ATP-sensitive Potassium Channel Opener Research Articles

Cytosolic calcium accumulation and delayed repolarization associated with ventricular arrhythmias in a guinea pig model of Andersen-Tawil syndrome

Andersen-Tawil syndrome (ATS1)-associated ventricular arrhythmias are initiated by frequent, hypokalemia-exacerbated, triggered activity. Previous ex vivo studies in drug-induced Andersen-Tawil syndrome (DI-ATS1) models have proposed that arrhythmia propensity in DI-ATS1 derives from cytosolic Ca(2+) ([Ca(2+)](i)) accumulation leading to increased triggered activity. The purpose of this study was to test the hypothesis that elevated [Ca(2+)](i) with concomitant APD prolongation, rather than APD dispersion, underlies arrhythmia propensity during DI-ATS1. DI-ATS1 was induced in isolated guinea pig ventricles by perfusion of 2 mM KCl Tyrode solution containing 10 μM BaCl(2). APD and [Ca(2+)](i) from the anterior epicardium were quantified by ratiometric optical voltage (di-4-ANEPPS) or Ca(2+) (Indo-1) mapping during right ventricular pacing with or without the ATP-sensitive potassium channel opener pinacidil (15 μM). APD gradients under all conditions were insufficient for arrhythmia induction by programmed stimulation. However, 38% of DI-ATS1 preparations experienced ventricular tachycardias (VTs), and all preparations experienced a high incidence of premature ventricular complexes (PVCs). Pinacidil decreased APD and APD dispersion and reduced VTs (to 6%), and PVC frequency (by 79.5%). However, PVC frequency remained significantly greater relative to control (0.5% ± 0.3% of DI-ATS1). Importantly, increased arrhythmia propensity during DI-ATS1 was associated with diastolic [Ca(2+)](i) accumulation and increased [Ca(2+)](i) transient amplitudes. Pinacidil partially attenuated the former but did not alter the latter. The study data suggest that arrhythmias during DI-ATS1 may be a result of triggered activity secondary to prolonged APD and altered [Ca(2+)](i) cycling and less likely dependent on large epicardial APD gradients forming the substrate for reentry. Therefore, therapies aimed at reducing [Ca(2+)](i) rather than APD gradients may prove effective in treatment of ATS1.

Akt involved in diazoxide preconditioning against rat hippocampal neuronal apoptosis induced by anoxia-reoxygenation injury

Investigate whether preconditioning with diazoxide (DZ), a mitochondrial ATP-sensitive potassium channel opener, could enhance Akt protein to up-regulate Bcl-2/Bax protein ratio against apoptosis. Cultured for 9 - 10 d in vitro, the hippocampal neurons of Sprague-Dawley rats were assigned to the following 5 groups randomly: Control (Group A), Anoxia (Group B), Anoxia + DZ100 micromol/L (Group C), Anoxia + DZ100 micromol/L + 5-hydroxydecanoate (Group D), Anoxia + DZ100 micromol/L + LY294002 50 micromol/L (Group E). Prior to oxygen deprivation, the hippocampal neurons were treated with DZ for 1 h per day and this treatment was persisted for 3 d. Each experiment was repeated for three times and each group contained 16 wells or 2 dishes of neurons for each time. Thereafter, neurons were derived of oxygen for 4 h. At 48 h of reoxygenation the neuronal optical density (A) were measured by MTT method, while the apoptotic rates were assayed by annexin V-FITC staining. The expressions of Akt, Bcl-2 and Bax proteins were detected and evaluated by Western blot. Compared with other pretreatment, DZ 100 micromol/L led to the elevation of A, whereas promoted the decrease of apoptotic rate (P < 0.05). Compared with those in other anoxic groups, the expressions of Akt protein and Bcl-2 protein in Group C were increased significantly (P < 0.05), whereas the Bax density were reduced significantly (P < 0.05). Preceding administration of 100 micromol/L DZ took protective effects on the neurons induced by anoxia-reoxygenation. DZ 100 micromol/L increased Akt protein to up-regulate Bcl-2/Bax protein ratio against apoptosis induced by anoxia-reoxygenation.

Mitochondria as a source of mechanical signals in cardiomyocytes

The myofibrillar and nuclear compartments in cardiomyocytes are known to be sensitive to extracellular mechanical stimuli. Recently, we have shown that alterations in the mitochondrial ionic balance in cells in situ are associated with considerably increased mitochondrial volume. Theoretically, this swelling of mitochondria could impose mechanical constraints on the myofibrils and nuclei in their vicinity. Thus, we studied whether modulation of mitochondrial volume in cardiomyocytes in situ has a mechanical effect on the myofibrillar and nuclear compartments. We used the measurement of passive force developed by saponin-permeabilized mouse ventricular fibres as a sensor for compression of the myofibrils. Osmotic compression induced by dextran caused an increase in passive force. Similarly, mitochondrial swelling induced by drugs that alter ionic homeostasis (alamethicin and propranolol) markedly augmented passive force (confirmed by confocal microscopy). Diazoxide, a mitochondrial ATP-sensitive potassium channel opener known to cause moderate mitochondrial swelling, also increased passive force (by 28 +/- 5% at 10% stretch, P < 0.01). This effect was completely blocked by 5-hydroxydecanoate (5-HD), a putative specific inhibitor of these channels. Mitochondrial swelling induced by alamethicin and propranolol led to significant nuclear deformation, which was visualized by confocal microscopy. Furthermore, diazoxide decreased nuclear volume, calculated using three-dimensional reconstructed images, in a 5-HD-dependent manner by 12 +/- 2% (P < 0.05). This corresponds to an increase in intracellular pressure of 2.1 +/- 0.3 kPa. This study is the first to demonstrate that mitochondria are able to generate internal pressure, which can mechanically affect the morphological and functional properties of intracellular organelles.

Transplantation of Mesenchymal Stem Cells Preconditioned with Diazoxide, a Mitochondrial ATP-Sensitive Potassium Channel Opener, Promotes Repair of Myocardial Infarction in Rats

Myocardial infarction (MI) causes myocardium injury and scar formation, and the transmural infarction is associated with ventricular hypofunction. Stem cell transplantation therapy has improved cardiac function in animal models of MI. However, the poor survival of the donor cells in the host myocardium hampers the therapeutic efficacy of stem cell transplantation. Diazoxide, a mitochondrial ATP-sensitive potassium channel opener, has been applied to suppress cell apoptosis and promote cell survival. We therefore assessed the effects of diazoxide on the selected mesenchymal stem cells (SMSCs). Pretreatment of SMSCs with diazoxide (200 micromol/L) for 30 min protected cells from oxidative stress injury by upregulating the expression of basic fibroblast growth factor and hepatocyte growth factor mRNAs and phospho-Akt and by preventing mitochondral cytochrome c translocation into the cytoplasm. Expression of mRNAs and proteins was detected by RT-PCR and western blot analyses. Thirty min after establishment of MI (the ligation of the left anterior descending of coronary artery) in female rats, the male rat SMSCs preconditioned with diazoxide were injected at four sites on the edge of the infarcted area. At 4 weeks after cell tranplantation, the donor cells in the recipient myocardium were tracked with Y chromosome. Preconditioning with diazoxide improved the survival rate of the transplanted SMSCs, compared to the untreated SMSCs. Moreover, transplantation of the diazoxide-pretreated SMSCs reduced the infarct size and increased left ventricular function, as judged by transthoracic echocardiography. In conclusion, diazoxide preconditioning is effective to promote SMSCs survival under oxidative stress and attenuates cardiac injury in MI.

Noradrenaline Protects In Vivo Rat Heart Against Infarction and Ventricular Arrhythmias Via Nitric Oxide and Reactive Oxygen Species

Our previous study showed that pretreatment with noradrenaline via opening of the mitochondrial ATP-sensitive potassium channel protects myocardium against ischemia/reperfusion injuries. We have hypothesized that production of nitric oxide (NO) and generation of reactive oxygen species (ROS) are involved in noradrenaline-induced cardioprotection in rat heart. All anesthetized rats underwent 25 min of regional ischemia followed by 120 min of reperfusion. Animals were randomized to receive one of the following treatment: saline, noradrenaline (2 μg/kg, i.v.), noradrenaline plus prazosin (an α(1)-adrenoceptor blocker, 0.5mg/kg, i.v.), noradrenaline plus L-NAME (a nonspecific NOS inhibitor, 10mg/kg, i.v.), noradrenaline plus tempol (a membrane-permeable radical scavenger, 30 mg/kg, i.v.), Prazosin alone, only L-NAME and tempol alone. Infarct size (% of risk area) was reduced from 49.6 ± 2.4 in saline-control group to 18.2 ± 1.5 in noradrenaline preconditioned group. Administration of prazosin, L-NAME, or tempol prior to noradrenaline injection abolished the observed cardioprotection of noradrenaline (45.5 ± 3, 41.7 ± 4.5 and 38.7 ± 5.4, respectively) and restored infarct size to saline-control rats' level. Incidences and severity of ventricular arrhythmia during ischemia and early reperfusion significantly decreased in noradrenaline preconditioned group compared with saline-control group. This cardioprotective effect of noradrenaline against ventricular arrhythmia was abrogated by administration of prazosin, L-NAME, or tempol. Cardioprotection effect of the α(1)-adrenoceptor stimulation by noradrenaline was inhibited by L-NAME or tempol in anesthetized rat heart.

Opening of ATP-sensitive Potassium Channel by Insulin in the Brain-induced Insulin Secretion in Wistar Rats

Cerebral insulin can regulate glucose homeostasis via activation of the parasympathetic nervous system, which results in the reduction of hepatic glucose output. However, the precise mechanism(s) through which cerebral insulin directly exerts an effect on insulin secretion remains unclear. In the present study, we found that cerebral administration of insulin caused an increase of plasma insulin concentration and a concomitant decrease in plasma glucose levels within one hour. These effects were blocked by vagotomy or intraperitoneal injection of 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide, a specific M (3) antagonist. The mediating influence of parasympathetic activation can thus be considered. The adenosine triphosphate-sensitive potassium (K-ATP) channel is a key mediator of the cerebral action of insulin. The plasma glucose-lowering action of insulin was abolished by cerebral administration of glibenclamide or repaglinide at concentrations sufficient to block K-ATP channels. In conclusion, our findings suggest that cerebral insulin may induce insulin release by stimulating the opening of K-ATP channels, which in turn activate parasympathetic tone in pancreatic tissue.

Coupling of Liquid Chromatography/Tandem Mass Spectrometry and Liquid Chromatography/Solid-Phase Extraction/NMR Techniques for the Structural Identification of Metabolites following In Vitro Biotransformation of SUR1-Selective ATP-Sensitive Potassium Channel Openers

SUR1-selective ATP-sensitive potassium channel openers (PCOs) have been shown to be of clinical value for the treatment of several metabolic disorders, including type I and type II diabetes, obesity, and hyperinsulinemia. Taking into account these promising therapeutic benefits, different series of 3-alkylamino-4H-1,2,4-benzothiadiazine 1,1-dioxides structurally related to diazoxide were developed. In view of the lead optimization process of the series, knowledge of absorption, distribution, metabolism, excretion, and toxicity parameters, and more particularly the metabolic fate of these compounds, is a fundamental requirement. For such a purpose, two selected promising compounds [7-chloro-3-isopropylamino-4H-1,2,4-benzothiadiazine 1,1-dioxide (BPDZ 73) and 7-chloro-3-(3-pentylamino)-4H-1,2,4-benzothiadiazine 1,1-dioxide (BPDZ 157)] were incubated in the presence of phenobarbital-induced rat liver microsomes to produce expected mammal in vivo phase I metabolites. The resulting major metabolites were then analyzed by both mass spectrometry (MS) and NMR to completely elucidate their chemical structures. The two compounds were also further incubated in the presence of nontreated rats and human microsomes to compare the metabolic profiles. In the present study, the combined use of an exact mass liquid chromatography (LC)/tandem MS platform and an LC/solid-phase extraction/NMR system allowed the clarification of some unresolved structural assessments in the accurate chemical structure elucidation process of the selected PCO drugs. These results greatly help the optimization of the lead compounds.

Modeling Cardiac Action Potential Shortening Driven by Oxidative Stress-Induced Mitochondrial Oscillations in Guinea Pig Cardiomyocytes

Ischemia-induced shortening of the cardiac action potential and its heterogeneous recovery upon reperfusion are thought to set the stage for reentrant arrhythmias and sudden cardiac death. We have recently reported that the collapse of mitochondrial membrane potential (ΔΨ m) through a mechanism triggered by reactive oxygen species (ROS), coupled to the opening of sarcolemmal ATP-sensitive potassium (K ATP) channels, contributes to electrical dysfunction during ischemia-reperfusion. Here we present a computational model of excitation-contraction coupling linked to mitochondrial bioenergetics that incorporates mitochondrial ROS-induced ROS release with coupling between the mitochondrial energy state and electrical excitability mediated by the sarcolemmal K ATP current ( I K,ATP). Whole-cell model simulations demonstrate that increasing the fraction of oxygen diverted from the respiratory chain to ROS production triggers limit-cycle oscillations of ΔΨ m, redox potential, and mitochondrial respiration through the activation of a ROS-sensitive inner membrane anion channel. The periods of transient mitochondrial uncoupling decrease the cytosolic ATP/ADP ratio and activate I K,ATP, consequently shortening the cellular action potential duration and ultimately suppressing electrical excitability. The model simulates emergent behavior observed in cardiomyocytes subjected to metabolic stress and provides a new tool for examining how alterations in mitochondrial oxidative phosphorylation will impact the electrophysiological, contractile, and Ca 2+ handling properties of the cardiac cell. Moreover, the model is an important step toward building multiscale models that will permit investigation of the role of spatiotemporal heterogeneity of mitochondrial metabolism in the mechanisms of arrhythmogenesis and contractile dysfunction in cardiac muscle.

The role of cGMP and PKG in cardioprotection

Interrupting blood flow to a heart (ischemia) for several minutes followed by several minutes of restored flow renders it very resistant to cell death from a subsequent much more prolonged ischemic episode. This phenomena, termed ischemic preconditioning, is the result of a complex signal transduction pathway which ultimately prevents formation of lethal mitochondrial permeability transition pores when blood flow is restored. In mapping this pathway we found that NOS was in the pathway [1] and proposed that it was involved in opening ATP-sensitive potassium channels (KATP) in the mitochondrial inner membrane. Those channels play a key role in preconditioning's mechanism. Incubating isolated heart mitochondria in cGMP + PKG + ATP opened the channel as indicated by mitochondrial swelling [2]. To test this hypothesis further the cGMP analog 8-CPT-cGMP (10 μM) was infused for 30 min starting 5 min prior to the end of 30 min of regional ischemia in an isolated rabbit heart [3]. It greatly attenuated the resulting infarction by an amount similar to that seen with preconditioning (infarct size approximately 50% of that in untreated hearts). 8-CPT's protection could be blocked by the KATP blocker 5 hydroxydecanoate confirming its position in the pathway. Because there is a medical need for an intervention that prevents myocardial infarction when given at reperfusion we looked for a method of raising cGMP that would be clinically feasible. We gave the direct guanylyl cyclase activator BAY 58-2667 (cinaciguat) to open-chest rabbits at 53 μg/kg bolus just prior to reperfusion followed by a 60 min 1.25 μg/min infusion. It reduced infarct size from 40.6% of the ischemic zone to only 16.0%. In another experiment we tested Vardenafil infusion of 140 μg/kg/h starting 5 min before coronary occlusion in the open-chest rabbit. It also greatly protected the hearts against infarction. Because it is controversial as to whether heart muscle expresses PDE 5, we transfected isolated rabbit ventricular myocytes with CNG channels selective for cGMP. In the resting cells cGMP as monitored with patch clamping was near zero and did not increase when they were exposed to 1 μM vardenafil. However, when cGMP was first increased with 1 μM BAY 58-2667 addition of vardenafil greatly increased the cGMP-dependent membrane current indicating that a vardenafil-sensitive PDE was present which is presumed to be PDE5. We conclude that raising cGMP in the reperfused heart is an effective anti-infarct strategy.

Predictive models, based on classification algorithms, for compounds potentially active as mitochondrial ATP-sensitive potassium channel openers

Heart mitochondrial ATP-sensitive potassium channel (mito-K ATP channels) are deeply implicated in the self-defense mechanism of ischemic preconditioning. Therefore, exogenous molecules activating these channels are considered as a promising pharmacological tool to reduce the myocardial injury deriving from ischemia/reperfusion events. In our laboratory, a series of 4-spiro-substituted benzopyran derivatives were earlier synthesized and some of them exhibited anti-ischemic properties. In this study, the above compounds are exploited in order to develop QSAR models, based on classification approaches, capable of discriminating between the ones acting as cardioprotective agents and those that are unable to elicit such a property. Molecules belonging to the whole dataset were subjected to CODESSA and E-Dragon calculations in order to compute a large number of molecular descriptors enabling the construction of classification models. Based on the two program packages used, two different experiments were carried out, with the aim of identify batteries of models to be exploited for designing new cardioprotective agents from libraries of new chemical entities. Both model batteries satisfy the rigorous criteria adopted for the validation, either when tested on the training and test set, according to the most straightforward protocol, and when tested on an additional prediction set. They were proven to ensure successful applications in the field of cardioprotective agent design.

To prevent, protect and save the ischemic heart: antioxidants revisited

Reactive oxygen species (ROS) and reactive nitrogen species (RNS) formation increases in the post-ischemic myocardium and represents a mechanism of post ischemic injury. ROS, which are formed during reperfusion, trigger lipid peroxidation, oxidize proteins and cause DNA strand breaks, all interfering with and potentially damaging normal cellular function. Oxidative stress is associated with poor recovery of left ventricular function after a sustained period of ischemia and, according to several studies, it contributes significantly to the acceleration of necrosis and thus extension of infarction, apoptosis, arrhythmiogenesis and endothelial dysfunction. Accordingly, targeting the generation of ROS with various antioxidants has been shown to improve left ventricular function after the restoration of flow. Apart from mechanical or pharmacological interventions that open the occluded artery, the heart has endogenous mechanisms of protection, called ischemic pre-conditioning and ischemic post-conditioning. Opening of the mitochondrial ATP-sensitive potassium channels and subsequent generation of ROS is considered to be a pivotal step in the mechanisms of pre- and post-conditioning. Notably, ROS play an ambiguous role in the protection of myocardium and can be both effective and harmful. Thus, the role of antioxidants in the attenuation of the effects of pre-and post-conditioning in vivo remains controversial.

Inhibition of hypothermic preservation‐induced Smac/DIABLO protein expression by diazoxide in donor rat myocardium

AIMTo investigate the effect of a mitochondrial ATP‐sensitive potassium channel (mitoKATP) opener diazoxide (DE) on Smac/DIABLO protein expressions in rat heart suffered from different duration of hypothermic preservation.METHODSThe Langendorff model of isolated rat heart was used. After stored in 4ºC Celsior solution with or without DE (30 μmol/L) for different time (0, 3, 6, 9 or 12 h). Cell apoptosis was detected by TUNEL technique. The expression of Smac/ DIABLO protein was also analyzed by Western blotting.RESULTS(1) Compared with the hypothermic preservation groups, DE reduced the percentage of apoptotic cells. (2) The peak level of Smac/DIABLO protein expression appeared at 6 h after hypothermic preservation, which was postponed to 9 h by DE. (3) The above effects of DE were attenuated by a mitoKATP channel inhibitor 5‐hydroxydecanoate (5‐HD).CONCLUSIONSThe findings indicate that in the isolated rat heart DE protects myocardium against different duration of hypothermic preservation injury via opening of mitoKATP channel and inhibition of Smac/DIABLO protein expression.

Neuroprotective effect of adenoviral catalase gene transfer in cortical neuronal cultures

Reduced availability of reactive oxygen species is a key component of neuroprotection against toxic stimuli. Recently, we have shown that the hydrogen peroxide scavenger catalase plays a central role in delayed preconditioning induced by the mitochondrial ATP‐sensitive potassium channel opener BMS‐191095 (Gáspár et al. JCBFM 28 (6): 1090‐1103, 2008). The aim of our present experiments was to investigate the neuroprotective effect of catalase in vitro using an adenoviral catalase gene transfer protocol. To induce catalase overexpression, cultured rat cortical neurons were incubated with the adenoviral vector Ad5CMVcatalase and control cells with Ad5CMVntLacZ for 24 h. Gene transfer increased catalase protein levels and activity, but did not influence other antioxidants. Catalase transfection did not protect against glutamate excitotoxicity and oxygen‐glucose deprivation, but conferred a dose‐dependent neuroprotection against 6 h exposure to 80 μM hydrogen peroxide (viability: control, 20.70±1.96%; LacZ 10 pfu/cell, 15.00± 1.66%; catalase 1 pfu/cell, 57.60±5.46%*; catalase 2 pfu/cell, 104.90±1.74%*; catalase 10 pfu/cell, 111.20±1.92%*; *p&lt;0.05 vs. control; mean±SEM). Finally, the protection could be antagonized using the catalase inhibitor 3‐aminotriazole. Our results support the view that enhancing cellular antioxidant capacity may play a crucial role in neuroprotective strategies.

KATP activation prevents progression of cardiac hypertrophy to failure induced by pressure overload via protecting endothelial function

We investigated the effects of iptakalim, a new ATP-sensitive potassium channel (K(ATP)) opener providing endothelial protection, on the progression of cardiac hypertrophy to failure in a rat model of pressure overloading caused by abdominal aortic banding (AAB). Endothelial dysfunction is central to cardiac hypertrophy and failure induced by pressure overload. It would be useful to clarify whether iptakalim could prevent this. The effects of pressure overload were assessed in male Sprague-Dawley rats 6 weeks after AAB using progression of cardiac hypertrophy to heart failure as the endpoint. The AAB-treated rats had significantly elevated blood pressure, systolic and diastolic cardiac dysfunction, evidence of left ventricular hypertrophy (LVH), and transition to heart failure. LVH was characterized by increases in the ratios of heart and left ventricular weights to body weight, increased myocyte cross-sectional areas, myocardial and perivascular fibrosis, and elevated cardiac hydroxyproline. These could be prevented by treatment with iptakalim at daily oral doses of 1, 3, and 9 mg/kg for 6 weeks. Progression to cardiac failure, demonstrated by increases in relative lung and right ventricular weights, cardiac function disorders and overexpression of atrial and B-type natriuretic peptide mRNA, could also be prevented. The downregulated nitric oxide signalling system was enhanced, whereas the upregulated endothelin signalling system was inhibited, resulting in normalization of the balance between these two systems. Iptakalim protected the endothelium and prevented progression of cardiac hypertrophy to failure induced by a pressure overload.

Neuroprotective effect of adenoviral catalase gene transfer in cortical neuronal cultures

Reduced availability of reactive oxygen species is a key component of neuroprotection against various toxic stimuli. Recently we showed that the hydrogen peroxide scavenger catalase plays a central role in delayed preconditioning induced by the mitochondrial ATP-sensitive potassium channel opener BMS-191095. The purpose of the experiments discussed here was to investigate the neuroprotective effect of catalase in vitro using a recombinant adenoviral catalase gene transfer protocol. To induce catalase overexpression, cultured rat cortical neurons were infected with the adenoviral vector Ad5CMVcatalase and control cells were incubated with Ad5CMVntLacZ for 24 h. Gene transfer effectively increased catalase protein levels and activity, but did not influence other antioxidants tested. Ad5CMVcatalase, with up to 10 plaque forming units (pfu) per neuron, did not affect cell viability under control conditions and did not protect against glutamate excitotoxicity or oxygen–glucose deprivation. In contrast, catalase overexpression conferred a dose-dependent protection against exposure to hydrogen peroxide (viability: control, 33.02 ± 1.09%; LacZ 10 pfu/cell, 32.85 ± 1.51%; catalase 1 pfu/cell, 62.09 ± 4.17%⁎; catalase 2 pfu/cell, 98.71 ± 3.35%⁎; catalase 10 pfu/cell, 99.68 ± 1.99%⁎; ⁎ p < 0.05 vs. control; mean ± SEM). Finally, the protection could be antagonized using the catalase inhibitor 3-aminotriazole. Our results support the view that enhancing cellular antioxidant capacity may play a crucial role in neuroprotective strategies.

Effects of ATP-sensitive potassium channels on the expression of P21, P27 and leptin

This study investigated the effects of ATP-sensitive potassium channels on the expression of P21, P27 and leptin. The expression of receptor of ATP-sensitive potassium channels (sulphonylurea receptor, SUR) mRNA in the preadipocytes and leptin mRNA was detected by PCR after rat preadipocytes were treated with the opener (diazoxide) or inhibitor (glibenclamide) of ATP-sensitive potassium channels during the process of inducing differentiation. The expression of P21 and P27 in preadipocytes treated with diazoxide or glibenclamide was assayed by Western blot. The results showed that the expression of SUR2, not SUR1 was detected in adipose tissue, preadipocytes and adipocytes. After treatment of preadipocytes with diazoxide, the expression levels of P21 and P27 were obviously higher than those in control group, but the expression levels of P21 and P27 in glibenclamide-treated group were lower than those in control group. During the process of inducing differentiation, the expression of leptin mRNA in preadipocytes treated with diazoxide was increased greatly, but the expression of leptin mRNA in glibenclamide-treated group decreased obviously. It was concluded that ATP-sensitive potassium channels might be involved in the proliferation and differentiation of rat preadipocytes by changing the expression of P21, P27 and leptin.

Levosimendan pre-treatment improves outcomes in patients undergoing coronary artery bypass graft surgery

The calcium sensitizer levosimendan has anti-ischaemic effects mediated via the opening of sarcolemmal and mitochondrial ATP-sensitive potassium channels. These properties suggest potential application in clinical situations where cardioprotection would be beneficial, such as cardiac surgery. We thus decided to investigate whether pharmacological pre-treatment with levosimendan reduces intensive care unit (ICU) length of stay in patients undergoing elective myocardial revascularization under cardiopulmonary bypass. One hundred and six patients undergoing elective coronary artery bypass grafting were randomly assigned in a double-blind manner to receive levosimendan or placebo. Levosimendan (24 microg kg(-1)) or placebo was administered as a slow i.v. bolus over a 10 min period before the initiation of bypass. Tracheal intubation time and the length of ICU stay were significantly reduced in the levosimendan group (P<0.01). The number of patients needing inotropic support for >12 h was significantly higher in the control group (18.0% vs 3.8%; P=0.021). Compared with control patients, levosimendan-treated patients had lower postoperative troponin I concentrations (P<0.0001) and a higher cardiac power index (P<0.0001). Pre-treatment with levosimendan in patients undergoing surgical myocardial revascularization resulted in less myocardial injury, a reduction in tracheal intubation time, less requirement for inotropic support, and a shorter length of ICU stay.

Neuroprotective Effects of Diazoxide and Its Antagonism by Glibenclamide in Pyramidal Neurons of Rat Hippocampus Subjected to Ischemia-Reperfusion-Induced Injury

Mitochondrial ATP-sensitive potassium channel opener, diazoxide, is shown to have protective effect on the heart and brain following ischemia-reperfusion-induced injury (IR/II). However, the detailed effect of diazoxide and its antagonist on neuronal death, mitochondrial changes, and apoptosis in cerebral IR/II has not fully studied. IR/II was induced in rats by the 4-vessel occlusion model. Neuronal cell death and mitochondrial changes in CA1–CA4 pyramidal cells of the hippocampus were studied by light and electron microscopy, respectively. Apoptosis was assessed by measuring the amount of protein expressed by Bax and Bcl-2 genes. In light microscopy studies, the number of total and normal cells were increased only following 18 mg/kg of diazoxide. Lower doses (2 and 6 mg/kg) failed to change the cell numbers. All three doses of glibenclamide (1, 5, and 25 mg/kg) decreased the number of total and normal cell populations. In electron microscopy studies, different doses of diazoxide and glibenclamide prevented and aggravated the IR-induced morphological changes, respectively. Western blot analysis showed that diazoxide and glibenclamide inhibited and enhanced Bax protein expression respectively. Regarding Bcl-2 expression, only diazoxide showed a significant enhancement of gene expression. In conclusion, the results show that diazoxide can exhibit neuroprotective effects against IR/II in hippocampal regions, possibly through the opening of mitochondrial ATP-sensitive K+ channels.

Levosimendan: from basic science to clinical practice

Levosimendan is a new cardiac enhancer that exerts positive inotropic effects on the failing heart mediated by calcium sensitization of contractile proteins as well as peripheral vasodilatory effects mediated by opening of ATP-sensitive potassium channels in vascular smooth-muscle cells. Levosimendan is the most well-studied calcium sensitizer in the real clinical practice, producing greater hemodynamic and symptomatic improvement in patients with acute heart failure syndromes (AHFS) than those with traditional inotropes. Immunomodulatory and anti-apoptotic properties of levosimendan may be an additional biologic mechanism that prevents further cytotoxic and hemodynamic consequences of abnormal immune and neurohormonal responses in AHFS. Recent mortality trials showed that levosimendan does not improve short- and long-term prognosis in AHFS in comparison to dobutamine or placebo. However, in patients with a previous history of CHF and on beta-blocker on admission, levosimendan seems to have a beneficial effect on short-term mortality. According to the recent guidelines of the European Society of Cardiology, levosimendan is indicated in patients with symptomatic low cardiac output HF secondary to cardiac systolic dysfunction without severe hypotension (Class IIa, Level of Evidence B).

Activation of ATP-sensitive potassium channels protects vascular endothelial cells from hypertension and renal injury induced by hyperuricemia

It has been demonstrated that hyperuricemia induces reno-cardiovascular damage resulting in hypertension and renal injury because of vascular endothelial dysfunction. The pathogenesis of hyperuricemia, endothelial dysfunction, hypertension, and renal injury is progressive, and develops into a vicious cycle. It is reasonable to suggest that an antihypertensive drug with endothelial protection may block this vicious cycle. Iptakalim, a novel antihypertensive drug undergoing phase-three clinical trials, is a new ATP-sensitive potassium channel opener and can ameliorate endothelial dysfunction. We hypothesized that iptakalim could prevent hypertension and retard the pathogenesis of endothelial dysfunction and renal injury in hyperuricemic rats. In rats with hyperuricemia induced by 2% oxonic acid and 0.1 mmol/l uric acid, iptakalim prevented increases in systolic blood pressure, reduced the impairment of endothelial vasodilator function, and attenuated renal dysfunction and pathological changes in glomerular and renal interstitial tissue at 0.5, 1.5, and 4.5 mg/kg orally daily for 4 weeks. Serum levels of nitric oxide and prostacyclin, and gene expression of endothelial nitric oxide synthase in the aortic and intrarenal tissue, were increased, whereas the serum levels of endothelin-1 and gene expression of endothelin-1 in aortic and intrarenal tissue were decreased. However, serum levels of angiotensin II and renin remained unchanged in the hyperuricemic rats treated with iptakalim. In cultured rat aortic endothelial cells, amelioration of endothelial dysfunction by iptakalim was suggested by inhibition of the overexpression of intercellular adhesive molecule-1, vascular cell adhesive molecule-1, and monocyte chemoattractant protein-1 mRNA induced by uric acid, and reversal of the inhibitory effects of uric acid on nitric oxide release in a concentration-dependent manner, which could be abolished by pretreatment with glibenclamide, an ATP-sensitive potassium channel blocker. Iptakalim ameliorated hyperuricemia in this rat model by decreasing renal damage through its antihypertensive and endothelial protective properties, and it had no direct effects on anabolism, catabolism and excretion of uric acid. These findings suggest that the activation of ATP-sensitive potassium channels by iptakalim can protect endothelial function against hypertension and renal injury induced by hyperuricemia. Iptakalim is suitable for use in hypertensive individuals with hyperuricemia.