Acta Physiologica Sinica,   April  25, 2003, 55(2): 115-120

Received 2002-10-31

Accepted 2003-02-10

The study was supported by the Research Grants Council,  Hong Kong and a Cardiovascular Physiology Research Fund donated by L.C.S.T.  (Holdings) Ltd.

Corresponding author. Tel: 852-28199194; Fax: 852-28559730; E-mail: wongtakm@hkucc.hku.hk

 

Brief Review

Roles of kappa opioid receptors in cardioprotection against ischemia the signaling mechanisms

Tak Ming WONG*,   Song WU

Department of Physiology,  The University of Hong Kong,  4/F,  Laboratory Block,  Faculty of Medicine Building,  21 Sassoon Road,   Hong Kong SAR,  China

 

Abstract:  There is evidence that the myocytes produce dynorphin and dynorphin-like peptides,  which are kappa opioid receptor (κ-OR) agonists. Activation of κ-OR,  a dominant opioid receptor in the heart,  alters the cardiac function in vivo and in vitro. The observations suggest that the endogenous κ-opioid peptides may act as autocrines or paracrine in regulation of cardiac functions. Myocardial ischemia is a common cause of heart disorders,  which is manifested in decreased myocardial performance,  arrhythmia and infarct. When myocardial ischemia occurs,  the sympathetic discharge increases,  which in turn increases the work-load and oxygen consumption. This exacerbates the situation induced by ischemia. One of the mechanisms with which the body protects against ischemia-induced injury/arrhythmia is inhibition of stimulation of β-adrenoceptor (β-AR),  the receptor mediating the actions of sympathetic stimulation.   κ-Opioids inhibit the β-AR activation. The inhibition of the β-AR activation is due to inhibition of Gs-protein and to a lesser extent the adenylyl cyclase of the signaling pathway mediating β-AR stimulation by a pertussis sensitive G-protein that mediates κ-OR activation. Another mechanism against ischemia-induced injury is preconditioning,  which is defined as prior exposures to ischemia or other insults make the heart more tolerant to subsequent and more severe insults. Protection occurs immediately or 1-3 days after preconditioning. κ-OR mediates protection of preconditioning with ischemia or metabolic inhibition,  one of the consequences of ischemia,  in the heart. Activation of κ-OR by U50488H,  a selective κ-OR agonist (pharmacological preconditioning with U50488H,  UP),  activates protein kinase C (PKC),  opens KATP channels and increases the production of heat shock proteins. Blockade of PKC,  or closing of the KATP channels or inhibition of the synthesis of the heat shock protein abolishes the cardioprotection of UP. The findings indicate the important roles of PKC,  the KATP channels and the heat shock protein in cardioprotection of UP. In addition,  UP also attenuates the Ca2+ overload,  a precipitating cause of cardiac injury,  induced by ischemic insults,  indicating that UP may confer cardioprotection via at least partly  attenuating the Ca2+ overload. Most interestingly,  blockade of the KATP channels with channel blockers,  that abolishes the delayed cardioprotection of UP,  also attenuates the inhibitory effect of UP on Ca2+ overload,  suggesting that the cardioprotective effect of opening of the KATP channels may be due at least partly to the prevention/attenuation of Ca2+ overload.

 

Key words: kappa opioid receptor;   myocardial ischemia;   β-adrenoceptor;  ischemic preconditioning

 

Kappa 阿片受体的抗缺血性心脏保护作用信息机制

黄德明*,  吴淞

香港大学医学院生理系, 香港沙宣道21号

 

摘要:  有证据表明,  心脏细胞产生强腓肽和强腓肽类多肽,  它们是 kappa 阿片受体 (κ-OR)的激动剂。κ-OR是心脏一种优势的阿片受体,  其激活可改变在体和离体心脏的功能。在正常和病理情况下,  内源性κ-阿片肽可能通过自分泌或旁分泌的方式调节心脏功能。心肌缺血是导致心脏功能紊乱的一个常见原因,  主要表现为心肌功能减弱,  心律失常及心肌梗塞等。心肌缺血时, 交感神经发放增强, 从而增加作功负荷及氧消耗量; 而这又使缺血引发的状况更为恶化。机体抵抗缺血引发心肌损害/心律失常的保护机制之一是抑制β-肾上腺素受体 (β-AR) 的兴奋 。κ-OR 确实能抑制β-AR的激动。这种抑制主要是由于GS蛋白受到抑制, 也在较小程度上由于信息通路的腺苷酸环化酶的抑制。因为该种酶能通过对百日咳毒素敏感的G蛋白转导β-AR的激动。另一保护心肌对抗缺血性损害的机制是预处理。预处理是指预先受到缺血等损伤使心脏对随后更严重的损伤产生较强的耐受能力。 这种保护作用可以在预处理后即时产生,  也可延至预处理后1-3天。 在采用缺血或其产生的后果之一代谢抑制作为预处理而致的心脏保护中,  κ-OR 参与媒介预处理的作用。用 κ-OR 的特异性激动剂U50488H激活κ-OR (U50488H 药理性预处理,  UP) 可激活蛋白激酶C (PKC),  开放 ATP 敏感的钾通道(KATP channels)及增加热休克蛋白 (HSP) 的产生。阻断PKC的作用, 关闭KATP通道或抑制HSP的合成, 均可消除UP的心脏保护作用。这些发现表明,  PKC、  KATP通道和HSP在UP的心脏保护中均具重要作用。此外, UP也能减低缺血造成心肌损害的因素之一,  即Ca2+的超负荷。这个事实表明UP发挥心脏保护作用至少部分地是通过减低Ca2+的超负荷。最有趣的是, 以阻断剂阻塞KATP通道, 在消除UP的延迟性心脏保护作用的同时也降低了UP对Ca2+超负荷的抑制作用。这个事实揭示了KATP通道开放所致的心脏保护作用至少部分地可能是由于防止或减低了Ca2+的超负荷。

 

关键词: kappa 阿片受体;  心肌缺血;  β-肾上腺素受体;  缺血预处理

中图分类号: Q463

 

Receptor binding studies showed that kappa-opioid receptor (κ-OR) is a predominant opioid receptor in the heart[1,2].  Functional studies also showed that activation of κ-OR with its selective agonists triggers cardiac responses,  which is blocked by selective κ-OR antagonists in vitro[3-6].The observations indicate that κ-OR may play an important role in the regulation of cardiac function. The presence of dynorphin and dynorphin-like peptides,  which are selective κ-OR agonists,  in the heart[7],  and  the expression of  the mRNA of the precursor prodynorphin of these peptides in the cultured myocytes[8] indicate that the κ-opioid peptides are synthesized in the heart.  The findings suggest that the opioid peptides may play an important role in the regulation of cardiac functions as autocrines and/or paracrines via the κ-OR.

Myocardial ischemia leads to anoxia/hypoxia,  hyperkalaemia,  acidosis and metabolic inhibition,  which in turn triggers inflammatory responses and initiates apoptosis. Myocardial ischemia is a common cause of coronary heart diseases,  which is manifested by decreased myocardial performance,  arrhythmia and myocardial infarct. When myocardial ischemia occurs,  the sympathetic activity is increased,  a response to ischemia-induced stress. Increased sympathetic activity increases the beating rate and contractility of the heart,  which increases the work-load and oxygen consumption. An increase in oxygen consumption at a time when oxygen supply is insufficient exacerbates the damage induced by ischemia. There are mechanisms in the body that counteract the detrimental effects of myocardial ischemia. Of these mechanisms one is inhibition of the sympathetic activity and another cardioprotection of ischemic preconditioning. There is evidence that the κ-opioid peptides and their receptors are involved in both mechanisms. In this article we review the evidence demonstrating the roles of the κ-OR,  and the signaling mechanisms.

Inhibition of sympathetic stimulation

When an electrical stimulation is applied to a myocyte,  like the arrival of an action potential,  the sarcolemmal membrane is depolarized,  which leads to opening of the voltage gated L-type Ca2+ channel,  that allows influx of extracellular Ca2+ into the myocyte. The entry of Ca2+ triggers a massive release of Ca2+ from the sarcoplasmic reticulum (SR),  the intracellular Ca2+ store,  via a Ca2+-induced-Ca2+-release mechanism; the sudden increase in intracellular Ca2+ ([Ca2+]i) triggers contraction. The release of Ca2+ from SR and contraction is enhanced by stimulation of the β-drenoceptor (β-AR). The whole sequence of events could be demonstrated in a single isolated ventricular myocyte in laboratory conditions[9]. In addition it could be demonstrated that shortening of the myocyte in response to electrical stimulation is preceded by a [Ca2+]i transient,  which is enhanced by norepinephrine (NE),  an effect abolished by blockade of β-AR with its antagonist,  propranolol[9].

Fig.1. Inhibition of  β-drenoceptor by κ-opioids receptor during myocardial ischemia.       SNS-sympathetic nervous system; NE-noradrenaline; β-AR-β-drenoceptor;      κ-OR-kappa opioid receptor;    AC-adenylyl cyclase;     Gi/o-Gi/Go proteins;        Gs-Gs protein.

 

In keeping with the well established fact that the effect of β-AR stimulation is mediated via the Gs-protein/adenylyl cyclase (AC) pathway[10],  we also demonstrated that β-AR stimulation increases the cAMP accumulation in the rat ventricular myocyte[11]. When NE was administered together with a κ-opioid receptor (κ-OR) agonist,  U50488H,  at  10-8-10-6  mol/L,  a concentration range which itself has no effect,  the stimulatory effects of NE on electrically stimulated [Ca2+]i transient[9] and cAMP accumulation[11] were significantly reduced. The inhibitory effect of U50488H was abolished by  blockade of  κ-OR  with  a selective κ-OR antagonist,  nor-BNI[9,11].  The observation  indicates that β-AR stimulation is inhibited by κ-OR stimulation, cross-talk between β-AR and κ-OR (Fig.1).  A similar cross-talk between β-AR and κ-OR in rat heart has also been observed[12,13].  It was also shown that U50488H inhibited the effect of activation of Gs-protein with cholera toxin[14]. In addition,  U50488H inhibited slightly,  but significantly,  the effect of activation of AC with forskolin[9]. The observations indicate that the cross-talk results mainly from inhibition of the Gs-protein and to a lesser extent is  also due to inhibition of AC. The cross-talk was abolished by pertussis toxin (PTX)[9,14],  an agent known to inhibit the inhibitory G-proteins. The finding indicates that the inhibition of Gs-protein and AC results from activation of a PTX-sensitive G-protein,  known to mediate the action of κ-OR stimulation[6].

There is evidence suggesting that during myocardial ischemia there is an increased release of opioid peptides from the heart[3]. So we hypothesized that during myocardial ischemia the opioid peptide released may inhibit the effects of increased sympathetic activity,  thus attenuating cardiac arrhythmia. To test the hypothesis,  we induced arrhythmias with NE in the isolated rat heart perfused with a low flow,  a situation,  which mimics myocardial ischemia. We found that U50488H at 10-6 mol/L,  which itself had no effect on cardiac rhythm,  abolished the arrhythmias induced by NE[11]. The effects of U50488H were abolished by nor-BNI. The finding demonstrates that activation of κ-OR protects the heart against ischemia-induced arrhythmias. It is of interest to note that κ-OR agonists,  U50488H or dynorphin at concentrations higher than 10-6 mol/L induces cardiac arrhythmias[3]. It is therefore likely that during myocardial ischemia there is an increased release of the κ-opioid peptides. These peptides may inhibit the β-AR,  thus reducing arrhythmias. However,  if ischemia is severe and prolonged,  excessive amount of κ-opioid peptides may be released,  which may induce arrhythmias.

 Cardioprotection of ischemic preconditioning

In 1986 Murry[15] and co-workers first discovered that brief exposures of a heart to ischemia make the heart more tolerant to subsequent and more severe ischemic insults. This phenomenon is termed cardioprotection of ischemic preconditioning. Subsequent studies showed that preconditioning with one of the consequences of ischemia such as metabolic inhibition[16,17] or other insult such as heat[18,19] confers protection against ischemia and vice versa,  a cross tolerance phenomenon.  There are two windows of the protection,  namely immediate (1-3 h after preconditioning)[15] and delayed (12-72 h after preconditioning)[20]. The clinical implication of protection by preconditioning has aroused great enthusiasm in the research of the mechanisms involved. Up to now receptors to a number of endogenous humoral substances such as adenosine,  catecholamine,  acetylcholine and δ-opioid have been shown to mediate the cardioprotection of preconditioning[21]. We found that the cardioprotection of preconditioning with ischemia[22] or metabolic inhibiton[23] was mimicked by pretreatment with U50488H,  a selective κ-OR agonist,  but antagonized by administration of nor-BNI,  a selective κ-OR antagonist at the time of preconditioning. The observations indicate that κ-OR also mediates cardioprotection of preconditioning. We demonstrated that in vivo (Chen and Wong,  unpublished result) and in vitro[23,24] that prior treatment with a κ-OR agonist,  U50488H (UP),  conferred the same cardioprotection as with ischemia[22] or metabolic inhibition[23].

The signaling mechanism responsible for the immediate cardioprotection of preconditioning has been extensively studied. We found that the immediate cardioprotection of UP was abolished with blockade of either protein kinase C (PKC) or the mitochondrial (mito) KATP channel with selective blockers during preconditioning in the isolated perfused rat heart[22],  indicating that both PKC and mito-KATP channel act to trigger the heart in a preconditioned state,  leading to cardioprotection. The observation is in agreement with the well-established roles of these two messengers[25].

The duration of delayed cardioprotection is longer,  which is clinically more useful. However the signaling mechanism of delayed cardioprotection has not been as well studied as that of immediate cardioprotection. We have delineated the signaling mechanisms of UP,  hoping  to provide more information on the signaling mechanism of delayed cardioprotection of preconditioning.

Fig.2. Signaling mechanism of delayed cardioprotection of pharmacological preconditioning with U50488H against ischemic insult in cardiomyocytes.  κ-OR-kappa opioid receptor;  HSP 70-heat shock protein 70;  IP-ischemic preconditioning; PKC-protein kinase C;  KATP channels-ATP sensitive potassium channels.

 

Similar to immediate protection,  the delayed cardioprotection of preconditioning with metabolic inhibition (MIP) or with U50488H was abolished when an inhibitor of PKC was administered at the time of preconditioning[23]. The observation indicates that activation of PKC triggers the signaling mechanisms. In a subsequent study we found that both MIP and UP,  that conferred delayed cardioprotection,  induced an increased expression of PKC-ε and that blockade of the PKC isoform with a selective inhibitor,  εV1-2,  at the time of preconditioning,  abolished not only the increased expression of PKC-ε,  but more importantly delayed cardioprotection of preconditioning[26]. The observation indicates that PKC-ε is a trigger of delayed cardioprotection of MIP and UP. This is in agreement with the previous finding that PKC-ε triggers delayed cardioprotection of preconditioning with ischemia[27,28]. It seems that this PKC isoform is a common trigger of delayed cardioprotection of preconditioning of different kinds.

While the mito-KATP channel is widely believed to play an important role in cardioprotection of preconditioning[25,  29-31],  the role of sarcolemmal (sarc) KATP channel is controversial[32-34]. Recently we found that intravenous administration of U50488H to rats led to a reduction in infarct induced by ischemia 24 h later (Chen and Wong,  unpublished result),  confirming that UP confers delayed cardioprotection demonstrated in an in vitro isolated ventricular myocyte preparation[23]. The infarct sparing effect of UP was attenuated when either of the two channels was blocked by their selective blockers,  namely 5-HD,  a selective blocker of mito-KATP channel or HRM-1098,  a selective blocker of sarc-KATP channel,  at the time of preconditioning (Chen and Wong,  unpublished result),  indicating that both channels act as a trigger of delayed cardioprotection of UP. On the other hand,  blockade of mito-KATP channel,  but not sarc-KATP channel,  before ischemia abolishes the delayed cardioprotection of UP (Chen and Wong,  unpublished result),  indicating that mito-KATP channel,  but not sarc-KATP channel,  is also a mediator/end-effector of cardioprotection of UP. This is in agreement with the previous observation[35].

Heat shock proteins are known to play an important role in cardioprotection[20]. We also found that both UP and MIP increased the expression of an inducible heat shock protein 70 in the heart. More importantly,  we found that blockade of synthesis of the protein with a selective antisense also blocked the delayed protection[36]. The observations indicate a  mediating role of this protein.

It has been shown that cardiac injury is preceded by an intracellular Ca2+ ([Ca2+]i) overload upon ischemia[37,38]. We also found an increased [Ca2+]i following metabolic inhibition[24]. More importantly we found that pretreatment with an intraperitoneal injection of BAPTA-AM,  a Ca2+ chelate,  attenuated the infarct size induced by subsequent more sever myocardial ischemia and reperfusion in the rat (Yan and Wong,  unpublished observation). The observations confirmed the belief that [Ca2+]i overload is a precipitating cause of injury. Recently we observed that intravenous administration of U50488H into the rat conferred delayed cardioprotection against ischemic insults and that the delayed cardioprotection was accompanied by attenuation of [Ca2+]i overload induced by ischemic insults (Chen and Wong,  unpublished results). The observation suggests the delayed cardioprotection of UP may result,  at least partly,  from attenuation of Ca2+ overload induced by ischemic insults. Similar observations have also been reported in immediate cardioprotection of UP against ischemic insults[24]. 

Interestingly,  blockade of mito- or sarc-KATP channels during UP or mito-KATP channel before ischemic insults,  that abolished the delayed cardioprotection of UP,  also suppressed  the attenuating effect of UP on [Ca2+]i overload (Chen and Wong,  unpublished results). The observation suggests that the cardioprotective effect of the KATP channels may result at least partly from attenuation of Ca2+ overload (Fig.2).

 Conclusion

When myocardial ischemia occurs there may be an increased release of κ-opioid peptides in the heart,  that would inhibit the harmful action of increased sympathetic   activity via activating the κ-OR. Ischemia may also trigger another protective mechanism,  cardioprotection of preconditioning,  that confers protection when the heart is exposed to a more severe ischemic insult again. κ-OR is one of the receptors that mediate the protection of preconditioning. PKC,  KATP channels,  heat shock protein and intracellular Ca2+ are all involved.

 

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