Acetylcholine, Bradykinin, Opioids, and Phenylephrine, but not Adenosine, Trigger Preconditioning by Generating Free Radicals and Opening Mitochondrial K _ATP Channels

Abstract

Abstract-It has been assumed that all G i -coupled receptors trigger the protective action of preconditioning by means of an identical intracellular signaling pathway. To test this assumption, rabbit hearts were isolated and perfused with Krebs buffer. All hearts were subjected to a 30-minute coronary artery occlusion followed by 120 minutes of reperfusion. Risk area was measured with fluorescent particles and infarct size with triphenyltetrazolium chloride staining. Control hearts showed 29.1Ϯ2.8% infarction of the risk zone. A 5-minute infusion of acetylcholine (0.55 mmol/L) beginning 15 minutes before the 30-minute occlusion resulted in significant protection (9.2Ϯ2.7% infarction). This protection could be blocked by administration of 300 mol/L N-2-mercaptopropionyl glycine (MPG), a free radical scavenger, or by 200 mol/L 5-hydroxydecanoate (5-HD), a mitochondrial K ATP antagonist, for 15 minutes beginning 5 minutes before the acetylcholine infusion (35.2Ϯ3.9% and 27.8Ϯ2.4% infarction, respectively). Similar protection was observed with other known triggers, ie, bradykinin (0.4 mol/L), morphine (0.3 mol/L), and phenylephrine (0.1 mol/L), and in each case protection was completely abrogated by either MPG or 5-HD. In contrast, protection by adenosine or its analog N 6 -(2-phenylisopropyl) adenosine could not be blocked by either MPG or 5-HD. Therefore, whereas most of the tested agonists trigger protection by a pathway that requires opening of mitochondrial K ATP channels and production of free radicals, the protective action of adenosine is not dependent on either of these steps. Hence, it cannot be assumed that all G i -coupled receptors use the same signal transduction pathways to trigger preconditioning.

Post-ischemic early acidosis in cardiac postconditioning modifies the activity of antioxidant enzymes, reduces nitration, and favors protein S-nitrosylation

“…These data indicate that specific antioxidant enzymes (SOD or CAT) can or cannot abrogate PostC-triggering. However, large spectrum antioxidants (NAC or MPG) abolish both preconditioning and PostC protection in several settings [8][9][10]13,22,38,51,54,63]. Although the infused enzymes do not easily enter into the cells, enough SOD enters, at least into endothelial cells, to perturb the redox-environment created by PostC (i.e., SOD down-regulation), thus blocking protection.…”

“…Therefore a vicious cycle is likely to be established in the so-called ROS-induced ROS release (RIRR) [26,27,34]; thus inducing ROS stress and reperfusion injury [for reviews see 46,50]. Survival mechanisms in the heart can be triggered by short, non-lethal periods of ischemia and reperfusion, applied either before (preconditioning) or immediately after (postconditioning, PostC) the index ischemia [3,10,20,25,33,[44][45][46]62]. Both in vivo and in vitro ischemic-and pharmacologicalpreconditioning triggering require protective ROS signaling.…”

“…Both in vivo and in vitro ischemic-and pharmacologicalpreconditioning triggering require protective ROS signaling. In fact, either ROS-generators or ROS-scavengers, given before the index ischemia, trigger or abolish preconditioning [2,7,10,16,23,43,47,48,61]. Therefore, ROS are double edge swords (ROS stress vs ROS signaling).…”

“…This supports the idea that the delayed recovery of intracellular pH during initial reperfusion, is mandatory to limit infarct size in PostC mainly via prevention of mPTP opening and prevention of RIRR [8,9,17,29,30,34,55]. Both in vitro and in vivo studies demonstrated that large spectrum ROSscavengers such as N-acetyl-L-cysteine (NAC) and/or N-(2-mercaptopropionyl)-glycine (MPG) given either during the PostC maneuvers [8,51,54,63] or during AB infusion [8,9,10,13] prevented their protective effects [13,51,54,63]. Moreover, addition of alkaline buffer in early reperfusion abolishes both AB-and PostC-protection [8,9,17,29,55].…”

H2O2opens mitochondrial KATPchannels and inhibits GABA receptors via protein kinase C-ε in cardiomyocytes

“…However, Forbes et al (10) recently demonstrated that diazoxide, a selective mitochondrial K ATP channel opener, generates oxygen radicals and protects ischemic rat hearts. Acetylcholine, bradykinin, phenylephrine, and opioids generate oxygen radicals that were blocked by selective mitochondrial K ATP channel antagonist 5-HD (6,25,38). These data indicate that the mitochondrial K ATP channel is upstream of oxygen radicals in mediating cardioprotection of these agents.…”