KATP Openers Research Articles

K ATP opener-induced delayed cardioprotection: involvement of sarcolemmal and mitochondrial K ATP channels, free radicals and MEK1/2

Previous studies have demonstrated the importance of the sarcolemmal and/or the mitochondrial KATP channel (sKATP and mKATP, respectively) in mediating delayed cardioprotection induced by opioids, bradykinin, adenosine or the KATP-channel opener (KCO), diazoxide. However, the ability of other KCOs, such as the putative sKCO P-1075 or the putative mKCO BMS-191095, to induce delayed cardioprotection has not been tested. In this study, rats were randomly divided and treated with P-1075 (1 μg/kg), BMS-191095 (BMS, 1 mg/kg), diazoxide (3.5 mg/kg) or vehicle. Selective blockers were also administered 5 min prior to KCO or vehicle. These included the sKATP antagonist (HMR-1098 (HMR, 6 mg/kg)), the mKATP antagonist (5-hydroxydecanoic acid (5-HD, 10 mg/kg)), the oxygen radical scavenger (N-2-mecaptopropionyl-glycine (2-MPG, 20 mg/kg)) or the MEK1/2 inhibitor (PD-98059 (PD, 1 mg/kg)). Twenty-four hours later, rats were subjected to 30-min ischemia and 2-h reperfusion, followed by infarct size assessment. P-1075, BMS or diazoxide treatment produced similar reductions in infarct size as compared to vehicle (37.2 ± 2.3%*, 40.7 ± 1.5%*, 35.6 ± 3.6%* vs. 59.8 ± 1.8%, P < 0.05, respectively). HMR administration before P-1075 and diazoxide abolished delayed cardioprotection, but not BMS-induced infarct size reduction (56.1 ± 1.1%, 54.9 ± 3.3%, 41.0 ± 1.7%*, respectively). Administration of 5-HD, 2-MPG or PD prior to the three KATP openers abolished delayed cardioprotection. These data imply BMS-191095 has a selective mitochondrial site of action, perhaps the mKATP channel, whereas diazoxide and P-1075 affect both the sKATP and mKATP channels. All three KCO also require an initial oxygen-derived free radical (OFR) burst and MEK1/2 activation to trigger delayed cardioprotection.

Involvement of K+ channels in adenosine A2A and A2B receptor-mediated hyperpolarization of porcine coronary artery endothelial cells.

This study investigated the effects of the following adenosine agonists: 5;-ethylcarboxamidoadenosine (NECA), N6-cyclopentyadenosine (CPA) 2-[p-(2-carboxyethyl)]phenylamino-5;N-ethylcarboxamidoadenosine (CGS-21680), and 2-chloroadenosine (CAD) and its antagonist, 4-(2-[7-amino-2-[2-furyl]]1,2,4-triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM-241385), a selective A2A adenosine receptor antagonist, and the involvement of the K+ATP and KCa channels on the resting membrane potential (RMP) of confluent monolayers of cultured porcine coronary artery endothelial cells (PCAECs). Adenosine agonists and K+ATP channel openers (pinacidil, cromakalim) hyperpolarized cultured PCAECs. The average RMP was -32.31 +/- 1.2 mV. Adenosine agonists at 10-5 M caused a significant increase in RMP to -65.0 +/- 1.5 mV for CAD (a nonselective adenosine receptor agonist) to -75.9 +/- 1.6 mV for CGS-21680 (a selective A2A receptor agonist) and to -87.0 +/- 3.5 mV for NECA (a nonselective A1/A2A/A2B receptor agonist). Pinacidil and cromakalim at 10 microM increased the membrane potential to -76.2 +/- 1.2 mV and -75.22 +/- 0.12 mV, respectively. The hyperpolarization induced by adenosine receptor agonists and KATP openers was inhibited by an application of the K+ATP channel blocker glibenclamide (10 microM), indicating the involvement of the K+ATP channel in the adenosine-mediated hyperpolarization of PCAECs. Moreover, 1-EB10, a selective opener of the maxi-KCa channel, hyperpolarized PCAECs, and the effect of 1-EB10 was completely blocked by a selective, irreversible blocker of the high conductance KCa (maxi-K) channels (penitrem A), but it only partially blocked the effect of NECA. ZM-241385 has no effect on hyperpolarization elicited by K+ATP and KCa channel openers. However, ZM-241385 significantly blocked the hyperpolarization effect of CAD and CGS-21680. ZM-241385 partially blocked the hyperpolarizing effect of NECA, and a combination of ZM-241385 and penitrem A further blocked the hyperpolarizing effect of NECA. These results further support the involvement of K+ channels in adenosine A2A and A2B receptor-mediated hyperpolarization of PCAECs.

Effect of Adenosine Triphosphate–Sensitive Potassium Channel Openers on Lung Preservation

ATP-sensitive potassium channel (KATP) openers have been proven to be involved in ischemic preconditioning, which protects ischemic tissue. However, the effect of KATP openers on ischemia-reperfusion injury of the lungs remains unknown. We investigated whether a KATP opener, pinacidil, can attenuate ischemia-reperfusion injury using an ex vivo rat lung model. Heart-lung blocks were flushed and preserved with phosphate-buffered saline (control group) or with one of the solutions containing pinacidil (pinacidil group) or pinacidil + glibenclamide (a KATP blocker) (glibenclamide group). The control and glibenclamide groups showed significantly higher values with respect to shunt fraction, pulmonary arterial pressure, and peak inspiratory pressure than the pinacidil group. The concentrations of total adenine nucleotides and ATP in the lung after reperfusion became significantly lower in the control and glibenclamide groups than in the fresh group. Lipid peroxidation of the lungs increased significantly in the control and glibenclamide groups after reperfusion. State 3 mitochondrial respiration and State 3/4 ratios of mitochondrial respiration were significantly decreased in the lungs of the control and glibenclamide groups. These findings suggested that KATP openers would maintain the mitochondrial respiratory function during lung preservation, prevent lipid peroxidation after reperfusion, and attenuate ischemia-reperfusion injury.

Pharmacologic profile of the selective mitochondrial-K(ATP) opener BMS-191095 for treatment of acute myocardial ischemia.

ATP-sensitive potassium channel (K(ATP)) openers as a class protect ischemic myocardium. The protective effects are independent of vasodilator activity and effects on action potential shortening, actions typically associated with sarcolemmal K(ATP) activation. BMS-191095 is a novel mitochondrial K(ATP) opener which protects ischemic myocardium while having no electrophysiologic or vasodilator effects (determined in vitro and in vivo). The cardioprotective effects were determined in isolated rat hearts subjected to ischemia and reperfusion. Protective effects were deduced from increased time to contracture formation during ischemia, improved reperfusion recovery of contractile function, and reduced reperfusion LDH release. The cardioprotective effects of BMS-191095 were observed at concentrations at which this compound selectively opened cardiac mitochondrial K(ATP) channels. This effect was consistent with the pharmacologic profile of this agent. The protective effects were abolished by mitochondrial K(ATP) inhibition. Unlike first-generation K(ATP) openers, BMS-191095 is expected to protect ischemic myocardium with little hemodynamic sequelae and without any proarrhythmic potential. BMS-191095 is potentially useful clinically as a cardioprotective agent. It is also a useful tool for basic research.

K(ATP) channel opening during ischemia: effects on myocardial noradrenaline release and ventricular arrhythmias.

Cardioprotection by K(ATP) channel openers during ischemia is well documented although ill understood. Proarrhythmic effects may be an important drawback. K(ATP) channel modulation influences neurotransmitter release during ischemia in brain synaptosomes. Therefore, we studied the effects of K(ATP) channel modulation on myocardial noradrenaline release and arrhythmias in ischemic rabbit hearts. Isolated rabbit hearts were perfused according to Langendorff and stimulated. Local electrograms were recorded and K+-selective electrodes were inserted in the left ventricular free wall. Cromakalim (3 microM) or glibenclamide (3 microM) was added 20 min prior to induction of global ischemia. After 15, 20, or 30 min of ischemia, hearts were reperfused and noradrenaline content of the first 100 ml of reperfusate was measured. Cromakalim (n = 16) prevented the second rise of extracellular [K(+)] in accordance with its cardioprotective effect. Cromakalim significantly reduced noradrenaline release after 15 min (mean, 169 +/- SEM 97 pmol/gr dry weight vs. control 941 +/- 278; p < 0.05) and 20 min of ischemia (230 +/- 125 pmol/gr dry wt vs. control 1,460 +/- 433; p < 0.05), but after 30 min of ischemia, the difference in noradrenaline release was no longer significant (cromakalim 2,703 +/- 1,195 pmol/gr dry wt vs. control 5,413 +/- 1,310; p = 0.08). Ventricular fibrillation or ventricular tachycardia occurred in 10 of 13 control hearts (77%) (n = 19), in six of 10 glibenclamide-treated hearts (60%) (n = 15), and in six of 14 cromakalim-treated hearts (43%) (p = NS). Cromakalim significantly accelerated onset of ventricular tachycardia or fibrillation (mean +/- SEM onset after 12.5 +/- 1.6 min ischemia vs. control 16.2 +/- 0.7 min; p < 0.05). Noradrenaline release occurred only in cromakalim-treated hearts with early-onset arrhythmias whereas no noradrenaline release was observed in cromakalim-treated hearts without ventricular tachycardia or fibrillation. Our results show that activation of the K(ATP) channel by cromakalim during ischemia reduces myocardial noradrenaline release and postpones the onset of irreversible damage, contributing to the cardioprotective potential of K(ATP) openers during myocardial ischemia.

The role of minoxidil on endogenous opioid peptides in the spinal cord: a putative co-agonist relationship between K-ATP openers and opioids

ATP-gated K + channel openers produce antinociception that is attenuated by opioid receptor antagonists, indicating K-ATP openers produce antinociception, in part, via the release of endogenous opioid peptides. Utilizing the spinal perfusion method, male Sprague–Dawley rats were administered minoxidil intrathecally (i.t.) at doses ranging from 12.5 to 200 μg/rat for 3 min, tested for antinociception using the tail-flick test, and perfused with artificial cerebrospinal fluid (aCSF) to collect endogenous opioid peptides. Endogenous opioid peptide levels were measured by radioimmunoassay. Naltrindole, a δ-opioid receptor antagonist, at 4 mg/kg, subcutaneously (s.c.), blocked minoxidil-induced antinociception. β-Funaltrexamine, a μ-opioid receptor antagonist, at 100 μg/rat, partially blocked minoxidil, whereas the κ-opioid receptor antagonist nor-binaltorphimine, at a dose of 100 μg/rat, did not attenuate minoxidil. Although antagonists of the μ- and δ-opioid receptor attenuated minoxidil-induced antinociception, there was no increase in β-endorphin, an endogenous ligand with affinity for both μ- and δ-opioid receptors or [Leu 5]enkephalin, an endogenous ligand with affinity for δ-opioid receptors.

Effects of potassium channel modulators on myotropic responses of aortic rings of pregnant rats.

The contribution of potassium channels [ATP-sensitive potassium (K(ATP)) and high-conductance calcium-activated potassium (BK(Ca)) channels] in the resistance of aortic rings of term pregnant rats to phenylephrine (Phe), arginine vasopressin (AVP), and KCl was investigated. Concentration-response curves to tetraethylammonium (TEA), a nonselective K(+) channel inhibitor, were obtained in the absence or presence of KCl. TEA induced by itself concentration-dependent responses only in aortic rings of nonpregnant rats. These responses to TEA could be modulated in both groups of rings by preincubation with different concentrations of KCl. Concentration-response curves to Phe, AVP, and KCl were obtained in the absence or presence of cromakalim or NS-1619 (K(ATP) and BK(Ca) openers, respectively) and glibenclamide or iberiotoxin (K(ATP) and BK(Ca) inhibitors, respectively). Cromakalim significantly inhibited the responses to the three agonists in a concentration-dependent manner in both groups of rats. Alternatively, in the pregnant group of rats, glibenclamide increased the sensitivity to all three agonists. NS-1619 also inhibited the response to all agonists. With AVP and KCl, its effect was greater in aortic rings of pregnant than nonpregnant rats. Finally, iberiotoxin increased the sensitivity to all three agents. This effect was more important in aortic rings of nonpregnant rats and was accompanied by an increase of the maximal response to Phe and AVP. These results suggest that potassium channels are implicated in the control of basal membrane potential and in the blunted responses to these agents during pregnancy.

Cardioselective antiischemic ATP-sensitive potassium channel (KATP) openers. 5. Identification of 4-(N-aryl)-substituted benzopyran derivatives with high selectivity.

This paper describes our studies aimed at the discovery of structurally distinct analogs of the cardioprotective KATP opener BMS-180448 (2) with improved selectivity for the ischemic myocardium. The starting compound 6, derived from the indole analog 4. showed good cardioprotective potency and excellent selectivity compared to 2 and the first-generation KATP opener cromakalim (1). The structure-activity studies indicate that increasing the size of the alkyl ester leads to diminished potency as does its replacement with a variety of other groups (nitrile, methyl sulfone). Replacement of the ethyl ester of 6 with an imidazole gave the best compound 3 (BMS-191095) of this series which maintains the potency and selectivity of its predecessor 6. The results described in this publication further support that there is no correlation between vasorelaxant and cardioprotective potencies of KATP openers. Compound 3 is over 20- and 4000-fold more selective for the ischemic myocardium than 2 and cromakalim (1), respectively. The selectivity for the ischemic myocardium is achieved by reduction of vasorelaxant potency rather than enhancement in antiischemic potency. As for cromakalim (1) and 2, the cardioprotective effects of compound 3 are inhibited by cotreatment with the KATP blocker glyburide, indicating that the KATP opening is involved in its mechanism of cardioprotection. With its good oral bioavailability (47%) and plasma elimination half-life (3 h) in rats, compound 3 offers an excellent candidate to investigate the role of residual vasorelaxant potency of 2 toward its cardioprotective activity in vivo.

Pharmacology of ATP-sensitive potassium channel (KATP) openers in models of myocardial ischemia and reperfusion.

Recently, much interest has been focused on the pharmacology of ATP-sensitive potassium channels (KATP) in myocardial ischemia. KATP are thought to be involved with the mechanism of myocardial preconditioning, therefore further increasing the level of interest in these channels. Pharmacologic KATP openers have been shown by numerous investigators to protect ischemic-reperfused myocardium. These agents reduce necrosis, improve postischemic functional recovery, and inhibit contracture formation. These protective effects are abolished by KATP blockers. The cardioprotective effects of KATP openers are independent of vasodilator and cardiodepressant effects, but seem to be mediated by energy conservation (reduced ATP hydrolysis). Action potential shortening is also not correlated with cardioprotection, suggesting a role for intracellular (mitochondrial) KATP. Agents have been developed that retain the glyburide-reversible cardioprotective effects of cromakalim but are devoid of vasodilator and action potential shortening activity. Currently, studies are underway to determine the mechanism of cardioprotection. The potential role of mitochondrial KATP in the pathogenesis of ischemia is discussed in this review article.

Section Review Cardiovascular &amp; Renal: Recent advances in the pharmacology and therapeutic potential of potassium channel openers

Potassium channels constitute the largest group of ion channels, the opening and closing of which result in membrane hyperpolarisation and depolarisation, respectively. This effectively modulates cellular excitability by controlling the open probability of voltage-sensitive calcium and sodium channels. Potassium channel openers are a diverse collection of drugs which act on KATP or BKca. Since both channels are non-inactivating, long-lasting inhibitory effects ensue. The site and mode of action of these drugs is not known for certain but, in the case of the KATP openers, it may be the so-called sulfonyl-urea receptor which is effectively a channel β-subunit. Openers of BKca are in the early stages of scientific study but drugs active at KATP are in various phases of clinical trial. One of these, nicorandil, is currently available for use in angina pectoris and levcromakalim has completed Phase III trials in hypertension. Of other possible applications, their use in the treatment of bronchial asthma and in...

One-day cold perfusion of bimakalim and butanedione monoxime restores ex situ cardiac function.

Bimakalim (Bim), an opener of ATP-sensitive K+ (KATP) channels, was given alone or with 2,3-butanedione monoxime (BDM), a reversible uncoupler of contractility, to protect myocardial function during 1 day of hypothermia. Left ventricular pressure (LVP), coronary flow (CF), percent O2 extraction (%O2E), and cardiac efficiency were measured in 96 isolated, perfused guinea pig hearts divided into seven groups: 1) cold control (no drugs); 2) BDM; 3) Bim; 4) BDM + Bim; 5) BDM + glibenclamide (Glib, a blocker of KATP channels); 6) BDM + Bim + Glib; and 7) time control (6 h warm perfusion only). Drugs were given before, during, and initially after 22 h of low CF at 3.8 degrees C. At 26 h (cold groups) or 4 h (warm group) LVP (mmHg; means +/- SE) was similar for time control (94 +/- 4) and BDM + Bim (92 +/- 4) groups, lower and equivalent in the BDM (65 +/- 7) and BDM + Bim + Glib (64 +/- 7) groups, but LVP was higher than in the Bim group (46 +/- 3), and lowest in the cold control (30 +/- 8) group. In addition, only in the BDM + Bim group were basal CF, %O2E, and cardiac efficiency returned to values obtained in the time control group. Epinephrine increased LVP to that of the time control (106 +/- 3) group only in the BDM + Bim group (106 +/- 3) after hypothermia, and CF increases with adenosine, 5-hydroxytryptamine, and nitroprusside were similar to that of the time control group only in the BDM + Bim group after hypothermia. All of the effects of Bim were reversed by Glib. These results indicate that Bim, given with BDM, effectively preserves myocardial function and metabolism as well as inotropic and vasodilatory reserve during long-term hypothermic preservation as if the 1-day hypothermic state had not been instituted. Because the beneficial effects of Bim are blocked by Glib, the protective effect of Bim likely results from maintained KATP channel opening. Treatment with exogenous KATP openers may prove useful in preserving cardiac function in the transplanted heart.

Treatment of Myocardial Ischemia with ATP-Sensitive Potassium Channel (KATP) Openers

ATP-sensitive potassium channel (KATP) openers are potent antihypertensive agents due to their peripheral vasodilating properties. Although KATP openers have been shown to be direct cardioprotective agents, they carry a certain degree of liability for their use as cardioprotective agents. Their potent coronary and peripheral vasodilating properties can cause complications such as coronary artery steal and hypotension resulting in underperfusion of the tissue already at risk. Also, cardioprotection is not related to action potential shortening and the development of agents devoid of this activity would be desirable. Recent studies indicate that the beneficial effects of myocardial preconditioning might be mediated via opening of the KATP· Therefore, the opening of KATP might constitute an endogenous protective mechanism used by the heart under ischemic stress. Development of therapeutics to mimic this powerful protective mechanism is an attractive approach for the discovery of myocardial protecting agents. In this review, we describe the progress made towards understanding the cardioprotective properties of KATP openers including the discovery of agents that might offer a higher margin of safety for the treatment of myocardial ischemia with this class of agents. We found distinct structure-activity relationships for the cardioprotective and vasorelaxant potencies of KATP openers. Based on the structure-activity relationship studies, we were able to find cardioprotective KATP openers (BMS- 180448) which, despite being equipotent to the first generation agents (e.g., cromakalim) as cardioprotectants, had lower vasorelaxant potencies. The cardioprotecti ve effects of these selective KA TP openers in animal models of myocardial ischemia-reperfusion are also discussed in this review.

Allergic responses reduce the relaxant effect of beta-agonists but not potassium channel openers in guinea-pig isolated trachea

This study was conducted to determine whether the relaxant effect of adenosine triphosphate (ATP)-sensitive potassium channel (KATP) openers on airway smooth muscle are reduced during the hyporesponsiveness of beta-adrenergic receptors. Isometric tension was measured and dose-response curves were constructed for levcromakalim (a KATP opener), Y-26763 (another KATP opener), isoprenaline and theophylline in guinea-pig isolated trachea that was challenged with ovalbumin or pretreated in vitro with either isoprenaline or the KATP openers. Antigen challenge in vitro significantly reduced the potency but not the efficacy of isoprenaline in tracheal strips from actively sensitized guinea-pigs. The concentration of drug that produced a half-maximum inhibition (IC50) was 0.014 +/- 0.004 microM in the challenge group versus 0.006 +/- 0.001 microM in the control group (p < 0.05; n = 9). The IC50 of levcromakalim (1.73 +/- 0.17 microM; n = 6) and of theophylline (110 +/- 3.1 microM; n = 6) were unaffected. Exposure to 4 microM isoprenaline for 30 min evoked beta-adrenergic hyporesponsiveness: the IC50 of isoprenaline rose significantly from 0.010 +/- 0.001 to 0.017 +/- 0.002 microM (p < 0.01; n = 6). No effect was observed on the relaxant actions of levcromakalim and theophylline. In contrast, prior incubation with either levcromakalim or Y-26763 (300 microM for 30 min) significantly reduced the subsequent potency and efficacy of levcromakalim, but did not alter the effects of isoprenaline and theophylline. We conclude that the relaxant effect of the adenosine triphosphate-sensitive potassium channel openers was independent of beta-adrenergic hyporesponsiveness in airway smooth muscle of guinea-pigs.

Myocardial Protection with the ATP-Sensitive Potassium Channel Openers

Abstract: The ATP-sensitive potassium channel (KArP) openers (e.g., Cromakalim) were originally developed for the treatment of hypertension, primarily due to their potent peripheral vasodilating properties. However, the clinical studies have failed to demonstrate any advantage of these compounds over the more established antihypertensive agents. Experimen­ tal studies have shown that the KArP openers may be effective for the treat­ ment of a variety of diseases (asthma, urinary incontinence, ischemia etc.) besides hypertension. However, their clinical utility is limited due to lack of tissue selectivity. We have shown that KArP openers have direct cardioprotective properties which do ,!lOt require contribution from vasodilation. However, the first generation agents offer a narrow window of safety for the treatment of myocardial ischemia due to their potent peripheral vasodilating properties which can compromise the tissue already at risk. Our efforts to find cardioprotective KArP openers with a lower degree of vasorelaxant potency relative to the first generation agents have resulted in the identification of a new class of agents (5). Their structure-activity relationships for antiischemic activity and efficacy in vivo are summarized in this review. The structure-activity studies show that an electron withdrawing group at C6 of the benzopyran ring, a gem-dimethyl at C2, a trans-hydroxyl at C3, an sp3 carbon at C3/C4 and an aryl ring attached to the nitrogen of the cyanoguanidine moiety are required for antiischemic activity. The mechanism of action of these compounds appear to involve opening of the KArP as structurally distinct KATP blockers abolish the cardioprotective effects of these compounds. Unlike the first generation compounds (e.g., cromakalim), the more selective KATP openers can be administered iv to observe antiischemic efficacy. The mechanism of antiischemic activity of KArP openers does not appear to involve the sarcolemmal KArP as their antiischemic activity is independent of action potential shortening. Progress at understanding the molecular mechanism of action of KArP openers and opportunities for future research are also discussed in this article.

The role of ATP-sensitive potassium channels in myocardial ischemia: pharmacology and implications for the future.

Modulators of potassium channels are of great interest for their potential scientific as well as clinical value. These agents may be used for a variety of illnesses including asthma, hypertension, myocardial ischemia, and arrhythmias. The development of KATP openers and blockers has opened a large area of research, particularly on their potential role in the pathogenesis of myocardial ischemia. While much work has shown protective effects for KATP openers, it is unknown whether currently existing agents are optimal. It is also possible that KATP openers may be useful for other types of ischemia such as peripheral vascular disease and cerebral ischemia. It would be exciting to develop agents which not only would protect ischemic myocardium, but also reduce the severity of peripheral and cerebral ischemia. The convergence of the KATP opener studies and the preconditioning area of study was a classical intersection of two seemingly independent lines of research. This convergence has been largely responsible for the heightened interest in KATP. Our quest for knowledge on the role of KATP openers in myocardial ischemia and their potential utility has only just begun.

Cardioselective antiischemic ATP-sensitive potassium channel openers. 4. Structure-activity studies on benzopyranylcyanoguanidines: replacement of the benzopyran portion.

The results of our efforts aimed at the replacement of the benzopyran ring of the lead cardiac selective antiischemic ATP-sensitive potassium channel (KATP) opener (4) are described. Systematic modification of the benzopyran ring of 4 resulted in the discovery of a structurally simpler acyclic analog (8) with slightly lower antiischemic potency than the lead compound 4. Further structure-activity studies on the acyclic analog 8 provided the 2-phenoxy-3-pyridylurea analog 18 with improved antiischemic potency and selectivity compared to the benzopyran-based compound 4. These data demonstrate that the benzopyran ring of 4 and its congeners is not mandatory for antiischemic activity and cardiac selectivity. The results described in this paper also show that, as for the benzopyran class of compounds, the structure-activity relationships for the antiischemic and vasorelaxant activities of KATP openers are distinct. The mechanism of action of the acyclic analogs (e.g., 18) still appears to involve KATP opening as their cardioprotective effects are abolished by pretreatment with the KATP blocker glyburide.

Monophosphoryl Lipid A: A Novel Agent for Inducing Pharmacologic Myocardial Preconditioning.

Myocardial tissue appears to possess an endogenous protective mechanism whereby brief ischemic periods precondition cells to better withstand both reversible and irreversible injury associated with prolonged subsequent ischemic events. Protection develops within minutes of transient ischemia, dissipates within 1-2 hours, and then reappears 12-24 hours following the preconditioning ischemic event. This phenomena, known as ischemic preconditioning (IP), is associated with limitation of infarct size, contractile stunning, and ventricular arrhythmias in postischemic/reperfused hearts. Preconditioned myocardium displays reduced anerobic metabolism. ATPase function, and, hence, improved ATP preservation during ischemia, reduced cytosolic calcium concentrations during reperfusion, and preservation of ultrastructural and myofilament integrity. Efforts to dissect the intracellular signal transduction pathway operative in IP have met with some success. Ischemic preconditioning is associated w ith activation of myocyte Gi protein-coupled receptors such as adenosine and acetylcholine, activation of PKC, production of nitric oxide, and, eventually, opening of ATP-sensitive potassium (KATP) channels. Preconditioning can also be elicited by pharmacologic means using adenosine receptor agonists, activators of PKC, nitric oxide inducers, and KATP openers, among other strategies. Monophosphoryl lipid A (MLA), a nontoxic derivative of the endotoxin pharmacophore lipid A, has been evaluated for cardioprotective activity in numerous preclinical models of cardiac ischemia/reperfusion injury. MLA, when given as a single dose pretreatment in various canine and rabbit models 12-24 hours prior to ischemia, limits infarct size and reduces regional and global contractile dysfunction. Cardioprotection in various models is associated with preservation of ATP during ischemia, enhanced 5'-nucleotidase and adenosine kinase function during reperfusion, and in these aspects mimics ischemic pr econditioning. Priming of KATP channel for enhanced opening during ischemia may be a prerequisite for the cardioprotective activity of MLA and is another feature establishing a similarity between MLA and ischemia induced preconditioning. Efforts continue to further our understanding regarding how MLA may regulate KATP channel and thereby precondition myocardium. Ongoing studies include evaluation of a possible direct effect on the KATP channel, investigation of the ability of MLA to induce a secondary mediator of potassium channel modulation, and evaluation of MLA's ability to phosphorylate the KATP channel as a consequence of kinase activation. Pretreatment with MLA represents a novel method of pharmacologically preconditioning myocardium, displaying a time course for development similar to that of the second window of ischemic preconditioning. Prior clinical experience with MLA indicates that intravenous doses of up to at least 20 µg/kg may be given safely to humans. The drug is c urrently being evaluated in patients undergoing coronary artery bypass engraftment surgery and may prove to be a useful way to protect myocardium from anticipated ischemic events.

Protective Effect of KATP Openers in Ischemic Rat Hearts Treated with a Potassium Cardioplegic Solution

ATP-sensitive potassium channel (KATP) openers directly protect ischemic myocardium, which may make them useful for treating patients undergoing cardiopulmonary bypass, but whether high-potassium-containing cardioplegic solutions would inhibit their protective effects is not clear. We determined whether additional protection greater than that provided by cardioplegia could be found for KATP openers. We studied the effect of 10 microM cromakalim or BMS-180448 pretreatment (10 min before cardioplegia) on severity of ischemia in isolated rat hearts given normothermic or cold St. Thomas' cardioplegic solution (16 mM K+). After cardioplegic arrest, the hearts were subjected to 30-min (normothermic) or 150-min (hypothermic) global ischemia, each followed by 30-min reperfusion. The cardioplegic solutions significantly protected the hearts, as measured by increased time to onset of contracture, enhanced recovery of function, and reduced lactate dehydrogenase (LDH) release. Cromakalim and BMS-180448 both further significantly increased time to contracture in both normothermic and hypothermic arrested hearts; this was accompanied by enhanced recovery of reperfusion contractile function and reduced cumulative LDH release. This additional protective effect of the K ATP openers was abolished by glyburide. Because administration of the K ATP openers only with the cardioplegic solution (1 min before global ischemia) was not efficacious, >1-min pretreatment apparently is necessary. K ATP openers provide additional protection to that afforded by cold or normothermic potassium cardioplegia in rat heart, although the timing of treatment may be crucial.

Glyburide-reversible cardioprotective effect of BMS-180448 is independent of action potential shortening

We determine if action potential duration (APD) shortening and cardioprotection are separable phenomena in ATP-sensitive potassium channel (KATP) openers which protect ischemic myocardium via a glyburide-reversible mechanism. We determined the effect of the weakly vasodilating KATP opener, BMS-180448, and the less cardiac-selective cromakalim, on APD in normal, hypoxic or ischemic guinea pig papillary muscles or isolated hearts and compared their APD effects with their cardioprotective activity in isolated guinea pig hearts. In isolated ischemic guinea pig hearts, cromakalim and BMS-180448 had similar cardioprotective potencies (EC25 of 3.2 and 3.3 microM, respectively, for increasing time to the onset of contracture). At 10 microM, BMS-180448 produced no APD shortening, yet was equally protective at this concentration compared to cromakalim, which produced profound APD shortening under either hypoxic or ischemic conditions. The cardioprotective effects of both compounds at 10 microM were abolished by 0.3 microM glyburide. APD shortening is not correlated with cardioprotective activity for BMS-180448 and cromakalim while their cardioprotective effects are abolished by glyburide. These results suggest the possibility of reduced proarrhythmic activity in some KATP openers and that their cardioprotective activity is not associated with sarcolemmal KATP opening.

K-channel opening activity of ZD6169 and its analogs: effect on 86Rb efflux and 3H-P1075 binding in bladder smooth muscle.

Zeneca ZD6169, (S)-N-(4-benzoylphenyl)-3,3,3-trifluoro- 2-hydroxy-2-methylpropionamide, is a novel compound which relaxes urinary bladder smooth muscle in vitro. The effect of ZD6169 and two of its analogs on 86Rb efflux and 3H-P1075 binding in guinea pig bladder strips was investigated to characterize the K-channel opening properties of this compound. ZD6169 concentration dependently increased the rate of 86Rb efflux from guinea pig bladder strips. 86Rb efflux evoked by ZD6169 and its analogs was blocked by glibenclamide (30 muM) but not by charybdotoxin, apamin or alpha-dendrotoxin, suggesting that this compound activates KATP channels in guinea pig bladder. In addition, interaction of ZD6169 with KATP channels was also confirmed in human bladder smooth muscle cells. Specific binding of 3H-P1075, a potent opener of KATP channels, to guinea pig urinary bladder strips was observed. 3H-P1075 binding was inhibited by known KATP openers. ZD6169 inhibited binding of 3H-P1075 to urinary bladder strips like other structurally different KATP openers, e.g. cromakalim and pinacidil. Potencies for inhibition of 3H-P1075 binding by ZD6169 and other potassium channel openers correlate well with potencies for increase in 86Rb efflux and bladder muscle relaxation studies. It is concluded that Zeneca ZD6169 is a potassium channel opener which activates ATP-sensitive K-channels in guinea pig urinary bladder strips as well as in human bladder cells. Furthermore, binding studies suggest that the effects of ZD6169 and its analogs are mediated by binding to the site labeled by 3H-P1075 in guinea pig bladder strips.