mitoKATP Channel Research Articles

Serofendic Acid, a Substance Extracted from Fetal Calf Serum, as a Novel Drug for Cardioprotection

We previously identified a novel endogenous substance, serofendic acid, from a lipophilic extract of fetal calf serum. The compound, a low-molecular-weight sulfur-containing atisane-type diterpenoid, exhibited potent protective action against neurotoxicity induced by glutamate, nitric oxide, and oxidant stress. We investigated whether this substance has a cardioprotective effect. Primary cultures of neonatal rat cardiac myocytes were exposed to oxidant stress (H2O2) to induce cell death. Pretreatment with serofendic acid significantly suppressed cell death induced by H2O2, and the cytoprotective effect was closely associated with the preservation of mitochondrial function. Serofendic acid inhibited H2O2-induced loss of mitochondrial membrane potential in a concentration-dependent manner (with saturation by 100 microM), by attenuating matrix calcium overload and intracellular accumulation of reactive oxygen species. The protective effect of serofendic acid was comparable to that of a mitochondrial ATP-sensitive potassium (mitoK ATP) channel opener, diazoxide. Furthermore, mitoK ATP channel blocker, 5-hydroxydecanoate, abolished the protective effect of serofendic acid. Serofendic acid and diazoxide, administered together, at 100 microM each, had no additive effects. Thus, serofendic acid inhibited the oxidant-induced mitochondrial death pathway, presumably through activation of the mitoK(ATP) channel. In conclusion, serofendic acid appears to protect cardiac myocytes from oxidant-induced cell death by preserving the functional integrity of mitochondria. Our findings suggest that serofendic acid may represent a novel candidate for cardioprotective therapy in ischemia/reperfusion injury.

Opening of microglial KATP channels inhibits rotenone‐induced neuroinflammation

As activated microglia (MG) is an early sign that often precedes and triggers neuronal death, inhibition of microglial activation and reduction of subsequent neurotoxicity may offer therapeutic benefit. The present study demonstrates that rat primary cultured MG expressed Kir6.1 and SUR2 subunits of KATP channel, which was identical to that expressed in BV-2 microglial cell line. The classic KATP channel opener pinacidil and selective mitochondrial KATP (mito-KATP) channel opener diazoxide prevented rotenone-induc microglial activation and production of pro-inflammatory factors (tumour necrosis factor[TNF]-α and prostaglandin E2[PGE2]). And the effects of pinacidil and diazoxide were reversed by mito-KATP blocker 5-hydroxydecanoate (5-HD), indicating that mito-KATP channels participate in the regulation of microglial activation. Moreover, the underlying mechanisms involved the stabilization of mitocho drial membrane potential and inhibition of p38/c-Jun-N-terminal kinase (JNK) activation in microglia. Furthermore, the in vivo study confirmed that diazoxide exhibited neuroprotective effects against rotenone along with the inhibition of microglial activation and neuroinflammation. Thus, microglial mito-KATP channel might be a novel prospective target for the treatment of neuroinflammation-related degenerative disorders such as Parkinson's disease.

Abstract 1554: PX-18 - A Novel Inhibitor of Type IIA Secretory Phospholipase A 2 Induces Pre and Post-Conditioning against Myocardial Infarction in Rabbit: Role of Mitochondrial K ATP Channels.

Background: The low-molecular-weight, calcium-dependent type IIA secretory phospholipase A 2 (sPLA 2 ) has been implicated in irreversible cell damage following ischemia-reperfusion (I/R) injury. We examined the protective effect of PX-18, a novel sPLA 2 inhibitor, during I/R in the heart. Since mitochondrial K ATP (mitoK ATP ) channel opening is an essential component of ischemic tolerance induced by preconditioning in hearts, we also interrogated the role of these channels in PX-18-induced protection. Methods and Results: Rabbits were treated with PX-18 (60 mg/kg ip) 30 min prior to 30 min of regional ischemia and 3 hrs of reperfusion. MitoK ATP channel blocker 5-hydroxydecanoate (5-HD, 5 mg/kg iv) was given 10 min before I/R. Infarct size (IS) was measured by computer morphometry of tetrazolium stained sections. IS (mean ± SE) was reduced in rabbits treated with PX-18 as compared to vehicle or non-treated controls (43% reduction)[Fig.1 ]. 5-HD blocked the protection of PX-18 as shown by an increase in IS. 5-HD alone had no effect on IS. Risk areas were not different among groups (not shown). Furthermore, PX-18 administration as an iv bolus (30 mg/kg) further reduced IS, both when given post-ischemia (55% reduction), and, importantly, when given 1 hr after reperfusion (61% reduction) [Fig. 1 ]. PX-18 had no significant effect on hemodynamics. Conclusion: These data suggest that PX-18 induces pharmacological preconditioning in the heart which is mediated by opening of mitoK ATP channels. Also, the drug was cardioprotective when administered after ischemia or 1 hr after reperfusion. We conclude that inhibition of sPLA2 is a powerful strategy to induce protection against I/R injury.

HYPEROXIA CONFERS MYOCARDIAL PROTECTION IN MECHANICALLY VENTILATED RATS THROUGH THE GENERATION OF FREE RADICALS AND OPENING OF MITOCHONDRIAL ATP‐SENSITIVE POTASSIUM CHANNELS

1. One hour exposure to hyperoxia has been shown previously to limit a subsequent ischaemia-reperfusion injury in spontaneously breathing rats. We tested the cardioprotective effect of a shorter period of hyperoxia during mechanical ventilation and the possible contribution of reactive oxygen species (ROS) and mitochondrial ATP-sensitive potassium (mitoK(ATP)) channels. 2. Mechanically ventilated rats were exposed to normoxia (Fi O2 = 0.3) or hyperoxia (Fi O2 = 1.0) for 30 min and pH, P CO2, PO2, heart rate, airway and blood pressure were measured at baseline and after 30 min mechanical ventilation. Isolated hearts were subsequently subjected to 30 min ischaemia and 120 min reperfusion. Infarct size and left ventricular end-diastolic pressure (LVEDP), developed pressure (LVDP) and coronary flow (CF) were measured. In order to investigate the role of ROS and KATP channels within the mechanism leading to cardioprotection, the free radical scavenger N-acetylcysteine (NAC; 150 mg/kg) was infused in mechanically ventilated rats and the KATP channel blockers glibenclamide (200 mmol/L) or 5-hydroxydecanoate (10 mmol/L) were infused in isolated hearts immediately before ischaemia. 3. No differences were detected in P CO2, pH, heart rate, airway and blood pressure between the groups. However, the PO2 in hyperoxic groups was significantly higher compared with that in normoxic groups (P < 0.01). After 30 min ischaemia, we found that hyperoxic preconditioning significantly improved CF (P < 0.01), LVDP (P < 0.01) and LVEDP (P < 0.01) and reduced the extent of infarct size in the reperfused heart compared with the normoxic group (P < 0.01). When rats were pretreated either with NAC before hyperoxic ventilation or with K(ATP) channel blockers before ischaemia, myocardial protection was abolished. 4. Hyperoxic mechanical ventilation, prior to ischaemia, reduces myocardial reperfusion injury. This is likely to occur through the induction of oxidative stress, which leads to myocyte mitoKATP channel opening.

The Cardioprotective Effects of Levosimendan: Preclinical and Clinical Evidence

Levosimendan, a drug used in the treatment of acute and decompensated heart failure, has positive inotropic and antistunning effects mediated by calcium sensitization of contractile proteins, and vasodilatory and antiischemic effects mediated via the opening of ATP-sensitive potassium channels in vascular smooth-muscle cells. Recently, it also has been shown to act on mitochondrial ATP-sensitive potassium (mitoKATP) channels, an action thought to protect the heart against ischemia-reperfusion damage. This finding has suggested a possible application for levosimendan in clinical situations in which preconditioning would be beneficial (eg, in pre- and perioperative settings in cardiac surgery). The demonstration that levosimendan can prevent or limit myocyte apoptosis via the activation of mitoKATP channels provides a potential mechanism whereby this agent might protect cardiac myocytes during episodes of acute heart failure. This finding may explain why short-term treatment with levosimendan may improve longer-term survival. The present article reviews the literature on the cardioprotective actions of levosimendan, with particular emphasis on its recently recognized effects on mitoKATP channels and the putative preconditioning effects of that action. A therapeutic approach to acute heart failure that includes a cardioprotective strategy could have a clinically meaningful benefit on disease progression beyond alleviation of symptoms.

Morphine Postconditioning Protects Against Reperfusion Injury in the Isolated Rat Hearts

Postconditioning is a novel strategy of attaining cardioprotection. Previous studies have suggested morphine mimics the effects of ischemic preconditioning. Whether it is also capable of producing postconditioning or not is still unclear. The purpose of this study was to determine (1) whether morphine postconditioning (MPostcond) would protect the heart against reperfusion injury and the subtype(s) of opioid receptors (OR) involved, (2) whether combining MPostcond with morphine preconditioning (MPC) would afford additive cardioprotection, and (3) to evaluate the role mitochondrial adenosine triphosphate-sensitive potassium (mito-K atp) channel played in MPostcond. Isolated perfused rat hearts were subjected to 45 min of ischemia followed by 1 h of reperfusion. First, three morphine concentrations (0.3, 3.0 and 30 microM) were used to study the protective effect of MPostcond. Second, the effect of blockade of OR subtypes by three antagonists (nonselective OR antagonist naloxone, kappa-OR antagonist nor-binaltorphimine, and delta-OR antagonist naltrindole) on MPostcond was investigated. Third, the protective effects of MPC, MPostcond and the combining MPC with MPostcond on reperfusion injury were compared. Last, the effect of blockade of mito-K atp by 5-hydroxydecanoate on MPostcond was studied. MPostcond was induced by a 10-min perfusion of morphine in Krebs-Ringer's solution performed at the onset of reperfusion, and MPC was produced by a 20-min perfusion of morphine 10 min before ischemia. Infarct size (IS/AAR, as a percentage of the area at risk) was determined by 2,3,5-triphenyltetrazolium staining. IS/AAR was significantly reduced after MPostcond from 58% +/- 8% (control) to 37% +/- 6% (morphine 3.0 microM, P < 0.01). This effect was abolished by coadministering naloxone (58% +/- 7%), nor-binaltorphimine (52% +/- 5%), but not naltrindole (34% +/- 5%). MPC and MPostcond had similar extent of protective effect on IS/AAR, and combining MPC with MPostcond did not afford further cardiprotection. 5-Hydroxydecanoate also abolished the cardioprotection of MPostcond. Unexpectedly, all three OR antagonists and 5-hydroxydecanoate themselves also afforded some extent of cardioprotection. MPost confers cardioprotection via activating kappa-OR but not delta-OR and opening mito-K atp channels. MPost and MPC have no additive protection. kappa-OR and mito-K atp channel may play a dual role in protecting ischemia-reperfusion injury.

Resistin, an adipocytokine, offers protection against acute myocardial infarction

Resistin, an adipocyte-derived hormone, is thought to represent a link between obesity and insulin-resistant diabetes. The potential role of resistin as a cardioprotective agent has not been explored. Our hypothesis is that resistin has a cardioprotective effect that is mediated by the resistin receptor-coupled activation of PI3K/Akt/PKC/K ATP dependent pathways. Our studies demonstrated that pretreatment of mouse hearts with 10 nM resistin for 5 min protected the heart against I/R injury in a mouse heart perfusion model. When mouse hearts were subjected to 60 min of LAD ligation followed by 4 h of reperfusion, resistin pretreatment (33 μg/kg) for 30 min or 24 h before ligation was able to significantly reduce the infarct size/risk area. The protective effect of resistin was abolished by wortmannin, as well as by an Akt inhibitor, triciribine. Resistin’s protective effect was absent in Akt kinase-deficient mutant mice. The protective effect was also blocked by chelerythrine, a PKC inhibitor, and εV1-2, a PKCε inhibitor. Finally, the protective effect was blocked by 5-hydroxydecanoate, which blocks the opening of mitoK ATP channels. Resistin-induced Akt phosphorylation in HL-1 cells was inhibited by wortmannin and triciribine. Resistin also induced PKCε phosphorylation, which was blocked by triciribine. These studies demonstrate that resistin’s cardioprotective effect is mediated by PI3K/Akt/PKC dependent pathways. In addition to cardiomyocytes, resistin also induced Akt phosphorylation in endothelial cells and smooth muscle cells, suggesting that resistin receptors are present in these cells. The effect of resistin on apoptosis was assessed in hearts subjected to 30 min of ischemia and 3 h of reperfusion. There were significantly fewer in situ oligo ligation-positive myocyte nuclei in mice treated with resistin. Our results show that resistin can dramatically reduce apoptosis and infarct size, thus protecting the heart against I/R injury.

Mitochondria are targets for geranylgeranylacetone-induced cardioprotection against ischemia-reperfusion in the rat heart

It has been shown that orally administered geranylgeranylacetone (GGA), an anti-ulcer drug, induces expression of heat shock protein 72 (HSP72) and provides protection against ischemia-reperfusion in rat hearts. The underlying protective mechanisms, however, remain unknown. Mitochondria have been shown to be a selective target for heat stress-induced cardioprotection. Therefore, we hypothesized that preservation of mitochondrial function, owing to an opening of a putative channel in the inner mitochondrial membrane, the mitochondrial ATP-sensitive potassium (mitoK(ATP)) channel, could be involved in GGA- or heat stress-induced cardioprotection against ischemia-reperfusion. Rats were treated with oral GGA or vehicle. Twenty-four hours later, each heart was isolated and perfused with a Langendorff apparatus. GGA-treated hearts showed better functional recovery, and less creatine kinase was released during a 30-min reperfusion period, after 20 min of no-flow ischemia. Concomitant perfusion with 5-hydroxydecanoate (5-HD, 100 microM) or glibenclamide (10 microM) abolished the GGA-induced cardioprotective effect. GGA also showed preserved mitochondrial respiratory function, isolated at the end of the reperfusion period, which was abolished with 5-HD treatment. GGA prevented destruction of the mitochondrial structure by ischemia-reperfusion, as shown by electron microscopy. In cultured cardiomyocytes, GGA induced HSP72 expression and resulted in less damage to cells, including less apoptosis in response to hypoxia-reoxygenation. Treatment with 5-HD abolished the GGA-induced cardioprotective effects but did not affect HSP72 expression. Our results indicate that preserved mitochondrial respiratory function, owing to GGA-induced HSP72 expression, may, at least in part, have a role in cardioprotection against ischemia-reperfusion. These processes may involve opening of the mitoK(ATP) channel.

ATP-sensitive Potassium Channel in Mitochondria of the Eukaryotic Microorganism Acanthamoeba castellanii

We describe the existence of a potassium ion transport mechanism in the mitochondrial inner membrane of a lower eukaryotic organism, Acanthamoeba castellanii. We found that substances known to modulate potassium channel activity influenced the bioenergetics of A. castellanii mitochondria. In isolated mitochondria, the rate of resting respiration is increased by about 10% in response to potassium channel openers, i.e. diazoxide and BMS-191095, during succinate-, malate-, or NADH-sustained respiration. This effect is strictly dependent on the presence of potassium ions in an incubation medium and is reversed by glibenclamide (a potassium channel blocker). Diazoxide and BMS-191095 also caused a slight but statistically significant depolarization of mitochondrial membrane potential (measured with a TPP(+)-specific electrode), regardless of the respiratory substrate used. The resulting steady state value of membrane potential was restored after treatment with glibenclamide or 1 mM ATP. Additionally, the electrophysiological properties of potassium channels present in the A. castellanii inner mitochondrial membrane are described in the reconstituted system, using black lipid membranes. Conductance from 90 +/- 7 to 166 +/- 10 picosiemens, inhibition by 1 mM ATP/Mg(2+) or glibenclamide, and activation by diazoxide were observed. These results suggest that an ATP-sensitive potassium channel similar to that of mammalian mitochondria is present in A. castellanii mitochondria.

Cardioprotective effects of bms-180448, a prototype mitokatp channel opener, and the role of salvage kinases, in the rat model of global ischemia and reperfusion heart injury

To investigate the involvement of reperfusion-induced salvage kinases (RISK) as possible signaling molecules for the cardioprotective effects of BMS-180448, a prototype mitochondrial ATP-sensitive K+ (mitoK(ATP)) channel opener, we measured its cardioprotective effects in a rat model of ischemia/reperfusion (I/R) heart injury, together with western blotting analysis of five different signaling proteins. In isolated rat hearts subjected to 30-min global ischemia followed by 30-min reperfusion, BMS-180448 (1, 3 and 10 microM) significantly increased reperfusion left ventricular developed pressure (LVDP) and 30-min reperfusion double product (heart rate x LVDP) in a concentration-dependent manner, while decreasing left ventricular end-diastolic pressure (LVEDP) throughout reperfusion period in a concentration-dependent manner. SDS-PAGE/western blotting analysis of left ventricle reperfused for 30 min revealed that BMS-180448 significantly decreased phospho-GSK3beta at high concentration, whereas it tended to increase slightly phospho-eNOS and phospho-p70S6K with concentration. However, BMS-180448 had no effect on phospho-Akt and phospho-Bad. These results suggest that the cardioprotective effects of BMS-180448 against I/R heart injury may result from direct activation of mitoK(ATP) channel in cardiomyocytes, with the minimal role of RISK pathway in the activation of this channel and the cardioprotective effects of BMS-180448.

Activation of α1B-adrenoceptors alleviates ischemia/reperfusion injury by limitation of mitochondrial Ca2+ overload in cardiomyocytes

Activation of alpha(1)-adrenergic receptors (alpha(1)-ARs) mimics ischemic preconditioning (IP). However, the subtypes of alpha(1)-ARs involved and the protective mechanisms are not entirely clear. Here we tested the hypothesis that preservation of mitochondrial integrity, in particular, Ca(2+) homeostasis via the epsilon isoform of protein kinase C (PKCepsilon) and mitoK(ATP) channels, may underlie the basis of alpha(1B)-AR-triggered cardioprotection. Indo-1 fluorescence in adult rat cardiomyocytes was used as an index of cytosolic ([Ca(2+)](c)) or mitochondrial free Ca(2+) concentration ([Ca(2+)](m)), and cell shortening was measured simultaneously. Cells were subjected to 20 min of simulated ischemia followed by 30 min of reperfusion (I/R). Activation of a(1)-ARs by phenylephrine significantly decreased I/R-induced [Ca(2+)](c) and [Ca(2+)](m) overload, mitochondrial cytochrome c release and ATP reduction, and improved Ca(2+) transients and cell shortening. These protective effects were markedly inhibited by blockade of alpha(1B)-AR (chloroethylclonidine) but not alpha(1A)-AR (5'-methylurapidil) or alpha(1D)-AR (BMY 7378). Moreover, phenylephrine-afforded protection on the [Ca(2+)](m), [Ca(2+)](c), and cell shortening was lost when mitoK(ATP) channels were inhibited with 5-hydroxydecanoate and PKCepsilon with PKCepsilon V(1-2). However, PKCepsilon V(1-2) did not affect the mitoK(ATP) channel opener diazoxide-induced protection on these parameters. These findings indicate that phenylephrine-induced protection on [Ca(2+)](m) homeostasis is mediated by selective activation of alpha(1B)-AR via mitoK(ATP) channel opening and PKCepsilon activation. Mitochondrial function appears to be a determinant of [Ca(2+)](c) and contractile function during I/R injury.

Hyperoxia confers cardioprotection in rats through involvement of ROS and mitok-ATP channels opening

Hyperoxia confers cardioprotection in rats through involvement of ROS and mitok-ATP channels opening

Iptakalim Alleviates Rotenone-Induced Degeneration of Dopaminergic Neurons through Inhibiting Microglia-Mediated Neuroinflammation

Inhibition of microglia-mediated neuroinflammation has been regarded as a prospective strategy for treating neurodegenerative disorders, such as Parkinson's disease (PD). In the present study, we demonstrated that systematic administration with iptakalim (IPT), an adenosine triphosphate (ATP)-sensitive potassium channel (K(ATP)) opener, could alleviate rotenone-induced degeneration of dopaminergic neurons in rat substantia nigra along with the downregulation of microglial activation and mRNA levels of tumor necrosis factor-alpha (TNF-alpha) and cyclooxygenase-2 (COX-2). In rat primary cultured microglia, pretreatment with IPT suppressed rotenone-induced microglial activation evidenced by inhibition of microglial amoeboid morphological alteration, declined expression of ED1 (a marker for activated microglia), and decreased production of TNF-alpha and prostaglandin E2 (PGE(2)). These inhibitory effects of IPT could be reversed by selective mitochondrial K(ATP) (mitoK(ATP)) channel blocker 5-hydroxydecanoate (5-HD). Furthermore, pretreatment with IPT prevented rotenone-induced mitochondrial membrane potential loss and p38/c-jun N-terminal kinase (JNK) mitogen-activated protein kinase (MAPK) activation in microglia, which might in turn regulate microglial activation and subsequent production of TNF-alpha and PGE(2). These data strongly suggest that the K(ATP) opener IPT may be a novel and promising neuroprotective drug via inhibiting microglia-mediated neuroinflammation.

Mito-KATP channels are not involved in the antiarrhythmic effect of peroxynitrite

We have shown previously that exogenously administered peroxynitrite (PN) markedly reduces ventricular arrhythmias resulting from ischaemia/reperfusion (I/R). The aim of the present study was to examine whether this protection is mediated through mitochondrial KATP channels (mitoKATP). Control dogs (C; n=10) were subjected to a 25 min occlusion of the left anterior descending coronary artery (LAD) followed by reperfusion. In 15 dogs PN (100 nM) was given in intracoronary (i.c.) infusion two times for 5 min, 5 min before the occlusion. In 7 dogs out of these 15 dogs 5-hydroxydecanoate (5-HD) was administered (150 μg kg−1 min−1, i.c.) 10 min prior to, and during the PN infusion. The severity of arrhythmias was determined during I/R, and nitrotyrosine (3-NT) formation was assessed following reperfusion by Western blot analysis. Compared to the controls PN significantly reduced the number of ventricular premature beats (VPBs, 289±84 vs. 84±34) and episodes of ventricular tachycardia (VT, 9.6±3.4 vs. 1.9±1.7) as well as the incidence of ventricular fibrillation (VF, 50% vs. 13%) during occlusion, and increased survival (0% vs. 50%) from the combined I/R insult. In these hearts 3-NT formation was also reduced. These protective effects against arrhythmias (VPBs; 43±26, VT; 0.4±0.5, VF; 0%; survival 60%) and 3-NT formation were not substantially modified by 5-HD. Our results suggest that the antiarrhythmic effect of exogenously administered PN is unlikely mediated through mitoKATP channels. This work was supported by OTKA (TO 37520).

α1B-Adrenoceptor activation improves postischemic contraction in cardiomyocytes: Cellular mechanisms

α1-Adrenoreceptor (α1-AR) activation protects the heart against ischemia/reperfusion (I/R)-injury, but the subtype(s) involved and the mechanisms remain unclarified. Intracellular Ca2+ ([Ca2+]i) overload is associated with I/R-injury. However, whether α1-AR activation limits I/R-induced [Ca2+]i overload and the signal transduction pathways are unknown. In the present study, we identified the α1-AR subtype that confers cardioprotection and tested the hypothesis that preservation of mitochondrial function, in particular, Ca2+ homeostasis, via activating PKCε and mitoKATP channels may underlie the basis of α1-AR-triggered cardioprotection.

NADPH oxidase- and mitochondrion-derived superoxide at rostral ventrolateral medulla in endotoxin-induced cardiovascular depression

We evaluated the contribution of superoxide anion (O2−) generated by NADPH oxidase or mitochondria in the rostral ventrolateral medulla (RVLM), where sympathetic premotor neurons for arterial pressure maintenance are located, on cardiovascular depression induced by inducible nitric oxide synthase-derived NO after Escherichia coli lipopolysaccharide (LPS) treatment. In Sprague–Dawley rats maintained under propofol anesthesia, microinjection of LPS bilaterally into the RVLM induced progressive hypotension, bradycardia, and reduction in sympathetic vasomotor outflow over our 240-min observation period. This was accompanied by an increase in O2− production (60–240 min) in the RVLM, alongside phosphorylation of p47phox or p67phox, upregulation of gp91phox or p47phox protein, and increase in Rac-1 or NADPH oxidase activity (60–120 min), and a depression of mitochondrial respiratory enzyme activity (120–240 min). Whereas inhibition of NADPH oxidase or knockdown of the gp91phox or p47phox gene blunted the early phase (60–150 min), coenzyme Q10 or mitochondrial KATP channel inhibitor antagonized the delayed phase (120–240 min) of LPS-induced increase in O2− production in RVLM and cardiovascular depression. We conclude that, whereas NADPH oxidase-derived O2− in RVLM participates predominantly in the early phase, O2− generated by depression in mitochondrial respiratory enzyme activity or opening of mitoKATP channels mediates the delayed phase of LPS-induced cardiovascular depression.

Neuronal preconditioning with the antianginal drug, bepridil

It has recently been shown that the antianginal drug bepridil (BEP) activates mitochondrial ATP-sensitive potassium (mitoK(ATP)) channels and thus confers cardioprotection. Our aim was to investigate whether BEP could induce preconditioning in cultured rat cortical neurons. Although BEP depolarized isolated and in situ mitochondria and increased reactive oxygen species generation, no acute protection was observed. However, a 3-day BEP-treatment elicited dose-dependent delayed neuroprotection against 180 min of oxygen-glucose deprivation (cell viability: untreated, 52.5 +/- 0.85%; BEP 1 micromol/L, 59.6 +/- 1.53%*; BEP 2.5 micromol/L, 71.9 +/- 1.23%*; BEP 5 micromol/L, 95.3 +/- 0.89%*; mean +/- SEM; *p < 0.05 vs. untreated) and 60 min of glutamate excitotoxicity (200 micromol/L; cell viability: untreated, 54.1 +/- 0.69%; BEP 1 micromol/L, 61.2 +/- 1.19%*; BEP 2.5 micromol/L, 78.1 +/- 1.67%*; BEP 5 micromol/L, 91.2 +/- 1.20%*; mean +/- SEM; *p < 0.05 vs. untreated), and inhibited the reactive oxygen species surge upon glutamate exposure. The protection was antagonized with co-application of the superoxide dismutase mimetic M40401, but not with reduced glutathione, catalase, or with the mitoK(ATP) blocker 5-hydroxydecanoate. Furthermore, BEP treatment resulted in increased levels of phosphorylated protein kinase C, manganese-dependent superoxide dismutase, glutathione peroxidase, and Bcl-2. Our results indicate that BEP induces delayed neuronal preconditioning which is dependent on superoxide generation but perhaps not on direct mitoK(ATP) activation.

Myocardial KATP channels are critical to Ca2+ homeostasis in the metabolically stressed heart in vivo

myocardial atp-sensitive potassium (KATP) channels appear to play central roles in the adaptation of the heart to both physiological and pathological metabolic stress ([1][1], [3][2], [5][3], [7][4], [8][5], [14][6], [16][7], [21][8]). In the heart, KATP channels are thought to act as metabolic

Mechanisms by which epoxyeicosatrienoic acids (EETs) elicit cardioprotection in rat hearts

Cytochrome P450 (CYP) epoxygenases and their arachidonic acid (AA) metabolites, the epoxyeicosatrienoic acids (EETs), have been shown to produce reductions in infarct size in canine myocardium following ischemia–reperfusion injury via opening of either the sarcolemmal K ATP (sarcK ATP) or mitochondrial K ATP (mitoK ATP) channel. In the present study, we subjected intact rat hearts to 30 min of left coronary artery occlusion and 2 h of reperfusion followed by tetrazolium staining to determine infarct size as a percent of the area at risk (IS/AAR, %). The results demonstrate that the two major regioisomers of the CYP epoxygenase pathway, 11,12-EET (2.5 mg/kg, iv) and 14,15-EET (2.5 mg/kg, iv) significantly reduced myocardial infarct size (IS/AAR, %) in rats as compared with control (41.9 ± 2.3%, 40.9 ± 1.2% versus 61.5 ± 1.6%, respectively), whereas, a third regioisomer, 8,9-EET (2.5 mg/kg, iv) had no effect (55.2 ± 1.4). The protective effect of pretreatment with 11,12- and 14,15-EETs was completely abolished (61.9 ± 0.7%, 58.6 ± 3.1%, HMR; 63.3 ± 1.2%, 63.2 ± 2.5%, 5-HD) in the presence of the selective sarcK ATP channel antagonist, HMR 1098 (6 mg/kg, iv) or the selective mitoK ATP channel antagonist, 5-HD (10 mg/kg, iv) given 10 min after 11,12- or 14,15-EET administration but 5 min prior to index ischemia. Furthermore, concomitant pretreatment with 11,12- or 14,15-EET in combination with the free radical scavenger, 2-mercaptopropionyl glycine (2-MPG), at a dose (20 mg/kg, iv) that had no effect on IS/AAR (57.7 ± 1.3%), completely abolished the cardioprotective effect of 11,12- and 14,15-EETs (58.2 ± 1.6%, 61.4 ± 1.0%), respectively. These data suggest that part of the cardioprotective effects of EETs in rat hearts against infarction is the result of an initial burst of reactive oxygen species (ROS) and subsequent activation of both the sarcK ATP and mitoK ATP channel.

Mitochondrial ATP-sensitive K + channels are redox-sensitive pathways that control reactive oxygen species production

Pharmacological mitochondrial ATP-sensitive K + channel (mitoK ATP) opening protects against ischemic damage and mimics ischemic preconditioning. However, physiological and pathological signaling events that open this channel are still not fully understood. We found that catalase, which removes H 2O 2, is capable of reversing the beneficial effects of ischemic preconditioning but not of mitoK ATP agonist diazoxide. On the other hand, 2-mercaptopropionylglycine prevented cardioprotection in both cases, suggesting that this compound may present effects other than scavenging of reactive oxygen species. Indeed, 2-mercaptopropionylglycine and a second thiol-reducing agent, dithiothreitol, impair diazoxide-mediated activation of mitoK ATP in isolated heart mitochondria. This demonstrates that mitoK ATP activity is regulated by thiol redox status. Furthermore, stimulating the generation of endogenous mitochondrial reactive oxygen species or treating samples with H 2O 2 strongly enhances mitoK ATP activity, in a manner probably dependent on redox sensors located in the channel's sulfonylurea receptor. We also demonstrate that mitoK ATP channel activity effectively prevents mitochondrial reactive oxygen release. Collectively, our results suggest that mitoK ATP acts as a reactive oxygen sensor that decreases mitochondrial free radical generation in response to enhanced local levels of oxidants. As a result, these channels regulate mitochondrial redox state under physiological conditions and prevent oxidative stress under pathological conditions such as ischemia/reperfusion.