemia. U50488 H also attenuated the elevation in [Ca2+]i and reduction in electrically induced [Ca2+]i transient in cardiomyocytes subjected to ischaemic insults. The effects were reversed by blockade of KATP channel, which abolished the protective effect of preconditioning with U50488 H. The results indicated that mitochondrial KATP channel serves as both a trigger and a mediator, while sarcolemmal KATP channel as a trigger only, of delayed cardioprotection of kappa-OR stimulation. The effects of these channels may result from prevention/attenuation of [Ca2+]i overload induced by ischaemic insults.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Calcium; Calcium Signaling; Coronary Circulation; Coronary Vessels; Dose-Response Relationship, Drug; Heart; Heart Rate; Injections, Intravenous; Ischemic Preconditioning, Myocardial; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Myocytes, Cardiac; Naltrexone; Potassium Channel Blockers; Potassium Channels; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa; Time Factors

There is controversy in the literature regarding the involvement of opioid delta (DOP, OP1)- and kappa (KOP,OP2)-receptors in ischemic preconditioning (IPC). Previous studies on this subject in our laboratories and elsewhere have been performed on either isolated heart muscles of experimental animals, or in open-heart surgery rats. To highlight this problem, we introduced an in vivo model of myocardial infarction in rats, which not only allowed electrocardiographic and enzymatic evaluation, but also morphometric assessment of myocardial infarction. In addition to these parameters, a direct receptor ligand study was undertaken, using [3H]-DPDPE, a specific opioid delta-receptor ligand. In our pharmacodynamic studies, we used the selective opioid delta-receptor agonist D-Ala2,D-Leu5 enkephalin (DADLE) and antagonist natrindole. For the evaluation of opioid kappa-receptors, the selective opioid agonist U-50488H and antagonist nor-BNI were employed. Ischemic preconditioning showed the best beneficial effect, compared with pharmacological stimulation of either opioid delta- or kappa-receptors. In normal rat myocytes, two types of opioid delta-receptors exist, namely, low-affinity and high-affinity opioid receptors. In acute myocardial infarction (30-min ischemia), the low-affinity type opioid receptors disappeared, most likely as a result of receptor downregulation due to an excessive release of enkephalins. There was no change in the density of the high-affinity opioid receptor type, but their affinity significantly increased (p < 0.05) by 58%. The radioligand receptor studies showed that opioid delta 1-receptor type was involved not only in triggering, but also in maintaining, the preconditioned state. On the basis of our pharmacodynamic studies, we suggest that both opioid delta 1- and kappa-receptors are involved in the phenomenon of IPC, but with different effects. After 30 min of left coronary artery occlusion, opioid delta-receptor agonist DADLE decreased the infarct size/area at risk from 59.80% in control, untreated, infarcted rats, to 20.40% in treated rats, without a significant effect (p > 0.05) on the occurrence of early cardiac arrhythmias. Opioid kappa-receptor agonist U-50488H produced an opposite effect on the myocardium. It decreased the infarct size/area at risk by 44%, decreased occurrence of early arrhythmias by 77% and also decreased ventricular ectopic beats by 80%. The opioid delta- and kappa-receptor agonists used in this study signifi

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Analysis of Variance; Animals; Disease Models, Animal; Hemodynamics; In Vitro Techniques; Ischemic Preconditioning, Myocardial; Male; Myocardial Infarction; Myocardium; Radioligand Assay; Rats; Rats, Wistar; Receptors, Opioid, delta; Receptors, Opioid, kappa

Two series of experiments were performed in the isolated perfused rat heart to determine the role of kappa- and delta-opioid receptors (OR) in cardioprotection of ischemic preconditioning (IP). In the first series of experiments, it was found that IP with two cycles of 5-min regional ischemia followed by 5-min reperfusion each reduced infarct size induced by 30-min ischemia, and the ameliorating effect of IP on infarct was attenuated with blockade of either 5 x 10(-6) mol/l nor-binaltorphimine (nor-BNI), a selective kappa-OR antagonist, or 5 x 10(-6) mol/l naltrindole (NTD), a selective delta-OR antagonist. The second series showed that U50,488H, a selective kappa-OR agonist, or D-Ala(2)-D-leu(5)-enkephalin (DADLE), a selective delta-OR agonist, dose dependently reduced the infarct size induced by ischemia, which mimicked the effects of IP. The effect of 10(-5) mol/l U50,488H on infarct was significantly attenuated by blockade of protein kinase C (PKC) with specific PKC inhibitors, 5 x 10(-6) mol/l chelerythrine or 8 x 10(-7) mol/l calphostin C, as well as by blockade of ATP-sensitive K(+) (K(ATP)) channels with blockers of the channel, 10(-5) mol/l glibenclamide or 10(-4) mol/l 5-hydroxydecanoate. IP also reduced arrhythmia induced by ischemia. Nor-BNI, but not NTD, attenuated, while U50,488H, but not DADLE, mimicked the antiarrhythmic action of IP. In conclusion, the present study has provided first evidence that kappa-OR mediates the ameliorating effects of IP on infarct and arrhythmia induced by ischemia, whereas delta-OR mediates the effects only on infarct. Both PKC and K(ATP) channels mediate the effect of activation of kappa-OR on infarct.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Arrhythmias, Cardiac; Coronary Circulation; Enkephalin, Leucine-2-Alanine; Enzyme Inhibitors; Hemodynamics; In Vitro Techniques; Ischemic Preconditioning, Myocardial; Male; Myocardial Infarction; Potassium Channel Blockers; Protein Kinase C; Rats; Rats, Sprague-Dawley; Receptors, Opioid, delta; Receptors, Opioid, kappa; Signal Transduction