G. J. Gross

Effects of the KATP channel opener bimakalim on coronary blood flow, monophasic action potential duration, and infarct size in dogs.

BACKGROUND The major purpose of the present study was to determine the effect of the potassium channel opener bimakalim, administered intracoronary only during the initial 10 minutes of ischemia, on myocardial infarct size in anesthetized dogs. A second aim was to test the possibility that bimakalim mediates its cardioprotective effects by accelerating the rate of myocyte action potential shortening during early ischemia. A third aim was to determine the relative potency of bimakalim to open coronary vascular ATP-regulated potassium (KATP) channels versus myocyte KATP channels. METHODS AND RESULTS Barbital-anesthetized open-chest dogs were used. In the initial studies, bimakalim (0.1 to 10 micrograms/min) was infused into the left anterior descending coronary artery (LAD), and changes in coronary blood flow and monophasic action potential duration (MAPD) were used as indexes of coronary vascular KATP channel and myocyte KATP channel activity, respectively. In subsequent infarct studies, dogs were subjected to 60 minutes of LAD occlusion followed by 4 hours of reperfusion. Based on preliminary studies, two doses of bimakalim that did not shorten MAPD during nonischemic conditions (0.1 and 0.3 microgram/min) and one that markedly shortened MAPD during nonischemic conditions (3.0 micrograms/min) or an equal volume of vehicle were infused into the LAD during the initial 10 minutes of coronary artery occlusion. Transmural myocardial blood flow was measured at 5 and 30 minutes of occlusion by the radioactive microsphere technique, and infarct size was determined at the end of 4 hours of reperfusion by triphenyltetrazolium staining. The monophasic action potential duration at 50% repolarization (MAPD50) was measured by an epicardial probe placed in the center of the ischemic area. Bimakalim had an approximately 10-fold greater affinity for the coronary vascular than the myocardial KATP channel (ED50 coronary, approximately 0.3 microgram/min; ED50 myocyte, approximately 3.0 micrograms/min). Three doses of bimakalim (0.1, 0.3, and 3.0 micrograms/min) all markedly reduced infarct size expressed as percent of the area at risk (12.6 +/- 3.3%, 14.5 +/- 2.2%, and 14.2 +/- 5.3%, respectively, versus 27.2 +/- 5.7% in controls) to nearly equal extents. Subsequently, we found that the two higher doses of bimakalim (0.3 and 3.0 micrograms/min) markedly accelerated yet the 0.1-microgram/min dose of bimakalim did not significantly affect the ischemia-related shortening of the action potential during the initial 5 minutes of occlusion. In addition, 0.1 and 0.3 microgram/min bimakalim did not increase the incidence of ventricular fibrillation during the 60 minutes of occlusion (0 of 7 and 0 of 8 dogs, respectively), whereas 3.0 micrograms/min bimakalim had a profibrillatory effect (6 of 6) compared with the control group (1 of 8). There were no significant differences between groups in systemic hemodynamics, myocardial oxygen demand, ischemic bed size, or collateral blood flow to the ischemic region. CONCLUSIONS The results of the present study clearly reveal that a small dose (0.1 or 0.3 microgram/min) of the KATP channel opener bimakalim administered only during the initial 10-minute period of ischemia markedly reduced myocardial infarct size to an extent equal to that of a higher profibrillatory dose in barbital-anesthetized dogs. These data also suggest that bimakalim and other potassium channel openers may partially exert their cardioprotective effects by accelerating KATP channel activation during early ischemia as evidenced by an enhanced rate of ischemic myocyte action potential shortening. However, the results also suggest that other cellular mechanisms may be involved in mediating the cardioprotection produced by a low dose of bimakalim (0.1 microgram/min) because it did not significantly accelerate the shortening of the action potential duration, yet it had an efficacy to redu BACKGROUND The major purpose of the present study was to de

A comparison of adenosine-induced cardioprotection and ischemic preconditioning in dogs. Efficacy, time course, and role of KATP channels.

BACKGROUND Adenosine has been proposed to be an important mediator of ischemic preconditioning. Intracoronary administration of adenosine has recently been shown to mimic the effects of preconditioning in isolated rabbit hearts. However, it is not known whether this agent can duplicate the effects of preconditioning in vivo or in other species. Thus, the first objective of the present study was to determine whether adenosine can limit myocardial necrosis to the same extent as preconditioning in anesthetized dogs. A second objective was to determine whether the duration of the adenosine-induced cardioprotection persisted as long as that of ischemic preconditioning. Finally, a third aim was to determine whether adenosine mediates its cardioprotection via the KATP channel, which has been shown to be an important mediator of preconditioning in several animal species, including dogs. METHODS AND RESULTS Barbital-anesthetized open-chest dogs were subjected to 60 minutes of left anterior descending coronary artery (LAD) occlusion followed by 4 hours of reperfusion. Preconditioning was elicited by 10 minutes of LAD occlusion followed by 10 or 60 minutes of reperfusion before the 60-minute occlusion period. Adenosine (400 micrograms/min) or an equivalent volume of saline was infused into the LAD for 10 minutes, followed by a 10- or 60-minute drug-free period before the 60-minute ischemic insult. Glibenclamide (0.3 mg/kg i.v.), a selective KATP channel blocker, was given 15 minutes before adenosine administration, and another selective KATP channel blocker, 5-hydroxydecanoate (5-HD, 3 mg/min IC) was infused concomitantly with adenosine into the LAD for 10 minutes. Transmural myocardial blood flow was measured at 5 minutes of occlusion, and infarct size was determined by triphenyltetrazolium staining and expressed as a percent of the area at risk (AAR). There were no significant differences in hemodynamics, collateral blood flow, or AAR between groups. Preconditioning with either 10 or 60 minutes of reperfusion produced a marked reduction (P < .05) in infarct size (6.7 +/- 2.5% and 8.7 +/- 2.6%, respectively, versus 26.9 +/- 4.3% in controls). Administration of adenosine with a 10-minute drug-free period before 60 minutes of occlusion resulted in a marked decrease in infarct size similar to that seen with preconditioning (9.6 +/- 1.7% versus 26.9 +/- 4.3% in controls); however, the protection disappeared when a 60-minute drug-free period was allowed after adenosine administration (23.0 +/- 2.4% versus 26.9 +/- 4.3% in controls). In addition, treatment with either glibenclamide or 5-HD completely abolished the protective effects of adenosine (26.4 +/- 6.8 and 25.0 +/- 4.4%, respectively, versus 26.9 +/- 4.3% in controls). CONCLUSIONS These results clearly reveal that (1) a 10-minute intracoronary infusion of adenosine exhibits the same efficacy as ischemic preconditioning in reducing myocardial necrosis in dogs; (2) similar to preconditioning, adenosine mediates its cardioprotection via a cardiac KATP channel-linked mechanism; and (3) adenosine-induced cardioprotection is transient (disappearing within 60 minutes), whereas ischemic preconditioning persists for at least 60 minutes. These data support the hypothesis that endogenous adenosine released during ischemia is an important mediator of ischemic preconditioning; however, important differences exist between the time course of effects of exogenously administered adenosine and preconditioning, which suggests that other factors may also be involved.

The role of mitochondrial KATP channels in cardioprotection.

Ischemic Preconditioning (IPC) is a phenomenon in which single or mutiple short periods of ischemia or hypoxia (21) result in a marked reduction in infarct size, the extent of stunning and the incidence of cardiac arrhythmias following a more prolonged period of ischemia. Although a number of mediators and signaling molecules have been proposed to mediate this phenomenon, opening of the adenosine triphosphate sensitive potassium channel (KATP channel) seems to be a common effector molecule involved in IPC. In addition, KATP channel openers have been shown to produce beneficial effects in a number of models of ischemia/reperfusion injury (12). Currently there have been two KATP channels identified in the heart (13), the sarcolemmal channel (sarc KATP) and the more recently discovered mitochondrial channel (mito KATP). There is currently some controversy concerning the relative importance of these two channels in mediating the cardioprotective effect of IPC and following the administration of KATP openers (13). Therefore, it is the purpose of this review to discuss the evidence which supports a role for the mito KATP channel in IPC and cardioprotection in the presence of KATP openers.

ATP-sensitive potassium channels and myocardial preconditioning

The ATP-sensitive potassium channel (KATP) has been shown to serve an endogenous cardioprotective role in a number of experimental models of myocardial stunning and infarction. More importantly, a majority of evidence has also been obtained which suggests that the KATP channel may be intimately involved in both triggering and maintaining the cardioprotection afforded by the phenomenon of ischemic preconditioning particularly in large animal models such as dogs and pigs. Although the evidence for an involvement of KATP in ischemic pre-conditioning is equivocal in smaller animal species such as rabbits and rats, activation of this channel by KATP channel openers produces cardioprotection in all species studied. Whether this channel is an important mediator of ischemic preconditioning in all animal species including man and the mechanism by which this cardio-protective effect is obtained await further experimental studies. Nevertheless, the use of selective potassium channel openers to mimic preconditioning in selected clinical settings may be a desirable future therapeutic goal.

Glibenclamide antagonizes adenosine A1 receptor-mediated cardioprotection in stunned canine myocardium.

BackgroundThe main objective of the present study was to determine the role of adenosine in the development of myocardial stunning following multiple, brief periods of coronary artery occlusion as well as the subtype of adenosine receptor (A1 A2 or) involved. A second objective was to determine if there was an interaction between the adenosine A1 receptor and the ATP-dependent K channel (KATP). Methods and ResultsThe effects of the selective adenosine A1 receptor antagonist 8-cyclopentyl-1,3- dipropyIxanthine (DPCPX) and agonist cyclopentyladenosine (CPA), the selective A2 receptor agonist CGS 21680, and the KATP channel blocker glibenclamide on myocardial stunning produced by repetitive coronary artery occlusions were studied in barbital-anesthetized dogs. Regional segment function was measured with sonomicrometry. Under control conditions, six 5-minute periods of coronary occlusion interspersed with 10-minute periods of reperfusion and ultimately followed by 2 hours of reperfusion produced regional segment dysfunction. Pretreatment with intravenous infusion of CPA (2.0

Mechanism of coronary vasodilation produced by bradykinin.

μg kg−1 min−1) improved percent segment shortening throughout reperfusion, whereas pretreatment with DPCPX (1.0 mg/kg i.v. bolus) significantly worsened the recovery of postischemic contractile function. In contrast, neither DPCPX nor CPA had any effect on the recovery of contractile function when administered before the second coronary occlusion. Furthermore, pretreatment with CGS 21680 (0.2

The A3 adenosine receptor agonist CP-532,903 [N6-(2,5-dichlorobenzyl)-3'-aminoadenosine-5'-N-methylcarboxamide] protects against myocardial ischemia/reperfusion injury via the sarcolemmal ATP-sensitive potassium channel.

μg kg1 min-1) did not affect the recovery of percent segment shortening. In addition, pretreatment with a low dose of glibenclamide (0.1 mg/kg) had no effect on percent segment shortening by itself but completely abolished the beneficial effect of CPA. Importantly, the effects of the various agents on percent segment shortening were independent of difference in systemic hemodynamics, collateral blood flow, or ischemic bed size. ConclusionsThese results suggest that stimulation of myocardial adenosine A1 receptors, particularly when induced by the initial coronary artery occlusion, is cardioprotective during repetitive, brief periods of coronary artery occlusion and that these beneficial actions may be partially mediated via a glibenclamide-sensitive mechanism, possibly opening of myocardial KATP channels.

Changes in ischemic blood flow distribution and dynamic severity of a coronary stenosis induced by beta blockade in the canine heart.

BackgroundBradykinin has been demonstrated to be an endothelium-dependent vasodilator in the cerebral circulation of the mouse, but the actions of bradykinin on regional tissue perfusion in the canine coronary circulation have not been studied. Methods and ResultsThe mechanism of coronary vasodilation by bradykinin was studied in open-chest, anesthetized dogs. The role of cyclooxygenase stimulation, bradykinin B2 receptor activation, and endothelium-derived relaxing factor in bradykinin-mediated vasodilation was studied in separate groups of dogs. Bradykinin was infused intracoronarily so as to avoid changes in systemic hemodynamics capable of altering the regional distribution of coronary blood flow (radioactive microspheres). Bradykinin produced a preferential increase in subendocardial blood flow. Pretreatment with indomethacin had no effect on bradykinin-mediated increases in total left ventricular flow or the transmural distribution of coronary blood flow. Blockade of bradykinin B2 receptors with the competitive antagonist [This58,D-Phe7]-bradykinin attenuated both the increase in total flow and redistribution of perfusion to the subendocardium produced by bradykinin. Inhibition of endothelium-derived relaxing factor with quinacrine, occlusion/reperfusion, or NG-monomethyl L-arginine attenuated the total increase in left ventricular flow and blocked the redistribution of flow to the subendocardium produced by bradykinin. ConclusionsThe present results demonstrate that intracoronary infusion of bradykinin produces a preferential increase in blood flow to the subendocardium via stimulation of B2 receptors and the release of an endothelium-dependent relaxing factor that may be nitric oxide. (Circulation 1991;83:2048—2056)

The leukocyte cell adhesion cascade and its role in myocardial ischemia-reperfusion injury.

We examined the cardioprotective profile of the new A3 adenosine receptor (AR) agonist CP-532,903 [N6-(2,5-dichlorobenzyl)-3′-aminoadenosine-5′-N-methylcarboxamide] in an in vivo mouse model of infarction and an isolated heart model of global ischemia/reperfusion injury. In radioligand binding and cAMP accumulation assays using human embryonic kidney 293 cells expressing recombinant mouse ARs, CP-532,903 was found to bind with high affinity to mouse A3ARs (Ki = 9.0 ± 2.5 nM) and with high selectivity versus mouse A1AR (100-fold) and A2AARs (1000-fold). In in vivo ischemia/reperfusion experiments, pretreating mice with 30 or 100 μg/kg CP-532,903 reduced infarct size from 59.2 ± 2.1% of the risk region in vehicle-treated mice to 42.5 ± 2.3 and 39.0 ± 2.9%, respectively. Likewise, treating isolated mouse hearts with CP-532,903 (10, 30, or 100 nM) concentration dependently improved recovery of contractile function after 20 min of global ischemia and 45 min of reperfusion, including developed pressure and maximal rate of contraction/relaxation. In both models of ischemia/reperfusion injury, CP-532,903 provided no benefit in studies using mice with genetic disruption of the A3AR gene, A3 knockout (KO) mice. In isolated heart studies, protection provided by CP-532,903 and ischemic preconditioning induced by three brief ischemia/reperfusion cycles were lost in Kir6.2 KO mice lacking expression of the pore-forming subunit of the sarcolemmal ATP-sensitive potassium (KATP) channel. Whole-cell patch-clamp recordings provided evidence that the A3AR is functionally coupled to the sarcolemmal KATP channel in murine cardiomyocytes. We conclude that CP-532,903 is a highly selective agonist of the mouse A3AR that protects against ischemia/reperfusion injury by activating sarcolemmal KATP channels.

Criteria for a mediator or effector of myocardial preconditioning: Do KATP channels meet the requirements?

The effects of equipotent β1‐receptor‐blocking doses of propranolol, metoprolol and sotalol on distal coronary pressure, stenosis resistance and regional myocardial blood flow (endo/epi) were studied in anesthetized dogs with a severe noncircumferential stenosis of the left circumflex coronary artery. No significant differences between the three β blockers were observed for overall hemodynamics and regional myocardial blood flow. After drug treatment, subendocardial blood flow (0.47 ± 0.05 to 0.78 ± 0.05 ml/min/g) and endo/epi (0.67 ± 0.04 to 1.18 ± 0.04) increased significantly (p < 0.05) in the ischemic region. These changes were associated with a marked increase in distal coronary perfusion pressure and a decrease in heart rate. Resistance across the stenosis decreased significantly (p < 0.05) after, β‐receptor blockade (3.2 ± 0.3 to 1.4 ± 0.2 units). Atrial pacing to control heart rate only partially attenuated these changes. These results suggest that a favorable redistribution of ischemic blood flow after β blockade is the result of an increase in distal diastolic pressure‐time index and an autoregulation‐induced increase in distal bed vascular resistance due to a decrease in myocardial oxygen demand associated with β blockade. The latter effect also resulted in a decrease in the dynamic severity of a proximal coronary stenosis.