ATP-dependent Potassium Research Articles

Détecteurs de glucose et régulation de la prise alimentaire

Based on the glucostactic and lipostastic theories, it has been proposed that two dependent phenomena could be involved in the control of food intake. At short term, the regulation of food intake is part of the of the glucoregulatory mechanism, i.e. to the mechanism of use and replenishment of the glucose stores. This mechanism which is mainly involved in meal initiation is modulated by a second mechanism linked to the use and storage of lipids i.e. fat mass. Some molecules have been proposed to play the role of signal informing some brain structures of the metabolic status of the individual, among them leptin, insulin and glucose seems to play a major role. New insights into the cellular and molecular mechanism that support the fact that the electrical activity of some neurones depends on the glucose and/or insulin concentration, have been gained recently. Making the analogy with the beta cell of the islet of Langerhans, it has been proposed that proteins such as glucose transporter 2, glucokinase and the ATP-dependent potassium channel, could assure this sensing. Alteration in this sensing system could be at the origin of some alterations in feeding behavior.

Expression and function of ATP-dependent potassium channels in late post-infarction remodeling

Myocardial remodeling late after infarction is associated with increased incidence of fatal arrhythmias. Heterogeneous prolongation of the action potential in the surviving myocardium is one of the predominant causes. Sarcolemmal ATP-dependent potassium ( K ATP) channels are important metabolic sensors regulating electrical activity of cardiomyocytes and are capable of considerably shortening the action potential. We determined whether ATP-dependent potassium channels generate or, on the contrary prevent the heterogeneity in action potential prolongation. Cardiomyocytes were obtained from the infarct border zone, the septum and the right ventricle of rat hearts 20 weeks after coronary occlusion when rats developed signs of heart failure. Expression of the conductance subunit Kir6.1, but not Kir6.2, and of all SUR regulatory subunits was increased up to 3-fold in cardiomyocytes from the infarct border zone. Concomitantly, there was a prominent response of the K ATP current to diazoxide that was not detectable in control cardiomyocytes. The action potential was prolonged in cardiomyocytes from the infarct border zone (74 ms) relative to sham (41 ms). However, activation of the K ATP channels by diazoxide reduced action potential duration to 42 ms. In myocytes of the septum and right ventricle, expression of channel subunits, duration of action potential, and sensitivity to diazoxide were only slightly increased relative to shams. In conclusion, the myocardium remodeled after infarction displays alterations of K ATP expression and function with spatial heterogeneity matching that of the action potential prolongation. Drugs selectively activating diazoxide-sensitive sarcolemmal K ATP channels should be considered in the prevention of arrhythmias in post-infarction heart failure.

Metabolic and ionic coupling factors in amino acid-stimulated insulin release in pancreatic β-HC9 cells

Fuel stimulation of insulin secretion from pancreatic beta-cells is thought to be mediated by metabolic coupling factors that are generated by energized mitochondria, including protons, adenine nucleotides, and perhaps certain amino acids (AA), as for instance aspartate, glutamate, or glutamine (Q). The goal of the present study was to evaluate the role of such factors when insulin release (IR) is stimulated by glucose or AA, alone or combined, using (31)P, (23)Na and (1)H NMR technology, respirometry, and biochemical analysis to study the metabolic events that occur in continuously superfused mouse beta-HC9 cells contained in agarose beads and enhanced by the phosphodiesterase inhibitor IBMX. Exposing beta-HC9 cells to high glucose or 3.5 mM of a physiological mixture of 18 AA (AAM) plus 2 mM glutamine caused a marked stimulation of insulin secretion associated with increased oxygen consumption, cAMP release, and phosphorylation potential as evidenced by higher phosphocreatine and lower P(i) peak areas of (31)P NMR spectra. Diazoxide blocked stimulation of IR completely, suggesting involvement of ATP-dependent potassium (K(ATP)) channels in this process. However, levels of MgATP and MgADP concentrations, which regulate channel activity, changed only slowly and little, whereas the rate of insulin release increased fast and very markedly. The involvement of other candidate coupling factors was therefore considered. High glucose or AAM + Q increased pH(i). The availability of temporal pH profiles allowed the precise computation of the phosphate potential (ATP/P(i) x ADP) in fuel-stimulated IR. Intracellular Na+ levels were greatly elevated by AAM + Q. However, glutamine alone or together with 2-amino-2-norbornanecarboxylic acid (which activates glutamate dehydrogenase) decreased beta-cell Na levels. Stimulation of beta-cells by glucose in the presence of AAM + Q (0.5 mM) was associated with rising cellular concentrations of glutamate and glutamine and strikingly lower aspartate levels. Methionine sulfoximine, an inhibitor of glutamine synthetase, blocked the glucose enhancement of AMM + Q-induced IR and associated changes in glutamine and aspartate but did not prevent the accumulation of glutamate. The results of this study demonstrate again that an increased phosphate potential and a functional K(ATP) channel are essential for metabolic coupling during fuel-stimulated insulin release but illustrate that determining the identity and relative importance of all participating coupling factors and second messengers remains a challenge largely unmet.

Decreased expression of aortic KIR6.1 and SUR2B in hypertension does not correlate with changes in the functional role of KATP channels

ATP-dependent potassium (KATP) channels are the target of multiple vasoactive factors and drugs and their functional role in hypertension is controversial. We aimed to analyze the possible changes of KATP channels expression and function during hypertension. We used endothelium-denuded aorta segments from Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) to analyze the 1) expression of KATP subunits Kir6.1, Kir6.2 and SUR2B, 2) the KATP currents recorded using the patch-clamp technique and 3) the vasodilator response to the KATP channel opener pinacidil. While Kir6.2 was not detected in aorta from either strain, Kir6.1 and SUR2B were expressed in the medial layer of the aorta from WKY and SHR, and their expression decreased in hypertension. However, pinacidil increased K+ currents and induced a vasodilator response which was not modified in hypertension. These effects were also similarly inhibited by the KATP channel blocker glybenclamide. Our data demonstrate direct evidence of decreased aortic Kir6.1/SUR2B subunit expression in hypertension, but preserved functional responses to a KATP channel opener. Supported by AGL2004-06685/ALI and SAF2005-03770

Glibenclamide Action on Myocardial Function and Arrhythmia Incidence in the Healthy and Diabetic Heart

Myocardial sarcolemmal ATP-dependent potassium (KATP) channels, which are normally closed by high ATP concentration, open during ischemia when ATP generation decreases favoring K(+) efflux. This reduces action potential duration (APD) decreasing the time of Ca(2+) influx and Ca(2+) overload. This behavior suggested that they might be involved in the protection against stunning and arrhythmias and in the mechanism of ischemic preconditioning. Sulfonylureas, used as hypoglycemic agents for the treatment of type 2 diabetes also block myocardial KATP channels prolonging APD during ischemia, which by allowing Ca(2+) entry for a longer period of time, is potentially harmful to the heart. Controversial findings have been reported regarding the protective effect of sulfonylureas. Due to their importance in the clinical setting, their action on the heart of large conscious animal models is relevant. The effect of glibenclamide, a representative sulfonylurea, has been studied in a conscious sheep model submitted to regional 12 min ischemia. Glibenclamide (0.4 mg/kg) completely blocked KATP channels, as assessed by monophasic APD, producing a deleterious effect on reperfusion-induced arrhythmias and myocardial recovery from stunning in normal animals. This adverse effect was more noticeable in alloxan-induced diabetic sheep, where a lower dose (0.1 mg/kg) inhibited KATP channel opening worsening mechanical recovery and arrhythmia incidence. However, glibenclamide did not abolish ischemic late preconditioning against stunning and arrhythmias in normal animals. Because diabetic sheep do not develop this cardioprotective phenomenon, probably due to KATP channel dysfunction, it was not possible to assess glibenclamide effect on preconditioning in this pathological condition. In conclusion, in large conscious animals, glibenclamide interferes with the beneficial action of KATP channel opening during acute ischemia-reperfusion events both in normal and diabetic animals. Therefore, despite some studies claiming no added cardiovascular risk due to glibenclamide treatment, this pharmacological agent should be further investigated to ensure its safe administration in patients with concurrent heart disease.

Preconditioning protects endothelium by preventing ET-1-induced activation of NADPH oxidase and xanthine oxidase in post-ischemic heart

The hypothesis was tested that endothelin-1 (ET-1)-induced superoxide (O 2 −) generation mediates post-ischemic coronary endothelial injury, that ischemic preconditioning (IPC) affords endothelial protection by preventing post-ischemic ET-1, and thus O 2 −, generation, and that opening of the mitochondrial ATP-dependent potassium channel (mK ATP) triggers the mechanism of IPC. Furthermore, the study was aimed at identifying the source of O 2 − mediating the endothelial injury. Langendorff-perfused guinea-pig hearts were subjected either to 30 min ischemia/35 min reperfusion (IR) or were preconditioned prior to IR with three cycles of either 5 min ischemia/5 min reperfusion or 5 min infusion/5 min washout of mK ATP opener diazoxide (0.5 mM). Coronary flow responses to acetylcholine (ACh) served as a measure of endothelium-dependent vascular function. Myocardial outflow of ET-1 and O 2 − and functional recoveries were followed during reperfusion. NADPH oxidase and xanthine oxidase (XO) activities were measured in cardiac homogenates. IR augmented ET-1 and O 2 − outflow and impaired ACh response. All these effects were attenuated or prevented by IPC and diazoxide, and 5-hydroxydecanoate (a selective mK ATP blocker) abolished the effects of IPC and diazoxide. Superoxide dismutase and tezosentan (a mixed ET-1-receptor antagonist) mimicked the effects of IPC, although they had no effect on the ET-1 generation. IR augmented also the activity of NADPH oxidase and XO. Apocynin treatment, that resulted in NADPH oxidase inhibition, prevented XO activation and O 2 − generation in IR hearts. The inhibition of XO, either by allopurinol or feeding the animals with tungsten-enriched chow, prevented post-ischemic O 2 − generation, although these interventions had no effect on the NADPH activity. In addition, the post-ischemic activation of NADPH oxidase and XO, and O 2 − generation were prevented by IPC, tezosentan, thenoyltrifluoroacetone (mitochondrial complex II inhibitor), and tempol (cell-membrane permeable O 2 − scavenger). In guinea-pig heart: (i) ET-1-induced O 2 − generation mediates post-ischemic endothelial dysfunction; (ii) IPC and diazoxide afford endothelial protection by attenuating the ET-1, and thus O 2 − generation, and the mK ATP opening triggers the protection; (iii) the NADPH oxidase maintains the activity of XO, and the XO-derived O 2 − mediates the endothelial injury, and (iv) ET-1 and O 2 − (probably of mitochondrial origin) are upstream activators of the NADPH oxidase–XO cascade, and IPC prevents the cascade activation and the endothelial dysfunction by preventing the ET-1 generation.

Розробка та підготовка до впровадження нового вітчизняного кардіопротекторного препарату – фторвмісного активатора АТФ-залежних калієвих каналів флокалін

Розробка та підготовка до впровадження нового вітчизняного кардіопротекторного препарату – фторвмісного активатора АТФ-залежних калієвих каналів флокалін

Role of ATP-Dependent Potassium Channels in Pulmonary Vascular Tone of Fetal Lambs With Congenital Diaphragmatic Hernia

High mortality in newborn babies with congenital diaphragmatic hernia (CDH) is principally due to persistent pulmonary hypertension. ATP-dependent potassium (K(ATP)) channels might modulate pulmonary vascular tone. We have assessed the effects of Pinacidil, a K(ATP) channel opener, and glibenclamide (GLI), a K(ATP) channel blocker, in near full-term lambs with and without CDH. In vivo, pulmonary hemodynamics were assessed by means of pressure and blood flow catheters. In vitro, we used isolated pulmonary vessels and immunohistochemistry to detect the presence of K(ATP) channels in pulmonary tissue. In vivo, pinacidil (2 mg) significantly reduced pulmonary vascular resistance (PVR) in both controls and CDH animals. GLI (30 mg) significantly increased pulmonary arterial pressure (PAP) and PVR in control animals only. In vitro, pinacidil (10 microM) relaxed, precontracted arteries from lambs with and without CDH. GLI (10(-5) microM) did not raise the basal tone of vessels. We conclude that activation of K(ATP) channels could be of interest to reduce pulmonary vascular tone in fetal lambs with CDH, a condition often associated with persistent pulmonary hypertension of the newborn.

CAMP-mediated signaling normalizes glucose-stimulated insulin secretion in uncoupling protein-2 overexpressing β-cells

We investigated whether an increase in cAMP could normalize glucose-stimulated insulin secretion (GSIS) in uncoupling protein-2 (UCP2) overexpressing (ucp2-OE) beta-cells. Indices of beta-cell (beta-TC-6f7 cells and rodent islets) function were measured after induction of ucp2, in the presence or absence of cAMP-stimulating agents, analogs, or inhibitors. Islets of ob/ob mice had improved glucose-responsiveness in the presence of forskolin. Rat islets overexpressing ucp2 had significantly lower GSIS than controls. Acutely, the protein kinase A (PKA) and epac pathway stimulant forskolin normalized insulin secretion in ucp2-OE rat islets and beta-TC-6f7 beta-cells, an effect blocked by specific PKA inhibitors but not mimicked by epac agonists. However, there was no effect of ucp2-OE on cAMP concentrations or PKA activity. In ucp2-OE islets, forskolin inhibited ATP-dependent potassium (K(ATP)) channel currents and (86)Rb(+) efflux, indicative of K(ATP) block. Likewise, forskolin application increased intracellular Ca(2+), which could account for its stimulatory effects on insulin secretion. Chronic exposure to forskolin increased ucp2 mRNA and exaggerated basal secretion but not GSIS. In mice deficient in UCP2, there was no augmentation of either cAMP content or cAMP-dependent insulin secretion. Thus, elevating cellular cAMP can reverse the deficiency in GSIS invoked by ucp2-OE, at least partly through PKA-mediated effects on the K(ATP) channel.

Growth-dependent changes in endothelial factors regulating arteriolar tone

Previous studies from this laboratory suggest that during maturation, rapid microvascular growth is accompanied by changes in the mechanisms responsible for regulation of tissue blood flow. To further define these changes, we studied isolated gracilis muscle arterioles from weanling ( approximately 25 days) and juvenile ( approximately 44 days) Sprague-Dawley rats to test the hypothesis that endothelial mechanisms for the control of arteriolar tone are altered with growth. Responses to the endothelium-dependent dilator acetylcholine (ACh) were greater in weanling arterioles (WA) than in juvenile arterioles (JA), whereas there were no consistent differences between age groups in arteriolar responses to other endothelium-dependent agonists (A-23187, vascular endothelial growth factor, and simvastatin). Inhibition of nitric oxide synthase (NOS) with N(omega)-nitro-l-arginine methyl ester (l-NAME) attenuated ACh-induced dilation in JA but not in WA. In JA, combined inhibition of NOS and cyclooxygenase (with indomethacin) reduced the dilator responses to ACh and simvastatin by approximately 90% and approximately 70%, respectively, but had no effect in WA. Cytochrome P450 epoxygenase inhibition [with 2-(propargyloxyphenyl) hexanoic acid] had no effect on responses to ACh or simvastatin in either age group. Inhibition of Ca(2+)-activated or ATP-dependent potassium channels (with tetraethylammonium or glibenclamide, respectively) reduced these arteriolar responses in JA but not those in WA. These findings suggest that in fully grown microvascular networks, endothelium-dependent arteriolar dilation is mediated by the combined release of endothelial nitric oxide and vasodilator prostanoids, and in part through activation of Ca(2+)-activated and ATP-dependent potassium channels. However, during earlier microvascular growth, this dilation is mediated by other factors yet to be identified. This may have significant implications for the regulation of tissue perfusion during microvascular development.

The Knudson’s Two-Hit Model and Timing of Somatic Mutation May Account for the Phenotypic Diversity of Focal Congenital Hyperinsulinism

Congenital hyperinsulinism (CHI) is associated with focal hyperplasia of endocrine tissue in 40-65% of patients. Focal CHI is sporadic and is caused by a germline, paternally inherited, mutation of the SUR1 (ABCC8) or KIR6.2 (KCNJ11) genes (encoding subunits of the pancreatic ATP-dependent potassium channel) together with somatic maternal haploinsufficiency for 11p15.5. Plurifocal or large forms of focal CHI are a cause of apparent failure of surgery, and their underlying mechanism has not been thoroughly investigated. We here report two patients with bifocal CHI as evidenced by relapsing hypoglycemia after removal of the first focal lesion and the detection of a second, distinct, focal adenomatous hyperplasia during later surgery (patients 1 and 2) and a patient with a giant focal lesion involving the major part of the pancreas (patient 3). In the three patients, a germline, paternally inherited, mutation of SUR1 was found. In patients 1 and 2, haploinsufficiency for the maternal 11p15.5 region resulted from a somatic deletion specific for each of the focal lesions, as shown by the diversity of deletion break points. In patient 3, an identical somatic maternal 11p15 deletion demonstrated by similar break points was shown in two independent lesion samples, suggesting a very early event during pancreas embryogenesis. Individual patients with focal hyperinsulinism may have more than one focal pancreatic lesion due to separate somatic maternal deletion of the 11p15 region. These patients and those with solitary focal lesions may follow the two-hit model described by Knudson. The stage of embryogenesis at which the somatic event occurs may account for the observed histological diversity (early event giant lesion, later event small lesion).

Preconditioning modulates pulmonary endothelial dysfunction following ischemia-reperfusion injury in the rat lung: Role of potassium channels

Ischemic preconditioning (IP) may protect the lung from ischemia-reperfusion (I/R) injury following cardiopulmonary by-pass and lung or heart transplantation. The present study was undertaken to investigate the role of ATP-dependent potassium channels (K ATP) in IP in the isolated buffer-perfused rat lung (IBPR) under conditions of elevated pulmonary vasoconstrictor tone (PVT). Since pulmonary arterial perfusion flow and left atrial pressure were constant, changes in pulmonary arterial pressure (PAP) directly reflect changes in pulmonary vascular resistance (PVR). When compared to control value, the pulmonary vasodilator responses to histamine and acetylcholine (ACh) following 2 h of hypothermic ischemia were significantly attenuated, whereas the pulmonary vasodilator response to sodium nitroprusside (SNP) was not altered. IP in the form of two cycles of 5 min of ischemia and reperfusion applied prior to the two-hour interval of ischemia, prevented the decrease in the pulmonary vasodilator responses to histamine and ACh. Pretreatment with glybenclamide (GLB) or HMR-1098, but not 5-hydroxydecanoic acid (5-HD), prior to IP abolished the protective effect of IP. In contrast, GLB or 5-HD did not significantly alter the pulmonary vasodilator response to histamine without IP pretreatment. The present data demonstrate that IP prevents impairment of endothelium-dependent vasodilator responses in the rat pulmonary vascular bed. The present data further suggest that IP may alter the mediation of the pulmonary vasodilator response to histamine and thereby trigger a mechanism dependent on activation of sarcolemmal, and not mitochondrial, K ATP channels to preserve endothelial-dependent vasodilator responses and protect against I/R injury in the lung.

P4-164: Neuroprotective role of ATP dependent potassium channel regulators following ischemia-reperfusion in parietal cerebral cortex of rat

P4-164: Neuroprotective role of ATP dependent potassium channel regulators following ischemia-reperfusion in parietal cerebral cortex of rat

Levosimendan: A Novel Agent in Heart Failure

Heart failure is characterised by decreased cardiac output, which results in the development of both peripheral hypoperfusion and pulmonary congestion and can lead to the development of acute pulmonary edema. The primary objective in treating a patient with decompensated heart failure is hemodynamic stabilization, which is usually achieved by inotropic support. Classic inotropic agents provide short-term hemodynamic improvement, but their use has been correlated with poor prognosis. Levosimendan, a new calcium sensitizer, offers hemodynamic and symptomatic improvement without increasing cAMP and intracellular calcium concentrations. This agent improves contractility without increasing the risk of cardiac events such as arrhythmias. By combining a positive inotropic action mediated via calcium sensitization and a vasodilatory effect via ATP-dependent potassium channels, it appears to be superior than classic positive inotropic agents. Furthermore, it seems to have prolonged benefit in heart failure patients, and it also has anti-inflammatory and antiapoptotic properties. In conclusion, levosimendan seems to be a particularly promising agent for the treatment of decompensated heart failure, as in addition to improving cardiac output, it has a more favorable side-effect profile than classic inotropic agents, and it affects multiple pathways with key role in the pathophysiology of heart failure.

Mitochondrial ATP-sensitive potassium channels and mitochondrial protein kinase C: sometimes it's good to have a close neighbor

the fine study by Jiang et al. ([8][1]) in this issue of the American Journal of Physiology Heart and Circulatory Physiology is an important milestone in the ongoing studies on the role of the mitochondrial ATP-sensitive K+ (mitoKATP) channels in myocardial ischemia. It is not particularly

An overview of potassium channel activators for the treatment of overactive bladder: a survey of new structures 2000 – 2005

For over 25 years, the muscarinic receptor antagonists have been the first-line treatment for overactive bladder (OAB). However, severe dry mouth and exacerbation of disease symptoms with prolonged use can result in compliance rates as low as 30%. Approximately 10 years ago, investigators began to look for alternative approaches to the treatment of overactive bladder. In the recent past, it was demonstrated that ATP-dependent potassium channel openers designed as antihypertensive agents inhibited the hyperactivity associated with the symptoms of overactive bladder. Newer agents targeting other bladder potassium channels and/or bladder-specific channel subtypes represent an exciting new frontier for the effective treatment of OAB, with the potential of having an improved side effect profile compared to the antimuscarinic agents. In this review the authors examine the patent literature surrounding the discovery and development of several novel classes of compounds targeting the various potassium channels associated with urinary bladder activity and excitability. When appropriate, the primary literature is also touched upon to highlight pertinent structure–activity relationships (SAR) associated with these newly disclosed chemotypes.

The cardioprotective effect of uridine and uridine-5′-monophosphate: The role of the mitochondrial ATP-dependent potassium channel

The activity of mitochondrial ATP-dependent potassium channel (mitoK ATP) of rat heart and liver mitochondria was shown to decrease during aging. This partially explains the increase of risk of ischemia at a mature age since mitoK ATP activation provides cardioprotection. We demonstrated that uridine-5′-diphosphate (UDP) possesses the property to activate mitoK ATP. At a concentration of 30 μM, it reactivated mitoK ATP in mitochondria, and 5-hydroxydecanoate (5-HD) eliminated this effect. In experimental animals, UDP precursors uridine and uridine-5′-monophosphate (UMP) (both 30 mg/kg, administered intravenously 5 min before coronary occlusion) decreased the myocardium ischemic alteration index (1.9 and 3.5 times, respectively) and the T-wave amplitude within 60 min after occlusion. Both effects were inhibited by Glibenclamide (Glib) and 5-HD. UMP and uridine decreased the number of premature ventricular beats 5.6 and 1.9 times and the duration of ventricular tachycardia 9.4 and 4.1 times, respectively. Glib and 5-HD inhibited the anti-arrhythmic parameters, 5-HD being less effective. Uridine and UMP decreased the duration of fibrillation 10.8 and 3.6 times, respectively, and this effect was not abolished by Glib and 5-HD. Thus, uridine and UMP, which are the precursors of UDP in the cell, possess cardioprotective properties. MitoK ATP prevents mainly ischemic injuries and partially rhythm disorders.

The Role of Potassium Channels in Relaxant Effect of Levosimendan in Rat Small Mesenteric Arteries

We investigated both the effect of levosimendan and the role of various potassium channels in KCl-precontracted rat small mesenteric arteries. Levosimendan (10(-6)-10(-3) M) or cromakalim (CRO, 10(-7)-10(-4) M) produced concentration-dependent relaxation responses in small mesenteric arteries precontracted by 30 mM KCl. The relaxant responses to levosimendan in KCl-precontracted arteries did not differ significantly between endothelium-intact and endothelium-denuded preparations. Incubation of rat small mesenteric arterial segments with ATP-dependent potassium channel (KATP) blocker glibenclamide (GLI, 10(-6) M) for 30 min significantly inhibited the relaxant responses to both levosimendan and CRO. Neither the Ca(2+)-activated potassium channel (KCa) blocker iberiotoxin (10(-7) M) nor the voltage-dependent potassium channel (KV) blocker 4-aminopyridine (5 mM) incubation for 30 min caused significant alterations in relaxant responses to levosimendan in KCl-precontracted small mesenteric arteries. These findings suggested that levosimendan-induced relaxation responses in isolated rat small mesenteric arteries were neither depended on endothelium nor inhibited by the blockers of KV or KCa but, they rather seem to depend on the activation of KATP.

Combined modulation of the mitochondrial ATP-dependent potassium channel and the permeability transition pore causes prolongation of the biphasic calcium dynamics

The permeability transition pore (PTP) and the ATP-dependent potassium (mtK-ATP) channel of mitochondria are known to play key roles in mitochondrially mediated apoptosis. We investigated how modulation of the permeability transition pore (PTP) and the ATP-dependent potassium (mtK-ATP) channel, either as single elements or in combination, affects the proapoptotic intracellular calcium ([Ca 2+] i) transients and the mitochondrial membrane potential ( ψ m). For this purpose a model was established exploring the [Ca 2+] i transients in N2A cells using continuous application of ATP that causes a biphasic [Ca 2+] i response. This response was sensitive to endoplasmatic reticulum (ER) Ca 2+ depletion and a smooth ER Ca 2+-ATPase (SERCA) antagonist. PTP inhibition by cyclosporine A (CsA) or its non-immunosuppressive derivative NIM811 caused an amplification of the secondary [Ca 2+] i peak and induced a hyperpolarization of ψ m. Both the putative mtK-ATP channel inhibitor 5-hydroxydecanoate (5-HD) and the opener diazoxide ameliorated the ATP-induced secondary [Ca 2+] i peak. The effect of diazoxide was accompanied by a depolarization of ψ m whereas 5-HD had no effect on ψ m. When diazoxide and CsA or NIM811 were applied together the secondary [Ca 2+] i rise did not return to baseline and a not significant hyperpolarization of ψ m was observed. So, simultaneous inhibition of PTP and activation of the mtK-ATP channel prevents the increased slope of the secondary [Ca 2+] i peak induced by CsA (or NIM811) and also the depolarization after diazoxide application. Hence, we propose that modulation of one of these channels leads to functional changes of the other channel by means of Δ[Ca 2+] i and Δ ψ m.

Brief antecedent anoxia preserves mitochondrial function after sustained undersupply: A subcellular correlate to ischemic preconditioning?

There is increasing evidence that mitochondria - owning a high degree of autonomy within the cell - might represent the target organelles of the myocardial protection afforded by ischemic preconditioning. It was the aim of the study to investigate a possible subcellular correlate to ischemic preconditioning at the mitochondrial level. In addition, we tested whether this protection depends on mitochondrial ATP-dependent potassium channels (K (ATP)) and an might involve an attenuation of mitochondrial ATP hydrolysis during sustained anoxia. Sustained anoxia (A, 14 min) and reoxygenation (R) completely inhibited state 3 and state 4 respiration of isolated ventricular mitochondria from Wistar rats. An antecedent brief anoxic incubation (4 min) followed by reoxygenation (2 min) prevented this loss of mitochondrial function. The protection afforded by anoxic preconditioning could be mimicked by the K (ATP) opener diazoxide (30 micromol/l) and was completely inhibited by the K (ATP) blocker 5-hydroxydecanoic acid (300 micromol/l). Structural mitochondrial integrity, as estimated from externalization of the mitochondrial enzymes creatine kinase and glutamateoxalacetate transaminase, remained unchanged between the groups, as did mitochondrial ATP loss during anoxia. For the first time, we provide direct evidence for a subcellular preconditioning-like functional mitochondrial adaptation to sustained anoxia. This effect apparently depends on opening of K(ATP) but is independent of ATP preservation.