Comparative Analysis of Infarct Size Limiting Activity of δ-Opioid Receptor Agonists in Reperfused Heart In Vivo.

The study examined the effects of δ-opioid receptor (OR) agonists on infarct size on cardiac ischemia (45 min) and reperfusion (120 min) in vivo model in rats narcotized with α-chloralose. The OR agonists were injected intravenously 5 min prior to reperfusion, OR antagonists were administered 10 min before reperfusion. A selective δ-OR agonist BW373U86 (1 mg/kg) reduced the infarct size/area at risk ratio. A selective δ1-OR agonist DPDPE injected in the doses of 0.1 or 0.969 mg/kg produced no effect on infarct size. The selective δ2-OR agonist deltorphin II (0.12 mg/kg) reduced infarct size/area at risk ratio by 2 times. The δ-OR agonist p-Cl-Phe-DPDPE (1 mg/kg) reduced infarct size/area at risk ratio by 40%. Naltrexone and naloxone methiodide, the peripheral OR antagonists, and selective δ2-OR antagonist naltriben prevented the infarct size limiting effect of deltorphin II. Therefore, activation of peripheral δ2-OR enhanced cardiac resistance against toxic action of reperfusion. During reperfusion, deltorphin II demonstrated the most pronounced cardioprotective activity.

[ROLE OF OPIOID RECEPTORS IN THE REGULATION OF RESISTANCE OF HEART TO IMPACT OF ISCHEMIA-REPERFUSION].

Activation of m-, d1-, d2- and k1-opioid receptors increases cardiac resistance to ischemia-reperfusion. The cardioprotective effect of opioids in many cases appears to be associated with the activation of the peripheral OR. However, when it comes to non-peptide agonists OR able to cross the blood-brain barrier, we cannot exclude the involvement of central opioid receptors in cardioprotection. Endogenous opioids are not involved in the regulation of cardiac tolerance to ischemia- reperfusion in non-adapted animals. Stimulation of k1- and d1-OP may exert delayed cardioprotective effect. Activation d- and k1-OP reduces the intensity of cardiomyocyte apoptosis after reperfusion. The results of studies related to the inotropic effect of opioids during reperfusion of the heart remain highly controversial.

[THE ROLE OF THE ENDOGENOUS CANNABINOID SYSTEM IN THE FORMATION OF THE GENERAL ADAPTATION SYNDROME].

Nonopioid stress-induced analgesia is the consequence of activation of CB1 receptors by the increased level of 2-arachidonoyl glycerol, anandamide in the periaqueductal gray matter in the midbrain. The activation of cannabinoid CB1 receptors inhibits stress-induced ulcerogenesis due to the strengthening of the antioxidant defense of the gastric mucosa. CB1 receptor antagonists promote an increase in ACTH and corticosterone concentrations in the blood of intact animals, the knockout of the gene encoding the CB1 receptor exhibits the same effect. Antagonists of CB1 receptors enhance the stressor elevation of ACTH and corticosterone levels in the blood of experimental animals. It was found an increase in stress-induced elevation of corticosterone and ACTH levels in the blood of mice with a knockout of the gene encoding the CB1 receptor. An increase in the endogenous anandamide level or disturbance of the reuptake of endogenous cannabi-noids after application of pharmacological agents promotes reducing corticosterone level in stressed animals. Consequently, endogenous cannabinoids inhibit basal and suppress stress-induced activity of the hypothalamic-pituitary-adrenal axis. The indicated regulation is carried out on the level of the hypothalamus, pituitary and adrenal cortex. Stimulation of central cannabinoid receptors leads to an activation of the sympathetic system. The activation of peripheral CB1 receptors leads to inhibition of norepinephrine release from sympathetic terminals and epinephrine release from the adrenal glands. The endogenous CB1 receptor agonists play an anxiolytic role and prevent the occurrence of pathological anxiety.

[MIMICKING ISCHEMIC PRECONDITIONING PHENOMENON THROUGH THE IMPACT ON THE CANNABINOID RECEPTORS: ROLE OF PROTEIN KINASE AND NO-SYNTHASE].

It was established that CB 1-receptors stimulation mimic preconditioning phenomena. Since the cardioprotective effect of cannabinoid HU-210 is occurred both in the experiments in vivo and in the experiments in vitro there are reasons to believe that the protective effect of HU-210 is me- diated via an activation of cardiac CB1-receptors. It is established that the cardioprotective effect of cannabinoid HU-2 10 is depends upon a stimulation ofprotein kinase C whereas NO-synthase is not involved in protective impact of CB1-receptor stimulation.

Prospects of Using of κ-Opioid Receptor Agonists U-50,488 and ICI 199,441 for Improving Heart Resistance to Ischemia/Reperfusion.

We studied the ability of the agonist of κ1-opioid receptors U-50,488 in doses of 0.1 and 1 mg/kg to simulate ischemic pre- and postconditioning of the heart and κ-opioid receptors ICI 199,441 in a dose of 0.1 mg/kg to simulate the antiarrhythmic effect of heart preconditioning. The duration of ischemia was 10 or 45 min and the duration of reperfusion was 10 min or 2 h. Administration of 1 mg/kg U-50,488 both before ischemia and 5 min before reperfusion produced a pronounced antiarrhythmic effect. U-50,488 injected 5 min before reperfusion 2-fold reduced the ratio of infarction to risk area. Administration of ICI 199,441 in a dose of 0.1 mg/kg 15 min before ischemia produced a potent antiarrhythmic effect. Antiarrhythmic effect of κ-opioid receptor agonists depended on activation of κ-opioid receptors.

Role of cyclic nucleotides and NO synthase in mechanisms of cardioprotective effects of cannabinoid HU-210.

Isolated perfused rat heart was subjected to global ischemia (45 min) followed by reperfusion (30 min). Under these conditions, the level of creatine phosphokinase in the perfusate increased by 4.5 times. Perfusion (10 min) of the isolated heart with a solution containing cannabinoid HU-210 (0.1 or 1.0 µmol/liter) was followed by a 2-fold decrease in creatine phosphokinase level in the perfusate. The cardioprotective effect of HU-210 remained unchanged under condition of NO synthase inhibition. Cannabinoid HU-210 reduced the concentration of cAMP in the myocardium by 2 times during reperfusion, but did not affect it before and during ischemia. This agent also did not change the level of cGMP in the myocardium before and during ischemia and during reperfusion. The results of the experiment suggest that the cardioprotective effect of HU-210 can be determined by a decrease in cAMP level in the myocardium during reperfusion. cGMP and NO synthase do not contribute to cytoprotective effect of HU-210.

Comparative analysis of early and delayed cardioprotective and antiarrhythmic efficacy of hypoxic preconditioning.

Hypoxic preconditioning produces an infarct-limiting effect both in the early and delayed periods. The increase in heart resistance to ischemia-repefusion was more pronounced after early preconditioning. Hypoxic preconditioning did not change heart resistance to the arrhythmogenic effect of coronary occlusion and reperfusion.

[Perspective of use of agonists of adenosine and opioid receptors for prevention of reperfusion damages of heart. Analysis of experimental and clinical data].

In Russia inhospital lethality after acute myocardial infarction is 16.5-16.7%. The part of patients perishes even after recanalisation of infarct-related coronary artery as a result of reperfusion cardiac injury. Experimental data indicate that adenosine receptor agonists and opioids can prevent reperfusion damages of heart that is mimic postconditioning phenomena. Data of clinical observation show that adenosine during intravenous infusion or intracoronary administration during thrombolysis or percutaneous coronary intervention exert infarct reducing effect and eliminate manifestation of of "no-reflow" phenomenon. Clinical data indicate that morphine is able to prevent cardiac reperfusion injury in human. Thus, analysis of published data testifies that adenosine and opioid receptor agonists can be prototype for development of drugs for prophylaxis of reperfusion heart injury.

[Receptor and signalling mechanisms of antiarrhythmic effects of ischemic pre-conditioning].

It has been established that ischemic preconditioning (IP) exerts significant antiarrhythmic effects, as revealed in experiments both in vivo and in vitro. Consequently, processes arising within the myocardium play a key role in adaptive tolerance to ischemia/reperfusion. Preconditioning enhances cardiac electrical stability both in animals and humans. The antiarrhythmic effect of preconditioning is transient, with enhanced tolerance to ischemia-reperfusion triggered arrhythmogenesis dissipating 2-3 after the IP stimulus. The basis of the antiarrhythmic and cardioprotective effects of IP may differ. Preconditioning improves conduction of the cardiac electrical impulse, thereby preventing occurrence of re-entrant arrhythmias. NO-synthase and peroxynitrite play an important role in evolution of the antiarrhythmic effects of IP. Furthermore, intracellular Ca2+ may be a trigger of improved cardiac electrical stability after IP. It has been established that G(i/o)-protein coupled receptors are not involved in antiarrhythmic effects of IP, whereas bradykinin B2 and alpha1 adrenergic receptor activities are involved in IP-dependent improvements in cardiac electrical stability. Adenosine receptors contribute only partially to these effects. In terms of signalling mechanisms, protein kinase C appears essential to the antiarrhythmic effects of IP, whereas PI3-kinase and cyclooxygenase do not appear to be significantly involved. It has also been established that cardiac mast cells are involved in IP effects. Some data indicate that increased cardiac electrical stability with preconditioning depends upon mitoK(ATP) channel opening. Other data provide evidence that antiarrhythmic effects of preconditioning depends upon sarcK(ATP) channel opening. Some data indicate that an increase in electrical stability of heart after preconditioning depends upon mitoK(ATP) channel opening. Other data are evidence that antiarrhythmic effect of preconditioning depends upon sarCK(ATP) channel opening. Further work is needed to fully delineate the mechanistic basis of antiarrhythmic effects of IP.

[Signaling mechanism of cardioprotective effects of opioids].

It has been established that G(i/o)-proteins are an intermediate link that provides intracellular signaling between opioid receptors and protein kinases. Our investigations have shown that protein kinase C is involved in realization of the anti-necrotic and anti-apoptotic effects of opioids. PI3 and Akt kinases are involved in the cardioprotective effect of opioids. MEK1/2, ERK1/2, Src and JAK2 kinases play an important role in the cardioprotective effect of opioids. Further study of the participation of JNK, p70s6K and GRK2 in the opioid-induced increase of cardiac tolerance to ischemia and reperfusion is required. NO-synthase plays an important role in the cardioprotective action of opioids. Transactivation of opioid and adenosine receptors is an important element in the development of cardiac tolerance to ischemia and reperfusion. Opioid transactivation of EGF receptor is a connecting link between opioid receptors and ERK1/2 and PI3 kinase cascades.

[Perspective of creation of drugs on basis of opioids increasing cardiac tolerance to pathogenic impact of ischemia reperfusion].

It was established that delta- and kappa1-opioid receptor (OR) stimulation both in vivo and in vitro promotes a decrease of infarct size/area at risk (IS/AAR) ratio during ischemia and reperfusion of heart. mu-OR activation increases a tolerance of isolated perfused heart to impact of ischemia and reperfusion but has no effect on IS/AAR index in vivo. The ORL1-receptor agonist nociceptin does not exert IS/AAR ratio in vivo. Delta- and kappa1-OR stimulation prevents cardiomyocyte apoptosis during ischemia and reperfusion of heart. The delta- and kappa1-OR agonists mimic infarct-reducing effect of postconditioning. The OR inhibition does not impact IS/AAR index both in vivo and in vitro. The delta1-, delta2- and kappa1-OR agonists are the most perspective group of opioids for creation of drugs increasing cardiac tolerance to pathogenic impact of ischemia and reperfusion.

[Preconditioning impact on coronary perfusion during ischemia and reperfusion of heart].

Recent studies have confirmed that ischemic preconditioning prevents appearance of reperfusion endothelial dysfunction. However, the issue of preconditioning impact on no-reflow phenomenon remains unresolved. The receptor mechanisms involved in the cardioprotective and vasoprotective effects of preconditioning are different. The ability of preconditioning in preventing reperfusion endothelial dysfunction is dependent upon bradykinin B2-receptor activation and not dependent upon adenosine receptor stimulation. The vasoprotective effect of preconditioning is mediated via mechanisms relying in part on activation of protein kinase C, NO-synthase, cyclooxygenase, mitochondrial K(ATP)-channel opening and an enhancement of antioxidative protection of the heart. The delayed preconditioning also exerts endothelium-protective effect. Peroxynitrite, NO* and O2* are the triggers of this effect but a possible end-effector involves endothelial NO-synthase.

Variability of the biological activity of oxidized titanium implants.

Oxidized titanium is a biologically inert material, but bioinertness reduces biomechanical characteristics of titanium implants. Modification of the structure of oxide surface layer of BT 5-1 titanium by increasing its thickness (by 1.7 times) and pore diameter (by 1.4 times) and by adding phosphorus, aluminum, and zinc oxides to its composition leads to radical modification of its biological characteristics. These implants acquire osteoinductive properties in in vivo systems not found in pure or oxidized BT 1-00 titanium and fairly well maintain in vitro growth of mesenchymal cells.

[Role of receptor transactivation in the cardioprotective effects of preconditioning and postconditioning].

Analysis of published data indicates that the activity of receptors for adenosine, opioids, bradykinin, calcitonin-gene related peptides (CGRP) and epidermal growth factor (EGF) play important role in triggering the cardioprotective effects of ischemic preconditioning. Cannabinoids mimic the infarct-sparing effects of preconditioning. Endogenous adenosine, opioids, bradykinin and CGRP have also been implicated in infarct-reduction with ischemic postconditioning. Again, cannabinoids also mimic the protective effect of postconditioning. Recent works support heterodimerization of G-protein coupled receptors (GPCRs), and GPCR transactivation of EGF receptors. It was found that cross-talk between delta(j)-opioid receptors and adenosine A(1)-receptors is essential to cardiac protection. Furthermore, evidence implicates EGF receptor transactivation in cardioprotective effect of multiple GPCrs including adenosine, acetylcholine, bradykinin, and opioid receptors. Such findings support a convergent pathway in which multiple GPCRs may interact (or function independently) to transactivate EGF receptor-dependent kinase signaling and cytoprotection.

[Mediated role of NO-syntase, protein kinase C and KATP-channels in realization of cardioprotective impact of cannabinoid HU-210].

It was shown that perfusion of the isolated heart of rat with solution containing the CB1- and CB2-receptor agonist HU-210 at concentrations of 0.1 or 1.0 microM/L for a duration of 10 min at 20 min before global ischemia (45 min) and reperfusion (30 min) promotes a twofold decrease in creatine kinase levels in coronary effluent. It was established that KATP channel blockade by glibenclamide (1 microM/L) or inhibition of protein kinase C (2 microM/L) by chelerythrine abolishes the cardioprotective effect of HU-210. The inhibitor of NO synthase L-NAME (1 microM/L) had no effect on the anti-necrotic effect of HU-210. Thus, the intracellular signaling mechanism of the cardioprotective effect of the CB-agonist HU-210 involves the activation of KATP channels and protein kinase C without the participation of NO-synthase.

Opioid peptide deltorphin II simulates the cardioprotective effect of ischemic preconditioning: role of δ2-opioid receptors, protein kinase C, and K(ATP) channels.

The cardioprotective properties of a δ2-opioid receptor agonist deltorphin II were studied in rats with coronary occlusion and reperfusion. Opioid receptor ligands and inhibitors (glybenclamide, chelerythrine, and 5-hydroxydecanoate) were injected intravenously before ischemia and reperfusion. A δ2-opioid receptor agonist deltorphin II significantly decreased the infarction zone/risk zone index. This effect was abolished by naltrexone, naloxone methiodide, and δ2-opioid receptor antagonist naltriben, but not by a δ1-opioid receptor antagonist BNTX. The infarct-limiting effect of deltorphin II was not observed after inhibition of protein kinase C or blockade of mitochondrial K(ATP) channels.

[Significance of opioid receptors in regulation of cardiac tolerance in pathogenic impact of long-term ischemia-reperfusion in vivo].

The study aimed at investigation of the role of opioid receptor (OR) in regulation of cardiac tolerance to ischemia-reperfusion. Opioid receptor ligands and inhibitors were administered in vivo prior to coronary artery occlusion (45 min) and reperfusion (2 hrs). Occurring infraction size/area at risk (IS/AAR) ratio was determined. Pretreatment with the micro-OR agonists DAMGO and dermorphin H exerted no effect on the IS/AAR ratio. Activation of delta 1-OR by DPDPE did not alter cardiac tolerance in ischemia-reperfusion either. Pretreatment with the delta 2-OR agonists deltorphin D and deltorphin E or ORL1 receptor agonist nociceptin exerted no effect on the IS/AAR ratio. Stimulation of K-OR by selective agonists did not modify cardiac tolerance to ischemia-reperfusion. The delta 2-OR agonist deltorphin II significantly reduced the IS/AAR index. This effect was prevented by treatment with naltrexone, naloxone methiodide and the delta 2-OR antagonist naltriben but not by the delta 1-OR antagonist BNTX. The infarction-limiting effect of deltorphin II was also abolished by inhibition of protein kinase C (PKC) and mitochondrial Katp channels. Thus, the agonists of micro, delta 1, kappa, and ORL1 receptors in used doses did not affect cardiac tolerance in ischemia-reperfusion injury in vivo. The peripheral delta 2-OR activation induces infarction size reduction. Its infarction-reducing effect of deltorphin II is mediated via PKC activation and mitochondrial Katp, channel opening.

[Phytoadaptogens-induced phenomenon similar to ischemic preconditioning].

The course administration (16 mg/kg per os for 5 days) of extracts of Panax ginseng or Rhodiola rosea induced a decrease in the infarction size/the area at risk (IS AAR) ratio during a 45-min local ischemia and a 2-hr reperfusion in artificially ventilated chloralose-anaesthetized rats. Single administration of ginseng or Rhodiola 24 h before ischemia did not affect the IS/AAR ratio. Chronic administration of Extracts of Eleutherococcus senticosus, Leuzea carthamoides and Aralia mandshurica had no effect on the IS/AAR ratio. Pretreatment with extract ofAralia mandshurica prevented appearance of ventricular arrhythmias during first 10 min coronary artery occlusion. Pretreatment with extract of Rhodiola rosea decreased the incidence of ventricular fibrillation during ischemia. Single administration of extracts of Panax ginseng or Rhodiola rosea in a dose of 16 mg/kg had no effect on the IS/AAR ratio. The authors conclude that extracts of ginseng or Rhodiola exhibit a powerful cardioprotective effect. Extract of Aralia exhibit a strong antiarrhythmic effect. Extracts of ginseng and Rhodiola do not mimic phenomena of ischemia preconditioning.

Antiarrhythmic activity of phytoadaptogens in short-term ischemia-reperfusion of the heart and postinfarction cardiosclerosis.

A course of treatment (16 mg/kg orally during 5 days) by Aralia mandshurica or Rhodiola rosea extracts reduced the incidence of ischemic and reperfusion ventricular arrhythmias during 10-min ischemia and 10-min reperfusion. Extracts of Eleutherococcus senticosus, Leuzea carthamoides, and Panax ginseng did not change the incidence of ischemic and reperfusion arrhythmias. Chronic treatment by aralia, rhodiola, and eleutherococcus elevated the ventricular fibrillation threshold in rats with postinfarction cardiosclerosis. Ginseng and leuzea did not change this parameter in rats with postinfarction cardiosclerosis.

Myocardial resistance to ischemic and reperfusion injuries under conditions of chronic administration of opioid receptor agonists and antagonists.

Chronic treatment with opioid receptor ligands: nonselective peptide opioid receptor agonist dalargin (intraperitoneally in a dose of 1 mg/kg), selective nonpeptide kappa-receptor agonist GR 89696 (subcutaneously in a dose of 0.03 mg/kg), nonselectrive nonpeptide antagonist quadazocine (subcutaneously in a dose of 3 mg/kg) or naltrexone (subcutaneously in a dose of 10 mg/kg) for 20 day had no effect of the incidence of ischemic ventricular arrhythmias and the size of necrotic zone after coronary occlusion and reperfusion in rats in vivo.