Morphine-induced Cardioprotection Research Articles

Effects of nalbuphine on the cardioprotective effect of morphine in rats

BackgroundWhen combined with morphine, nalbuphine not only does not affect the analgesic effect but also prevents opioid-induced side effects. The authors investigated whether nalbuphine interferes with morphine-induced cardioprotection in rats. MethodsAnesthetized male Sprague-Dawley rats were randomly assigned to 1 of 4 treatment groups. Nalbuphine (NAL, 1 mg/kg) and morphine (MOR, 0.3 mg/kg) were administered 10 and 5 min prior to myocardial ischemia, respectively. Additionally, the NAL + MOR group received the combination of NAL and MOR prior to myocardial ischemia. An in vivo animal model was established by occluding the left anterior descending artery for 30 min and reperfusing it for 2 h. After 2 h of reperfusion, the infarcted area of heart was measured by Evans blue/triphenyl tetrazolium staining, and the levels of creatine kinase isoenzymes (CK-MB) in serum were detected by enzyme-linked immunosorbent assay. ResultsNalbuphine had no protective effect against the infarct area compared with the control treatment (NAL, 52.5 ± 5% versus CON, 52.6 ± 4%; *P < 0.01), and the infarct size-sparing effects of morphine were not affected by nalbuphine (NAL + MOR, 42.6 ± 7% versus MOR, 40.4 ± 3%; P > 0.05). The nalbuphine group did not show a change the levels of serum CK-MB compared with the control group, and nalbuphine did not affect the levels of serum CK-MB in the MOR group. ConclusionsNalbuphine does not interfere with the cardioprotective effect of morphine in vivo. Therefore, nalbuphine could be safely used or combined with morphine in patients with acute myocardial infarction.

Spinal NGF induces anti-intrathecal opioid-initiated cardioprotective effect via regulation of TRPV1 expression

Evidences from previous studies confirmed that intrathecal morphine preconditioning (ITMP) reduces the cardiac injury of ischemia-reperfusion (IR) via the central nervous system. However, the molecular mechanism is not fully understood. The breath of central nerve growth factor (NGF) during nociceptive transmission has been well documented, and little is known about the significance of NGF in myocardial injury of IR and intrathecal morphine-induced cardioprotection. To address these questions, we over-expressed or silenced NGF in the spinal cord by using intrathecal injection of lentivirus-NGF or shRNA respectively, accompanied by ITMP in the IR rat model. The levels of NGF and tropomyosin receptor kinase A (Trka) as well as transient receptor potential vanilloid 1 (TRPV1) in the T2–6 spinal cord were evaluated. The results showed that cardiac damage indicators induced by IR, including the increased infarct size, arrhythmia score and serum troponin levels were attenuated after ITMP. However, overexpression of spinal NGF significantly reversed these decreases, as well as reduced the expression and phosphorylation of TRPV1 that was elicited by ITMP. Conversely, silencing of spinal NGF enhanced ITMP-induced cardioprotective effects. Phosphorylation and expression of TRPV1 in the spinal cord were significantly decreased after regional NGF silencing. These findings suggested that the cardioprotective effects of ITMP may implement by mediating through spinal NGF expression, wherein it involves the nociceptor TRPV1. NGF may act as a potential therapeutic target in the development of new agents for the treatment of cardiac injury induced by IR.

Transient receptor potential vanilloid 1 inhibitors block laparotomy- and opioid-induced infarct size reduction in rats.

In light of the opioid epidemic, physicians are increasingly prescribing non-opioid analgesics to surgical patients. Transient receptor potential vanilloid 1 (TRPV1) inhibitors are potentially alternative pain therapeutics for surgery. Here, we examined in rodents whether the cardioprotection conferred by two common procedures during surgery, a laparotomy or morphine delivery, is mediated by the TRPV1 channel. We further tested whether an experimental analgesic peptide (known as P5) targeted against the TRPV1 C-terminus region interferes with laparotomy- or morphine-induced cardioprotection. Male Sprague-Dawley rats were subjected to 30min coronary occlusion followed by 120min reperfusion. Before ischaemia, a laparotomy with or without capsaicin application (0.1% cream, a TRPV1 activator) was performed. Additional rats were given morphine (0.3mg·kg-1 ) with or without capsaicin. In addition, capsazepine (3mg·kg-1 , a classical TRPV1 inhibitor), or P5 (3mg·kg-1 , a peptide analgesic and TRPV1 inhibitor), was given either alone or prior to a laparotomy or morphine administration. Myocardial infarct size was determined. A laparotomy, in addition to combining a laparotomy with capsaicin cream, reduced infarct size versus control. Morphine, in addition to combining morphine administration with capsaicin cream, also reduced infarct size versus control. When TRPV1 inhibitors capsazepine or P5 were given, either TRPV1 inhibitor abolished the infarct size reduction mediated by a laparotomy or morphine. Inhibiting the TRPV1 channel blocks laparotomy- or morphine-induced cardioprotection. Impaired organ protection may be a potential pitfall of using TRPV1 inhibitors for pain control.

Transient Receptor Potential Vanilloid 1 Mediates Laporotomy and Opioid-induced Infarct Size Reduction in Rats

Background: In light of an opioid epidemic in the United States, physicians are increasingly prescribing non-opioid analgesics. However, newer pain medications such as transient receptor potential vanilloid 1 (TRPV1) inhibitors may impede organ protection. We examined in rodents whether a laparotomy- and morphine-induced infarct size reduction is mediated by TRPV1. Further, we tested if inhibiting TRPV1 interferes with laparotomy- or morphine-induced cardioprotection. Methods: Male Sprague-Dawley rats were subjected to 30 minutes of coronary occlusion followed by 120 minutes reperfusion. Before ischaemia, a laparotomy with or without application of capsaicin cream (0.1%, a TRPV1 activator), was performed. Additional sets of rats were given morphine (3 mg/kg) with or without capsaicin. Further, capsazepine (3 mg/kg, a classical TRPV1 inhibitor), and P5 (3 mg/kg, a peptide analgesic and TRPV1 inhibitor), were given prior to the laparotomy or morphine. Myocardial infarct size was determined. Results: A laparotomy or capsaicin reduced infarct size versus controls (to 44±2%* or 49±1%* vs. 66±1%, infarct size/area at risk %, n=6, *P<0.0001). When combining laparotomy and capsaicin, cardioprotection was not additive. Morphine also reduced infarct size (37±3%* vs. 61±2%, n=6, P<0.0001), with no additive effect when combined with capsaicin. The TRPV1 inhibitors capsazepine and P5 abolished cardioprotection induced by either a laparotomy (58±1%+ and 65±2%+ vs. 44±2%+, respectively) or morphine (62±3%+ and 58±2%+ vs. 37±3%, respectively, n=6, +P<0.0001). Conclusions: Infarct size reduction by a laparotomy or morphine is blocked by inhibiting TRPV1. Drugs given which inhibit TRPV1 may impede organ protection and this requires further study.

Spinal Neuronal NOS Signaling Contributes to Morphine Cardioprotection in Ischemia Reperfusion Injury in Rats.

Morphine has been widely used as rescue treatment for heart attack and failure in humans for many decades. Relatively little has been known about the role of spinal opioid receptors in morphine cardioprotection. Recent studies have shown that intrathecal injection of morphine can reduce the heart injury caused by ischemia (I)/reperfusion (R) in rats. However, the molecular and cellular mechanisms underlying intrathecal morphine cardioprotection has not been determined. Here, we report that intrathecal morphine postconditioning (IMPOC) rescued mean artery pressure (MAP) and reduced myocardial injury in I/R. Pretreatment with either naloxone (NAL), a selective mu-opioid receptor antagonist, or nitric oxide synthase (NOS) inhibitors via intrathecal delivery completely abolished IMPOC cardioprotection, suggesting that the spinal mu-opioid receptor and its downstream NOS signaling pathway are involved in the mechanism of the morphine-induced effect. Consistent with this, IMPOC significantly enhanced spinal neural NOS phosphorylation, nitric oxide, and cGMP content in a similar time course. Intrathecal application of 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, a specific inhibitor of guanylate cyclase, completely ablated IMPOC-induced enhancement of cardioprotection and spinal cGMP content. IMPOC rescue of MAP and ischemic injury is correlated with IMPOC enhancement of NOS signaling. Collectively, these findings strengthen the concept of spinal mu-opioid receptors as a therapeutic target that mediates morphine-induced cardioprotection. We also provide evidence suggesting that the activation of spinal NOS signaling is essential for morphine cardioprotection.

Morphine-Induced Preconditioning: Involvement of Protein Kinase A and Mitochondrial Permeability Transition Pore.

BackgroundMorphine induces myocardial preconditioning (M-PC) via activation of mitochondrial large conductance Ca2+-sensitive potassium (mKCa) channels. An upstream regulator of mKCa channels is protein kinase A (PKA). Furthermore, mKCa channel activation regulates mitochondrial bioenergetics and thereby prevents opening of the mitochondrial permeability transition pore (mPTP). Here, we investigated in the rat heart in vivo whether 1) M-PC is mediated by activation of PKA, and 2) pharmacological opening of the mPTP abolishes the cardioprotective effect of M-PC and 3) M-PC is critically dependent on STAT3 activation, which is located upstream of mPTP within the signalling pathway.MethodsMale Wistar rats were randomised to six groups (each n = 6). All animals underwent 25 minutes of regional myocardial ischemia and 120 minutes of reperfusion. Control animals (Con) were not further treated. Morphine preconditioning was initiated by intravenous administration of 0.3 mg/kg morphine (M-PC). The PKA blocker H-89 (10 μg/kg) was investigated with and without morphine (H-89+M-PC, H-89). We determined the effect of mPTP opening with atractyloside (5 mg/kg) with and without morphine (Atr+M-PC, Atr). Furthermore, the effect of morphine on PKA activity was tested in isolated adult rat cardiomyocytes. In further experiments in isolated hearts we tested the protective properties of morphine in the presence of STAT3 inhibition, and whether pharmacological prevention of the mPTP-opening by cyclosporine A (CsA) is cardioprotective in the presence of STAT3 inhibition.ResultsMorphine reduced infarct size from 64±5% to 39±9% (P<0.05 vs. Con). H-89 completely blocked preconditioning by morphine (64±9%; P<0.05 vs. M-PC), but H-89 itself had not effect on infarct size (61±10%; P>0.05 vs. Con). Also, atractyloside abolished infarct size reduction of morphine completely (65±9%; P<0.05 vs. M-PC) but had no influence on infarct size itself (64±5%; P>0.05 vs. Con). In isolated hearts STAT3 inhibitor Stattic completely abolished morphine-induced preconditioning. Administration of Stattic and mPTP inhibitor cyclosporine A reduced infarct size to 31±6% (Stat+CsA, P<0.05 vs. Con). Cyclosporine A alone reduced infarct size to 26±7% (CsA P<0.05 vs. Con). In cardiomyocytes, PKA activity was increased by morphine.ConclusionOur data suggest that morphine-induced cardioprotection is mediated by STAT3-activation and inhibition of mPTP, with STA3 located upstream of mPTP. There is some evidence that protein kinase A is involved within the signalling pathway.

MicroRNA-133b-5p Is Involved in Cardioprotection of Morphine Preconditioning in Rat Cardiomyocytes by Targeting Fas

BackgroundMicroRNAs (miRNAs) have been implicated in ischemia-reperfusion injury and ischemic preconditioning. Opioid pre- and postconditioning have powerful protective effects on the heart, but it is still not known whether miRNAs are involved in opioid-induced cardioprotection. The present study was designed to investigate the role of miRNAs in morphine preconditioning (MPC)-induced cardioprotection. MethodsMiRNA microarray analysis was performed to examine the differentially expressed miRNAs caused by MPC in adult rat cardiomyocytes. A dual-luciferase reporter assay was performed to confirm the direct regulation of miR-133b-5p on the target gene Fas. MiR-133b-5p mimic or inhibitor was separately transfected into myocardial H9c2 cells to examine the role of miR-133b-5p in morphine-induced cardioprotection. ResultsMPC protected adult rat cardiomyocytes against hypoxia/reoxygenation (H/R) injury by reducing cell injury and death. MiRNA microarray data showed that a total of 39 miRNAs were differentially expressed after MPC treatment. A Dual-luciferase reporter assay confirmed that miR-133b-5p directly targets the Fas gene. After H/R injury, the decrease in miR-133b-5p and a contemporaneous rise in Fas mRNA and protein levels in adult rat cardiomyocytes were prevented by MPC treatment. In H9c2 cardiomyocytes, overexpression of miR-133b-5p reduced H/R-induced cell injury and apoptosis by inhibiting Fas expression. Knockdown of miR-133b-5p blocked morphine-mediated cardioprotection by reducing miR-133b-5p levels while enhancing the expression of Fas mRNA and protein. ConclusionsMPC causes a change in miRNA expression in rat cardiomyocytes. Morphine may protect cardiomyocytes against H/R injury through upregulation of miR-133b-5p by targeting Fas.

Morphine Reduces Myocardial Infarct Size via Heat Shock Protein 90 in Rodents.

Opioids reduce injury from myocardial ischemia-reperfusion in humans. In experimental models, this mechanism involves GSK3β inhibition. HSP90 regulates mitochondrial protein import, with GSK3β inhibition increasing HSP90 mitochondrial content. Therefore, we determined whether morphine-induced cardioprotection is mediated by HSP90 and if the protective effect is downstream of GSK3β inhibition. Male Sprague-Dawley rats, aged 8–10 weeks, were subjected to an in vivo myocardial ischemia-reperfusion injury protocol involving 30 minutes of ischemia followed by 2 hours of reperfusion. Hemodynamics were continually monitored and myocardial infarct size determined. Rats received morphine (0.3 mg/kg), the GSK3β inhibitor, SB216763 (0.6 mg/kg), or saline, 10 minutes prior to ischemia. Some rats received selective HSP90 inhibitors, radicicol (0.3 mg/kg), or deoxyspergualin (DSG, 0.6 mg/kg) alone or 5 minutes prior to morphine or SB216763. Morphine reduced myocardial infarct size when compared to control (42 ± 2% versus 60 ± 1%). This protection was abolished by prior treatment of radicicol or DSG (59 ± 1%, 56 ± 2%). GSK3β inhibition also reduced myocardial infarct size (41 ± 2%) with HSP90 inhibition by radicicol or DSG partially inhibiting SB216763-induced infarct size reduction (54 ± 3%, 47 ± 1%, resp.). These data suggest that opioid-induced cardioprotection is mediated by HSP90. Part of this protection afforded by HSP90 is downstream of GSK3β, potentially via the HSP-TOM mitochondrial import pathway.

Abstract 340: Heat Shock Proteins HSP90 and HSP70 Mediate Opioid- and GSK3β-induced Cardioprotection

Previously, opioids were established to reduce myocardial injury in an ischemic preconditioning (IPC)-like manner, involving a central and downstream role of glycogen synthase kinase-3 beta (GSK-3β) inhibition. However, the mechanism of GSK-3β inhibition mediating cardioprotection and the protein partners involved has not been fully elucidated. Hence, we used a non-biased sequence scan of the proteome to determine potential GSK-3β protein partners and tested whether two candidate proteins, heat shock proteins (HSP) 70 and 90, are involved in the mechanism of opioid-induced cardioprotection. A non-biased BLAST search was performed for putative GSK-3β target substrates, based upon the sequence motif S/T-X-X-X-S/T. Approximately 700 proteins were identified to have this moiety, including many of the HSP protein class, including HSP70 and HSP90. To determine whether HSP70 or HSP90 are indeed important in opioid-induced cardioprotection, rats were subjected to an in vivo myocardial ischemia-reperfusion protocol consisting of 30 minutes of ischemia and 2 hours of reperfusion of the left anterior descending coronary artery followed by infarct size assessment. Either morphine (0.3mg/kg) or inhibition of GSK-3β using SB216763 (0.6mg/kg), reduced infarct size compared to control (42.21±1*% and 41.09±2*%, respectively versus control 60.38±1.2, *P&lt;0.01). Inhibition of HSP70 using desoxysperguanalin (DSG), or HSP90 using radicicol (RAD), abrogated morphine-induced protection (56.09±2 and 58.64±1, respectively). Either DSG or RAD partially inhibited protection in the presence of GSK-3β (47.28±1.071 and 49.88±3.09). Our results suggest that morphine-induced cardioprotection occurs by a HSP70 and HSP90- dependent mechanism, with this HSP machinery partially required for GSK3β-inhibition-induced cardioprotection. Further understanding of this mechanism is important, considering many agents targeting HSP are currently in development as novel cancer treatments, which may have detrimental effects on the myocardial salvage mediated by opioids or by GSK3β-inhibition.

Involvement of Adenosine Monophosphate-Activated Protein Kinase in Morphine-induced Cardioprotection

Adenosine monophosphate-activated protein kinase (AMPK) orchestrates the regulation of energy-generating and -consuming pathways, and protects the heart against ischemic injury and apoptosis. Recent progress shed light on various factors, including adiponectin, MIF, H11K, and metformin in the activation of AMPK. It is uncertain whether the activation of AMPK is contributed to cardioprotection of opioids. Here we show that morphine, an exogenous non-peptide opioid receptor agonist, can modulate the activation of the cardioprotective AMPK pathway during ischemia and exert anti-apoptotic effects through AMPK. Isolated rat hearts were perfused on a constant pressure Langendorff system and subjected to 30 min of global ischemia followed by 60 min of reperfusion. The hearts received vehicles, morphine, a combination of morphine and compound C, a combination of morphine and STO609, a combination of morphine and BAPTA-AM at the onset of ischemia. Hemodynamics parameters, infarct size, release of intracellular creatine kinase, expression of AMPK, and terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling staining were analyzed. Morphine significantly increased phosphorylation level of Thr172 site on AMPK, left ventricular function, and reduced infarct size as a percentage of the area at risk (IS/AAR from 63% ± 7% to 40% ± 5%), release of intracellular creatine kinase (from 319 ± 46 to 156 ± 42IU/60 min/gdw), apoptosis ratio (from 16% ± 2% to 5% ± 1.4%) during reperfusion in comparison with the control group. A inhibitor of AMPK, compound C abrogated phosphorylation of AMPK induced by morphine, the improvement in myocardial function, and the reduction of IS/AAR (58% ± 6%), release of intracellular creatine kinase (270 ± 40IU/60 min/gdw), apoptosis ratio (13% ± 1.5%). A Ca(2+)/calmodulin-dependent protein kinase kinase inhibitor STO609 and a chelator of intracellular Ca(2+) stores BAPTA-AM also abolished the cardioprotection of morphine. Morphine can ameliorate myocardial contractile dysfunction and limit infarct size following ischemia and reperfusion by a mechanism involving activation of AMPK, and activate AMPK by Ca(2+)-CaMKKβ-dependent phosphorylation.

Diabetes Abolishes Morphine-Induced Cardioprotection via Multiple Pathways Upstream of Glycogen Synthase Kinase-3β

The cardioprotective effect of opioids or glycogen synthase kinase (GSK) inhibitors given at reperfusion has not been investigated in diabetes models. Therefore, nondiabetic (NDBR) or streptozotocin-induced diabetic (DBR) rat hearts were subjected to 30 min of ischemia and 2 h of reperfusion. Groups of NDBR or DBR were administered either vehicle, morphine (0.3 mg/kg), or the GSK inhibitor SB216763 (0.6 mg/kg) 5 min before reperfusion. SB216763 (but not morphine) reduced infarct size in DBRs (44 +/- 1* and 55 +/- 2%, respectively), while both agents reduced infarct size in NDBRs versus untreated NDBRs or DBRs (44 +/- 3*, 42 +/- 3*, 60 +/- 2, and 56 +/- 2%, respectively, *P < 0.001). Morphine-induced phospho- (P-)GSK3beta was reduced 5 min after reperfusion in DBRs compared with NDBRs (0.83 +/- 0.29 and 1.94 +/- 0.12 [P < 0.05] pg/microg tissue, respectively). The GSK3beta mediators, P-Akt, P-extracellular signal-related kinase (ERK)1, and P-signal transducer and activator of transcription (STAT)3, were also significantly reduced in untreated DBR compared with NDBR rats. Morphine-induced elevations of P-Akt, P-ERK1, P-p70s6, P-janus-activated kinase-2, and P-STAT3 in NDBRs were also blunted in DBRs. H9C2 cells raised in 25 mmol/l compared with 5.56 mmol/l glucose media also demonstrated reduced morphine-induced P-GSK3beta, P-Akt, P-STAT3, and P-ERK1 after 15 min. Hence, acute GSK inhibition may provide a novel therapeutic strategy for diabetic patients during an acute myocardial infarction, whereas morphine is less effective due to signaling events that adversely affect GSK3beta.

COX-2 mediates morphine-induced delayed cardioprotection via an iNOS-dependent mechanism

Cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) have been shown to be mediators of cardioprotection induced by ischemic preconditioning and opioids. However, it is not known whether COX-2 is involved in morphine-induced cardioprotection accompanied with iNOS. Therefore, we investigated the role of COX-2 in morphine-induced cardioprotection and the effect of iNOS on COX-2. Myocardial ischemia was induced by a 45-min coronary artery occlusion in mice. Infarct size (IS) as a percentage of the area at risk (AAR) was determined by triphenyltetrazolium chloride staining. The COX-2-selective inhibitor NS-398 was used to investigate the role of COX-2. Expression of COX-2 was assessed by Western blotting, and the myocardial prostaglandin (PG)E 2 and 6-keto-PGF 1α contents were measured using enzyme immunoassays. The iNOS-selective inhibitor SMT and iNOS gene-knockout mice were used to investigate the effect of iNOS on COX-2. IS/AAR was reduced significantly 1 and 24 h after morphine preconditioning. The infarct-sparing effect 24 h after morphine administration, but not the cardioprotection 1 h later, was completely abolished by NS-398. Marked enhancement of myocardial COX-2 expression was measured 24 h after morphine preconditioning associated with up-regulation of myocardial contents of PGE 2 and 6-keto-PGF 1α. Neither the level of COX-2 nor the contents of PGE 2 and 6-keto-PGF 1α were enhanced 1 h later. Administration of SMT and targeted abrogation of iNOS gene blocked the enhancement of myocardial PGE 2 and 6-keto-PGF 1α 24 h after morphine administration but did not inhibit the up-regulation of COX-2 expression. We concluded that COX-2 mediates morphine-induced delayed cardioprotection via an iNOS-dependent pathway.

Acute aspirin treatment abolishes, whereas acute ibuprofen treatment enhances morphine-induced cardioprotection: role of 12-lipoxygenase.

Patients suffering an acute myocardial infarction routinely receive morphine and nonsteroidal anti-inflammatory drugs (NSAIDs) alone or in combination. However, the importance of the dose, timing, or the combined administration of both on infarct size reduction has not been assessed. Additionally, it is not known whether morphine or NSAIDs require 12-lipoxygenase (12-LO) to mediate infarct size reduction as found previously for ischemic preconditioning. Male Sprague-Dawley rats were subjected to 30 min of ischemia and 2 h of reperfusion, followed by infarct size assessment (mean +/- S.E.M.%, **P < 0.01). Morphine (0.3 mg/kg), ibuprofen (3 mg/kg), but not aspirin (3 mg/kg) reduced infarct size when administered 5 min before reperfusion compared with vehicle (42.3 +/- 1.5**, 40.8 +/- 2.8**, 60.7 +/- 2.3 versus 59.1 +/- 1.7%, respectively); however, none of these agents reduced infarct size when administered 10 s after reperfusion. Ibuprofen (3 mg/kg) administered with morphine (0.3 mg/kg) reduced infarct size (43.7 +/- 1.3%**), whereas aspirin (1 and 3 mg/kg) abolished morphine-induced infarct size reduction. Morphine (0.2 mg/kg) and ibuprofen (0.6 mg/kg) given at doses not effective individually reduced infarct size when given together (59.0 +/- 1.4, 57.6 +/- 2.8, and 43.9 +/- 1.6%**, respectively). Morphine- and ibuprofen-induced infarct size reduction was abolished by the 12-LO inhibitor baicalein (3 mg/kg) and mimicked by the 12-LO metabolite 12-(S)-hydroxyeicosa-5Z,8Z,10Z,14Z-tetraenoic acid (45.2 +/- 2.5%**). These data suggest that morphine and ibuprofen reduce infarct size individually or at subthreshold doses in combination by 12-LO when administered 5 min before reperfusion. Furthermore, acute aspirin administration has a detrimental interaction with morphine that abrogates morphine-induced infarct size reduction.

Pretreatment with delta-opioid receptor blockade attenuates the reduction of apoptosis associated with ischemic preconditioning or morphine-induced cardioprotection in rabbit ischemia/reperfusion model Shinji Okubo, Yujirou Tanabe, Nakaba Fujioka, Noboru Takehoski, Rakesh C. Kukreja. Kanzawa Medical University, Uchinada, Japan

Pretreatment with delta-opioid receptor blockade attenuates the reduction of apoptosis associated with ischemic preconditioning or morphine-induced cardioprotection in rabbit ischemia/reperfusion model Shinji Okubo, Yujirou Tanabe, Nakaba Fujioka, Noboru Takehoski, Rakesh C. Kukreja. Kanzawa Medical University, Uchinada, Japan

Tumour necrosis factor-α induces cyclo-oxygenase-2 expression in first trimester trophoblasts: Suppression by glucocorticoids and NSAIDs : Imseis H.M., Zimmerman P.D., Samuels P., Kniss D.A. Department of Obstetrics and Gynecology, Laboratory of Perinatal Research, Ohio State University College Medicine, Columbus, OH, USA [Placenta 18(7), 521–526; 1997

Cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) have been shown to be mediators of cardioprotection induced by ischemic preconditioning and opioids. However, it is not known whether COX-2 is involved in morphine-induced cardioprotection accompanied with iNOS. Therefore, we investigated the role of COX-2 in morphine-induced cardioprotection and the effect of iNOS on COX-2. Myocardial ischemia was induced by a 45-min coronary artery occlusion in mice. Infarct size (IS) as a percentage of the area at risk (AAR) was determined by triphenyltetrazolium chloride staining. The COX-2-selective inhibitor NS-398 was used to investigate the role of COX-2. Expression of COX-2 was assessed by Western blotting, and the myocardial prostaglandin (PG)E2 and 6-keto-PGF1α contents were measured using enzyme immunoassays. The iNOS-selective inhibitor SMT and iNOS gene-knockout mice were used to investigate the effect of iNOS on COX-2. IS/AAR was reduced significantly 1 and 24 h after morphine preconditioning. The infarct-sparing effect 24 h after morphine administration, but not the cardioprotection 1 h later, was completely abolished by NS-398. Marked enhancement of myocardial COX-2 expression was measured 24 h after morphine preconditioning associated with up-regulation of myocardial contents of PGE2 and 6-keto-PGF1α. Neither the level of COX-2 nor the contents of PGE2 and 6-keto-PGF1α were enhanced 1 h later. Administration of SMT and targeted abrogation of iNOS gene blocked the enhancement of myocardial PGE2 and 6-keto-PGF1α 24 h after morphine administration but did not inhibit the up-regulation of COX-2 expression. We concluded that COX-2 mediates morphine-induced delayed cardioprotection via an iNOS-dependent pathway.

Ischemic Preconditioning and Morphine-induced Cardioprotection Involve the Delta (δ)-opioid Receptor in the Intact Rat Heart

Several investigators have demonstrated that the opioid pathway is involved in tissue preservation during hypoxia or ischemia and that this protection is mediated via the delta (δ)-opioid receptor. Subsequently, we have shown that opioid receptors are involved in ischemic preconditioning (PC) in the rat heart and that morphine produces a cardioprotective effect; however, the class of opioid receptors involved in mediating these effects is still unknown. Therefore, the purpose of the present study was to test the hypothesis that ischemia- and morphine-induced cardioprotection are mediated via stimulation of theδ-opioid receptor in the rat heart. Anesthetized, open-chest Wistar rats were subjected to one of six protocols. The control group was subjected to 30 min of occlusion and 2 h of reperfusion. Ischemic PC was elicited by three 5 min occlusion periods interspersed with 5 min of reperfusion. Morphine-induced cardioprotection was produced by three 5 min morphine infusions (100μg/kg/infusion, i.v.) interspersed with a 5-min drug-free period. To determine if theδ-opioid receptor has a role in ischemic PC and morphine-induced cardioprotection, naltrindole (NTI), a selectiveδ-opioid receptor antagonist, was utilized. NTI (5 mg/kg, i.v.) was given 10 min prior to ischemic PC (NTI+PC) or morphine infusion (NTI+MOR). Also, NTI (5 mg/kg, i.v.) was given 10 min before the 30 min occlusion period in untreated rats. Infarct size (IS) as a percent of the area at risk (AAR) was determined by 2,3,5-triphenyltetrazolium chloride staining. Ischemic PC and morphine infusions resulted in similar reductions in IS/AAR from 51±4 to 11±3 and 15±4% (*P<0.05), respectively. NTI completely abolished the cardioprotective effect induced by ischemia and morphine. The results of the present study suggests a role ofδ;-opioid receptors in ischemic PC or morphine-induced myocardial protection in the rat.