The role of adrenergic and muscarinic receptors in stress-induced cardiac injury.

Takotsubo syndrome (TS) is a rare but dangerous disease that can be fatal. The pathogenesis of TS is not well understood because there is no animal model of TS that fully mimics TS. It has now been documented that stress exposure (24 h) of rats induced the state which is similar TS in human: contracture damage of myofibrils, elevation of the serum creatine kinase MB level, increased 99mTc-pyrophosphate (99mTc-PYP) accumulation in the heart, QTc interval prolongation, and contractility dysfunction of the heart. Immobilization stress resulted in an increase in coronary blood flow. Emotional stress increased the serum catecholamine level. Blockade of ß1-adrenergic receptor (AR) prevented stress-induced cardiac injury (SICI). Blockade of ß2-AR aggravated stress-induced cardiac injury. Stimulation of ß2-AR increased cardiac tolerance to stress. Inhibition of ß3-AR, α1-AR had no effect on SICI. Blockade of peripheral muscarinic receptors or α2-AR aggravated SICI. Pretreatment with the selective ß1-AR antagonist atenolol attenuates stress-induced cardiac contractility dysfunction, but recovery of cardiac contractility is not complete. There is indirect evidence that circulating catecholamines play an important role in SICI. Consequently, the activation of ß1-AR plays a significant role in SICI. However, there are other receptors which are also involved in SICI and require further investigation.

Impact of cold adaptation on cardiac tolerance to ischemia/reperfusion and glucocorticoid, thyroid hormone levels.

We have established that the continuous cold exposure (CCE, 4°C, 4 weeks) causes cold adaptation, increases systolic blood pressure, exerts infarct-limiting effect during coronary artery occlusion (45 min) and reperfusion (2 h). The CCE increases adrenal weight, heart weight and triiodothyronine (T3) level but does not change thymus, spleen weight, serum cortisol, corticosterone and thyroxin (T4) levels. The long-term (4°C, 8 h/day, 4 weeks) intermittent cold exposure (LICE) induces adaptation to the cold and increases T4 level. The brief (4°C, 1.5 h/day, 4 weeks) intermittent cold exposure (BICE) also evokes adaptation to the cold but had no effect on the blood pressure, the cardiac tolerance to ischemia/reperfusion, and does not change thymus, spleen weight, serum cortisol, corticosterone, T3 and T4 levels.

Pharmacology of mitochondrial permeability transition pore inhibitors.

It is now firmly established that an important event in the formation of reperfusion injury of the heart is the opening of mitochondrial permeability transition pores (mPTPs), which changes the permeability of the mitochondria. mPTP opening results in the death of cardiomyocytes through activation of apoptosis and necroptosis. Experimental studies have shown that pharmacological inhibition of mPTP opening promotes the reduction in the infarct size and the suppression of apoptosis. Indeed, studies have shown the efficacy of mPTP inhibitors in animal models of myocardial reperfusion and isolated human myocardial trabeculae. However, clinical trials of cyclosporin A and TRO40303 have not provided a clear answer to the question of the effectiveness of mPTP inhibitors in patients with acute myocardial infarction. This article presents an analysis of possible approaches for the pharmacological regulation of mPTP during reperfusion injury of the heart.

Role of protein kinase C, PI3 kinase, tyrosine kinases, NO-synthase, KATP channels and MPT pore in the signaling pathway of the cardioprotective effect of chronic continuous hypoxia.

It was established that adaptation to chronic continuous normobaric hypoxia (CCNH) increases cardiac tolerance to ischemia and reperfusion. It was performed coronary artery occlusion (20 min) and reperfusion (3 h) in Wistar rats. CCNH promoted a decrease in the infarct size/area at risk ratio in 2-fold. CCNH promoted an increase in the nitrite/nitrate levels in blood serum and myocardium. Pretreatment with protein kinase C (PKC) inhibitor chelerythrine, NO-synthase (NOS) inhibitor L-NAME, iNOS inhibitor S-methylisothiourea, KATP channel blocker glibenclamide, mitoKATP channel blocker 5-hydroxydecanoic acid abolished the infarct-reducing effect of CCNH. The non-selective tyrosine kinase inhibitor genistein attenuated but not eliminated infarct-sparing effect of CCNH. The nNOS inhibitor 7-nitroindazole, sarcKATP channel blocker HMR 1098, MPT pore inhibitor atractyloside, PI3 kinase inhibitor wortmannin did not reverse infarct-limiting effect of CCNH. It was concluded that infarct-reducing effect of CCNH is mediated via PKC, iNOS activation and mitoKATP channel opening. While nNOS, PI3 kinase, sarcKATP channel, MPT pore are not involved in the development of CCNH-induced cardiac tolerance to impact of ischemia-reperfusion.

Prospects for Creation of Cardioprotective and Antiarrhythmic Drugs Based on Opioid Receptor Agonists.

It has now been demonstrated that the µ, δ1 , δ2 , and κ1 opioid receptor (OR) agonists represent the most promising group of opioids for the creation of drugs enhancing cardiac tolerance to the detrimental effects of ischemia/reperfusion (I/R). Opioids are able to prevent necrosis and apoptosis of cardiomyocytes during I/R and improve cardiac contractility in the reperfusion period. The OR agonists exert an infarct-reducing effect with prophylactic administration and prevent reperfusion-induced cardiomyocyte death when ischemic injury of heart has already occurred; that is, opioids can mimic preconditioning and postconditioning phenomena. Furthermore, opioids are also effective in preventing ischemia-induced arrhythmias.

Reperfusion Cardiac Injury: Receptors and the Signaling Mechanisms.

It has been documented that Ca2+ overload and increased production of reactive oxygen species play a significant role in reperfusion injury (RI) of cardiomyocytes. Ischemia/reperfusion induces cell death as a result of necrosis, necroptosis, apoptosis, and possibly autophagy, pyroptosis and ferroptosis. It has also been demonstrated that the NLRP3 inflammasome is involved in RI of the heart. An increase in adrenergic system activity during the restoration of coronary perfusion negatively affected cardiac resistance to RI. Toll-like receptors are involved in RI of the heart. Angiotensin II and endothelin-1 aggravated ischemic/reperfusion injury of the heart. Activation of neutrophils, monocytes, CD4+ T-cells and platelets contributes to cardiac ischemia/reperfusion injury. Our review outlines the role of these factors in reperfusion cardiac injury.

Activation of peripheral δ2-opioid receptor prevents reperfusion heart injury.

Coronary artery occlusion (45 min) and reperfusion (2 h) was performed in rats anesthetized with α-chloralose. Opioid receptor agonists were administered intravenously 5 min before reperfusion, while opioid receptor antagonists were administered 10 min before reperfusion. The non-selective opioid δ-receptor agonist DADLE at a dose of 0.088 mg/kg had no effect the infarct size/area at risk ratio. The selective opioid δ-receptor agonist BW373 was administered at a dose of 1 mg/kg. This opioid at a dose of 1 mg/kg reduced infarct size. The selective opioid δ1-receptor agonist DPDPE at a dose of 0.1 mg/kg and 0.969 mg/kg did not affect infarct size. The selective opioid δ2-receptor agonist deltorphin II at a dose of 0.12 mg/kg reduced infarct size by one half. The opioid δ-receptor agonist p-Cl-Phe-DPDPE was administered at a dose of 0.105 mg/kg and 1.02 mg/kg. This opioid at a dose of 1.02 mg/kg reduced infarct size. The universal opioid receptor antagonists, naltrexone and naloxone methiodide acting on peripheral opioid receptor, as well as the selective opioid δ-receptor antagonist TIIP[ψ], the selective opioid δ2-receptor antagonist naltriben eliminated the infarct limiting effect of deltorphin II. The selective opioid κ receptor antagonist nor-binaltorphimine, the selective opioid µ receptor antagonist CTAP, and the selective opioid δ1-receptor antagonist BNTX did not abolish the protective effect of deltorphin II. Deltorphin II exhibited the most pronounced cardioprotective effect during reperfusion. These studies clearly indicate that the activation of opioid δ2-receptor located in cardiomyocytes increases the resistance of the heart to reperfusion injury.

High carbohydrate high fat diet causes arterial hypertension and histological changes in the aortic wall in aged rats: The involvement of connective tissue growth factors and fibronectin.

BACKGROUND: Age and diabetes are risk factors for arterial hypertension. However, the relationship between age, connective tissue growth factors, vascular aging and arterial hypertension while on a the high-carbohydrate high-fat diet (HCHFD) remains poorly understood. PURPOSE: To estimate the relationship between humoral factors, the morphological changes of aorta and impaired blood pressure regulation under the influence of age and a HCHFD. METHODS: A study was carried out in male Wistar rats, which were divided into the following groups: 1st (n = 15) - naive young rats; 2nd (n = 15) - young rats, exposed to HCHFD; 3rd (n = 14) - naive old rats; 4th (n = 12) - old rats exposed to HCHFD. The age of old rats was 540 days, and young rats 150 days at the end of the diet. HCHFD contained proteins 16%, fats 21%, carbohydrates 46%, including 17% fructose, 0.125% cholesterol, 90 days. Blood pressure and body weight were measured weekly, carbohydrate metabolism, histological signs of changes in the aorta, serum transforming growth factor-ß (TGF-ß), connective tissue growth factor (CTGF), fibronectin, and endothelin-1 levels were determined one week after the onset of diet. RESULTS: The severity of arterial hypertension and its histological signs in the aortic wall was found to be most pronounced in elderly rats kept on a HCHFD. In young rats kept on a HCHFD, arterial hypertension was transient. An increase in systolic blood pressure has a positive correlation with the degree of obesity, serum fibronectin, and endothelin-1 content, and impaired carbohydrate metabolism. The rise in diastolic blood pressure has a positive correlation with the serum CTGF, endothelin-1, fibronectin levels and aortic wall thickness, and impaired carbohydrate metabolism. A rise in the serum concentration of fibronectin was also associated with increased endothelin-1, TGFß and CTGF serum levels. CONCLUSION: This study indicated that an increase in blood pressure in old rats with a high-carbohydrate high-fat diet is due to a disturbance of a structure of the vascular wall, the release of fibronectin, which can occur under the influence of carbohydrate metabolism disorders, endothelin-1, TGFß and CTGF.

Takotsubo Syndrome: Clinical Manifestations, Etiology and Pathogenesis.

The purpose of the review is the analysis of clinical and experimental data on the etiology and pathogenesis of takotsubo syndrome (TS). TS is characterized by contractile dysfunction, which usually affects the apical region of the heart without obstruction of coronary artery, moderate increase in myocardial necrosis markers, prolonged QTc interval (in 50% of patients), sometimes elevation of ST segment (in 19% of patients), increase N-Terminal Pro-B-Type Natriuretic Peptide level, microvascular dysfunction, sometimes spasm of the epicardial coronary arteries (in 10% of patients), myocardial edema, and life-threatening ventricular arrhythmias (in 11% of patients). Stress cardiomyopathy is a rare disease, it is observed in 0.6 - 2.5% of patients with acute coronary syndrome. The occurrence of takotsubo syndrome is 9 times higher in women, who are aged 60-70 years old, than in men. The hospital mortality among patients with TS corresponds to 3.5% - 12%. Physical or emotional stress do not precede disease in all patients with TS. Most of patients with TS have neurological or mental illnesses. The level of catecholamines is increased in patients with TS, therefore, the occurrence of TS is associated with excessive activation of the adrenergic system. The negative inotropic effect of catecholamines is associated with the activation of ß2 adrenergic receptors. An important role of the adrenergic system in the pathogenesis of TS is confirmed by studies which were performed using 125I-metaiodobenzylguanidine (125I -MIBG). TS causes edema and inflammation of the myocardium. The inflammatory response in TS is systemic. TS causes impaired coronary microcirculation and reduces coronary reserve. There is a reason to believe that an increase in blood viscosity may play an important role in the pathogenesis of microcirculatory dysfunction in patients with TS. Epicardial coronary artery spasm is not obligatory for the occurrence of TS. Cortisol, endothelin-1 and microRNAs are challengers for the role of TS triggers. A decrease in estrogen levels is a factor contributing to the onset of TS. The central nervous system appears to play an important role in the pathogenesis of TS.

Hibernation-like state induced by an opioid peptide protects against experimental stroke.

BACKGROUND: Delta opioid peptide [D-ala2,D-leU5]enkephalin (DADLE) induces hibernation in summer ground squirrels, and enhances preservation and survival of isolated or transplanted lungs and hearts. In the present study, we investigated the protective effect of DADLE in the central nervous system. RESULTS: Adult Sprague-Dawley rats were pretreated with DADLE (4 mg/kg every 2 h x 4 injections, i.p.) or saline prior to unilateral occlusion of the middle cerebral artery (MCA). Daily behavioral tests revealed that ischemic animals treated with DADLE did not show any significant behavioral dysfunctions compared with saline-treated ischemic animals. Opioid antagonists only transiently inhibited the protective effect of DADLE, indicating the participation of non-opioid mechanisms in DADLE neuroprotection. Histological examination using triphenyltetrazolium chloride (TTC) revealed that brains from ischemic animals treated with DADLE, either alone or with adjuvant opioid blockers, exhibited almost completely intact striata. In contrast, brains from ischemic animals that received saline showed significant infarction in the lateral striatum. Analyses of apoptotic cell death revealed a significant increase in the p-53 mRNA expression in the striatum of ischemic animals that received saline, while those that received DADLE exhibited near normal striatal p-53 expression. This protective effect was accompanied by significant increments in protein levels of glial cell line-derived neurotrophic factor in the striatum of DADLE-treated ischemic animals. CONCLUSION: These results indicate that DADLE protected against necrotic and apoptotic cell death processes associated with ischemia-reperfusion injury. The present study demonstrates that delta opioids are crucially involved in stroke, suggesting that the opioid system is important in the study of brain injury and protection.

Is oxidative stress of adipocytes a cause or a consequence of the metabolic syndrome?

Metabolic syndrome is accompanied by oxidative stress in animals and humans. The main source of ROS in experimental metabolic syndrome is NADPH oxidase and possibly adipocyte mitochondria. It is now documented that oxidative stress induces insulin resistance of adipocytes and increases secretion of leptin, MCP-1, IL-6, and TNF-α by adipocytes. It was established that oxidative stress induces a decrease in adiponectin production by adipocytes. It has also been shown that obesity itself can induce oxidative stress. Oxidative stress can cause an alteration of intracellular signaling in adipocytes that apparently leads to the formation of insulin resistance of adipocytes. Chronic stress, glucocorticoids, mineralocorticoids, angiotensin-II, TNF-α also play an important role in the pathogenesis of oxidative stress of adipocytes. Oxidative stress is not only a consequence of metabolic syndrome, but also a reason and a foundational link in the pathogenesis of the metabolic syndrome.

Physiological and Pathological Role of TRPV1, TRPV2 and TRPV4 Channels in Heart.

Transient receptor potential vanilloid channel 2 (TRPV2) is required for normal cardiac contractility. The stimulation of TRPV1 in isolated cardiomyocytes can aggravate the effect of hypoxia/ reoxygenation (H/R) on H9C2 cells. The knockout of the TRPV1 gene promotes increased tolerance of the isolated perfused heart to the impact of ischemia/reperfusion (I/R). However, activation of TRPV1 increases the resistance of the heart to I/R due to calcitonin gene-related peptide (CGRP) release from afferent nerve endings. It has been established that TRPV1 and TRPV2 are involved in the pathogenesis of myocardial infarction and, in all likelihood, ensure the cardiac tolerance to the ischemia/reperfusion. It has also been documented that the activation of TRPV4 negatively affects the stability of cardiomyocytes to the H/R. The blockade of TRPV4 can be considered as a new approach to the prevention of I/R injury of the heart. Studies also indicate that TRPV1 is involved in the pathogenesis of cardiac hypertrophy and that TRPV2 channels participate in the pathogenesis of dilated cardiomyopathy. Excessive expression of TRPV2 leads to chronic Ca2+- overload of cardiomyocytes, which may contribute to the development of cardiomyopathy.

Trigger, Signaling Mechanism and End Effector of Cardioprotective Effect of Remote Postconditioning of Heart.

The hypothetical trigger of remote postconditioning (RPost) of the heart is the highmolecular weight hydrophobic peptide(s). Nitric oxide and adenosine serve as intermediaries between the peptide and intracellular structures. The role of the autonomic nervous system in RPost requires further study. In signaling mechanism RPost, kinases are involved: protein kinase C, PI3, Akt, JAK. The hypothetical end effector of RPost is aldehyde dehydrogenase-2, the transcription factors STAT, Nrf2, and also the BKCa channel.

Activation of peripheral delta opioid receptors increases cardiac tolerance to arrhythmogenic effect of ischemia/reperfusion.

OBJECTIVES: The objective of this study was to investigate the role of peripheral µ, δ1, δ2, and nociceptin opioid receptors agonists in the regulation of cardiac tolerance to the arrhythmogenic effect of ischemia/reperfusion in rats. METHODS: Anesthetized open-chest male Wistar rats were subjected to either 45 minutes of left coronary artery occlusion (phase 1a 10 minutes and phase 2b 35 minutes) and 2 hours of reperfusion in Experiment 1 or 10 minutes of ischemia and 10 minutes of reperfusion in Experiment 2. In Experiment 1, saline or vehicle controls and the mu-specific opioids dermorphin-H (Derm-H) and ([d-Ala2, N-Me-Phe4, Gly-ol5] enkephalin (DAMAGO); the delta-1-specific opioid d-Pen2,5enkephalin (DPDPE); nociceptin; and the delta-2-specific opioids deltorphin-II (Delt-II), Delt-Dvariant (Delt-Dvar), and deltorphin-E (Delt-E) were infused 15 minutes prior to ischemia. In Experiment 2, DPDPE, Delt-D, Delt-Dvar, and Delt-E were infused at 15 minutes prior to ischemia. The universal opioid receptor antagonist naltrexone, the peripherally acting antagonist naloxone methiodide, the selective δ1 antagonist 7-benzylidene naltrexone maleate, and the specific δ2 antagonist naltriben mesylate were infused 25 minutes prior to ischemia. RESULTS: In Experiment 1, pretreatment with the µ opioids Derm-H and DAMGO, DPDPE, and nociceptin at all doses tested did not reduce the incidence of ischemia-induced arrhythmias compared to controls during 45 minutes of ischemia. The δ2 opioids Delt-II (0.12 mg/kg), Delt-Dvar (0.3 mg/kg), and Delt-E (0.18 mg/kg) all demonstrated significant antiarrhythmic effects at the 150 nmol/kg dose compared to saline or vehicle controls. Nine of 19 animals treated with Delt-II were tolerant without ventricular arrhythmias to the arrhythmogenic effect of ischemia during the first 10 minutes of ischemia (phase 1a) and 11 of 19 were without ventricular arrhythmias during the following 35 minutes of ischemia (phase 1b). Delt-II also decreased the incidence of premature ventricular contractions and ventricular tachycardia by almost half during phase 1a. Delt-II did not affect the incidence of ventricular fibrillation (VF). Pretreatment with Delt-Dvar and Delt-E completely blocked the incidence of VF in phase 1b. Delt-E also decreased premature ventricular contractions by 50%, and the incidence of ventricular tachycardia decreased over twofold in phase 1b of ischemia. There was no enhanced tolerance by any of the delta-2 opioids to the arrhythmogenic effect of reperfusion after long-term ischemia. In Experiment 2, after 10 minutes of ischemia and 10 minutes of reperfusion, Delt-II (0.12 mg/kg) reduced the incidence of premature ventricular contractions and ventricular tachycardia compared to controls, and completely blocked the incidence of VF following 10 minutes of reperfusion. Delt-Dvar and Delt-E were without effect, as was DPDPE following 10 minutes of reperfusion. The antiarrhythmic effect of Delt-II during 10 minutes of ischemia and 10 minutes of reperfusion was completely blocked by the peripherally acting opioid receptor inhibitor naloxone methiodide and the selective delta-2 opioid receptor inhibitor naltriben mesylate, but not by the selective delta-1 inhibitor 7-benzylidene naltrexone maleate. The antagonists alone had no effect on arrhythmogenesis. CONCLUSIONS: Peripheral delta-2 opioid receptor activation by Delt-II, Delt-Dvar, and Delt-E enhanced cardiac tolerance to the arrhythmogenic effects of ischemia.

Comparative analysis of the cardioprotective properties of opioid receptor agonists in a rat model of myocardial infarction.

OBJECTIVES: This study was conducted to test the hypothesis that opioid receptor (OR)-mediated cardioprotection is agonist specific when administered prior to coronary artery occlusion and reperfusion in a rat model. METHODS: Anesthetized open-chest male Wistar rats were subjected to 45 minutes of left coronary artery occlusion and 2 hours of reperfusion. Opioid agonists were infused 15 minutes prior to coronary artery occlusion. Two control groups and 15 opioid-treated groups were studied. Controls were infused with either saline alone (n = 16) or dimethyl sulfoxide plus hydroxypropyl-ß-cyclodextrin in saline (n = 19). The µ-selective agonist DAMGO was infused at either 150 nmol/kg (n = 15) or 1500 nmol/kg (n = 14), and dermorphin-H was infused at 150 nmol/kg (n = 14). The δ1 -selective agonist d-Pen²(,)5 enkephalin (DPDPE) was infused at 150 nmol/kg (n = 16) or 1500 nmol/kg (n = 14). The δ2 -selective agonists deltorphin II (n = 16), deltorphin-D(variant) (n = 15), and deltorphin-E (n = 14) were infused at 150 nmol/kg. The selective κ1 opioid agonist U-50488 was infused at 240 nmol/kg (n = 14), 1500 nmol/kg (n = 14), and 2,400 nmol/kg (n = 14). The selective κ2 opioid agonist GR-89696 was infused at 150 nmol/kg (n = 14) and 1500 nmol/kg (n = 15). Orphinan FQ (nociceptin), also referred to as OR-like 1 (ORL1), was infused at 220 nmol/kg (n = 15) and 1500 nmol/kg (n = 15). The infarct size/area at risk (IS/AAR) ratio was determined after reperfusion by negative staining with patent blue violet dye. Hemodynamic parameters including heart rate, mean arterial blood pressure (MAP), and rate pressure product (RPP) were determined. RESULTS: Pretreatment with the δ2 OR agonist deltorphin II (150 nmol/kg) significantly reduced the IS/AAR ratio, while deltorphin-D(variant) and deltorphin-E did not exhibit an infarct-sparing effect at that treatment dose. Activation of δ1 OR by DPDPE, κ1 OR by U-50488, κ2 OR by GR-89696, µ OR by DAMGO, dermorphin-H, and nociceptin had no effect on the IS/AAR ratio. U-50488 at 2,400 nmol/L induced a bradycardic effect. All other opioids had no effect on hemodynamic parameters at the doses tested. CONCLUSIONS: Peripheral δ2 OR activation by deltorphin II induces infarct size reduction in this animal model. Agonists of µ, δ1, κ1, κ2, and nociceptin receptors at the doses tested did not induce cardiac tolerance to ischemia/reperfusion injury in vivo.

Activation of peripheral delta2 opioid receptors increases cardiac tolerance to ischemia/reperfusion injury Involvement of protein kinase C, NO-synthase, KATP channels and the autonomic nervous system.

AIMS: This study aims to investigate the role of peripheral delta(2) opioid receptors in cardiac tolerance to ischemia/reperfusion injury and to examine the contribution of PKC, TK, K(ATP) channels and the autonomic nervous system in delta(2) cardioprotection. MAIN METHODS: Deltorphin II and various inhibitors were administered in vivo prior to coronary artery occlusion and reperfusion in a rat model. The animals were monitored for the development of arrhythmias, infarct development and the effects of selected inhibitors. KEY FINDINGS: Pretreatment with peripheral and delta(2) specific opioid receptor (OR) antagonists completely abolished the cardioprotective effects of deltorphin II. In contrast, the selective delta(1) OR antagonist 7-benzylidenenaltrexone (BNTX) had no effect. The protein kinase C (PKC) inhibitor chelerythrine and the NO-synthase inhibitor L-NAME (N-nitro-L-arginine methyl ester) also reversed both deltorphin II effects. The nonselective ATP-sensitive K+ (K(ATP)) channel inhibitor glibenclamide and the selective mitochondrial K(ATP) channel inhibitor 5-hydroxydecanoic acid only abolished the infarct-sparing effect of deltorphin II. Inhibition of tyrosine kinase (TK) with genistein, the ganglion blocker hexamethonium and the depletion of endogenous catecholamine storage with guanethidine reversed the antiarrhythmic action of deltorphin II but did not change its infarct-sparing action. SIGNIFICANCE: The cardioprotective mechanism of deltorphin II is mediated via stimulation of peripheral delta(2) opioid receptors. PKC and NOS are involved in both its infarct-sparing and antiarrhythmic effects. Infarct-sparing is dependent upon mitochondrial K(ATP) channel activation while the antiarrhythmic effect is dependent upon TK activation. Endogenous catecholamine depletion reduced antiarrhythmic effects but did not alter the infarct-sparing effect of deltorphin II.

Delta 2-specific opioid receptor agonist and hibernating woodchuck plasma fraction provide ischemic neuroprotection.

OBJECTIVES: The authors present evidence that the delta opioid receptor agonist Deltorphin-D(variant) (Delt-D(var)) and hibernating woodchuck plasma (HWP), but not summer-active woodchuck plasma (SAWP), can provide significant neuroprotection from focal ischemia in mice by a mechanism that relies in part on reducing nitric oxide (NO) release in ischemic tissue. METHODS: Cerebral ischemia was produced in wild-type and NO synthase-deficient (NOS(-/-)) mice by transient, 1-hour middle cerebral artery occlusion (MCAO). Behavioral deficits were determined at 22 hours and infarct volume was assessed at 24 hours after MCAO. Mice were treated with saline or Delt-D(var) at 2.0 and 4.0 mg/kg, or 200 microL of HWP or SAWP. NOS(-/-) mice were treated with either saline or Delt-D(var) at 4.0 mg/kg. NO release was determined using an N9 microglial cell line pretreated with delta- or mu-specific opioids and HWP or SAWP prior to activation with lipopolysaccharide and interferon-gamma. Nitrate in the medium was measured as an indicator of NO production. RESULTS: Infusion of Delt-D(var) or HWP (but not SAWP) decreased infarct volume and improved behavioral deficits following 1 hour of MCAO and 24 hours of reperfusion. In NOS(-/-) mice, endothelial NOS(+/+) is required to provide Delt-D(var)-induced neuroprotection. Delt-D(var) and HWP dose-dependently decreased NO release in cell culture, while SAWP and other delta- and mu-specific opioids did not. CONCLUSIONS: Delt-D(var) and HWP, but not SAWP, are effective neuroprotectant agents in a mouse model of transient MCAO. In cell culture, the mechanism of this ischemic neuroprotection may rely in part on their ability to block NO release.

24-hour pretreatment with delta opioid enhances survival from hemorrhagic shock.

OBJECTIVES: Delta opioids have been shown to confer ischemic preconditioning and pharmacologic ischemic preconditioning to the myocardium. However, their role in providing extended pharmacologic ischemic preconditioning in hemorrhagic shock has not been explored. The authors examined the effects of 24-hour preinfusions of a selective delta opioid receptor agonist, Deltorphin-Dvariant (Delt-Dvar), on hemodynamic stability and duration of survival in a rat model of severe hemorrhagic shock. METHODS: Conscious Sprague-Dawley rats with indwelling catheters were hemorrhaged at a rate of 3.18 mL/l00 g over 20 minutes. Twenty-four hours before hemorrhage, the control group (n = 14) was infused with 1.0 mL lactated Ringer's solution, and the Delt-Dvar-treated group (n = 22) was infused with 5.0 mg/kg Delt-Dvar in 1.0 mL lactated Ringer's solution. Rats were continuously monitored for heart rate (HR), mean arterial pressure, and four-hour survival rates. Plasma lactate levels were determined at the beginning of hemorrhage and the end of hemorrhage. RESULTS: At 240 minutes, only one of 14 controls (7.1%) survived, while 16 (72.7%) of the 22 experimental rats survived. No significant differences in heart rate between controls and Delt-Dvar-treated rats were noted. Increases in mean arterial pressure of Delt-Dvar-treated rats at the beginning of hemorrhage and at the end of hemorrhage were found to be significant (p < 0.05). At 240 minutes, heart rate and mean arterial pressure were not different between the single surviving control and the Delt-Dvar group. At the end of hemorrhage, lactate levels in the Delt-Dvar-treated group were 8.5 (+/- 0.5) mmol/L versus 10.8 (+/- 0.6) mmol/L (p < 0.05) in the control group. CONCLUSIONS: Twenty-four-hour pretreatment with Delt-Dvar decreases plasma lactate levels and improves hemodynamic stability and survival during hemorrhagic shock. The use of delta-specific opioids may improve survival from hemorrhagic shock and have clinical utility in providing ischemic protection in scenarios of planned ischemia.

A delta2-opioid agonist inhibits p38 MAPK and suppresses activation of murine macrophages.

BACKGROUND: delta-Opioid agonists have been shown to attenuate ischemic organ injury in multiple models. The purpose of the present study was to determine if delta-opioid agonists could inhibit proinflammatory cytokine production by macrophages. MATERIAL AND METHODS: Murine macrophages (RAW 264.7) were pretreated for 4 h with media, a dose range (10(-4) to 10(-7) M) of DADLE ([D-Ala2], D-Leu5]-enkephalin, a nonspecific delta-opioid receptor agonist), a dose range (10(-4) to 10(-7) M) of DPDPE ([D2,5Pen]-enkephalin, a specific delta1-opioid receptor agonist), or a dose range (10(-4) to 10(-7) M) of Deltorphin-Dvariant (a specific delta2 opioid receptor agonist) and then incubated with 0.1 microg/ml lipopolysaccharide (LPS) for 1 or 4 h. Cytokine levels were measured by enzyme-linked immunosorbent assay. Activation of NF-kappaB, AP-1, and p38 MAPK were determined by mobility shift assays and Western blot. RESULTS: LPS induced significant increases in TNFalpha and MIP-2 production. Deltorphin-Dvariant, but not DADLE or DPDPE, dose-dependently reduced both TNFalpha and MIP-2 production. Deltorphin-Dvariant did not alter activation of the transcription factors NF-kappaB or AP-1, but greatly reduced activation of p38 MAPK. CONCLUSIONS: The data show that delta2- but not delta1-opioid agonists suppress LPS-induced p38 MAPK activation and expression of TNFalpha and MIP-2.

Hibernation induction trigger reduces hypoxic damage of swine skeletal muscle.

A link between the cardioprotective benefits of pharmacological preconditioning and natural mammalian hibernation is considered to involve the cellular activation of opioid receptors and subsequent opening of K(ATP) channels. In previous studies, we have demonstrated the protective effects of specific delta-opioid agonists against porcine cardiac ischemia/reperfusion injury. We hypothesize here that preincubation with hibernation induction trigger (HIT) should confer a similar protection in skeletal muscles. Therefore, muscle bundles from swine were pretreated with plasma from hibernating woodchucks (HWP) for 30 min, then exposed to hypoxia for 90 min and reoxygenation for 120 min. Stimulated twitch forces were assessed. The functional effects of pretreatment with nonhibernation (summer) woodchuck plasma, a K(ATP) blocker, or opioid antagonist were also studied. During the reoxygenation period, significantly greater force recoveries were observed only for bundles pretreated with HWP; this response was blocked by naloxone (P < 0.05). We conclude that HIT pretreatment could be used to confer protection against hypoxia/reperfusion injury of skeletal muscles of nonhibernators; it could potentially be utilized to prevent injury during surgical procedures requiring ischemia.