Kappa-opioid receptor activation during reperfusion limits myocardial infarction via ERK1/2 activation in isolated rat hearts

Abstract

Background: We investigated whether p42/p44 extracellular signal-regulated kinases (ERK1/2) and/or phosphatidylinositol-3-OH kinase (PI3K)-Akt play a crucial role in cardioprotection by κ-opioid receptor (KOP) activation.

Methods: Langendorff perfused rat hearts were subjected to 30 min of regional ischemia and 2 h of reperfusion. Antagonists of ERK1/2 and PI3K were perfused in hearts treated with the KOP agonist U50488H (U50). Infarct size was measured after 2 h of reperfusion. The phosphorylation states of ERK1/2 and Akt by Western immunoblots were determined. Drugs were perfused for a period of 5 min before and 30 min after reperfusion.

Results: Inhibition of ERK1/2 (26.8 ± 2.9%, P < 0.05 vs. U50) but not PI3K (15.5 ± 1.1%, P > 0.05 vs. U50) completely abrogated the anti-infarct effect of U50488H. Western blot analysis revealed a significant increase in ERK1/2 but not Akt phsophorylation in U50488H-treated hearts as compared to control hearts when measured immediately after reperfusion.

Conclusions: KOP activation effectively reduces myocardial infarction. The anti-infarct effect of U50488H is mediated by the ERK1/2, but not the PI3K-Akt pathway.

Keywords: Coronary occlusion; Heart; Myocardial function; Opioid receptors; Reperfusion.

Fig. 1

Area at necrosis (AN) as a percentage of area at risk (AR), evaluated by triphenyltetrazolium chloride staining following 30 min of occlusion and 2 h reperfusion in an isolated rat heart model. Hearts were subjected to either control (CON), 1 µM U50488H (U50), 15 µM LY294002 (LY), 20 µM PD98059 (PD), or U50 with LY or PD treatment. Each circle represents 1 heart. Horizontal bars depict mean of the group. AN: area of necrosis, AR: area at risk. Values are means ± SEM. ^*P < 0.05 vs. CON.

Fig. 2

Western immunoblotting analysis of phospho- and total-ERK1/2 (Thr²⁰²/Tyr²⁰⁴) in isolated rat hearts. Myocardial samples were obtained from the risk zone at 20 min of ischemia (I-20') and 5 min (R-5') after onset of reperfusion. Bar for R-5' is phosphorylation of proteins at 5 min of reperfusion relative to the value of I-20', expressed as means ± SEM of individual experimental observations. CON indicates control hearts. ^*P < 0.05 vs I-20', ^†P < 0.05 vs. R-5' (CON).

Fig. 3

Western immunoblotting analysis of phospho- and total-Akt (Ser⁴⁷³) in isolated rat hearts. Myocardial samples were obtained from the risk zone at 20 min of ischemia (I-20') and 5 min (R-5') after onset of reperfusion. Bar for R-5' is phosphorylation of proteins at 5 min of reperfusion relative to the value of I-20', expressed as means ± SEM of individual experimental observations. CON indicates control hearts. ^*P < 0.05 vs I-20'.