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PubMedDate not supplied

Subcellular heterogeneity in mitochondrial red-ox responses to KATP channel agonists in freshly isolated rabbit cardiomyocytes.

We have used the technique of fluorescent microscopy imaging supplemented with the refined analysis of temporal cartography of the cell fluorescence to investigate the mechanisms of regulation of mitochondrial function and its red-ox state in cardiac cells in vivo. Autofluorescence of flavoproteins of the respiratory chain in the isolated rabbit cardiomyocytes was registered before and after application of mitochondrial KATP channel opener diazoxide (100 and 400 microM). Diazoxide addition resulted in oxidation of flavoproteins. Detailed analysis of these responses showed that they were heterogeneous over space and time. The local responses show rapid jumps. In a few cells, metabolic oscillations developed and could be recorded for tens of minutes. Under these conditions the cells appeared divided into a small number of regions in which mitochondria function synchronously. Local pattern of oxidation switches again and again from a reduced state to the same level of oxidation. All these phenomena where absent when the cells were permeabilized by saponin giving a direct access to mitochondrial KATP channel opener. Cross-correlation analysis revealed a high degree of homogeneity for cells presenting metabolic oscillations, contrarily to those displaying a smooth increase in fluorescence in response to diazoxide. The results are consistent with the view that mitochondria form independent functional units whose behaviour can be synchronised by some unknown cellular factors or metabolites.

Animals↗
PubMed2006

Enhanced PIP3 signaling in POMC neurons causes KATP channel activation and leads to diet-sensitive obesity.

Leptin and insulin have been identified as fuel sensors acting in part through their hypothalamic receptors to inhibit food intake and stimulate energy expenditure. As their intracellular signaling converges at the PI3K pathway, we directly addressed the role of phosphatidylinositol3,4,5-trisphosphate-mediated (PIP3-mediated) signals in hypothalamic proopiomelanocortin (POMC) neurons by inactivating the gene for the PIP3 phosphatase Pten specifically in this cell type. Here we show that POMC-specific disruption of Pten resulted in hyperphagia and sexually dimorphic diet-sensitive obesity. Although leptin potently stimulated Stat3 phosphorylation in POMC neurons of POMC cell-restricted Pten knockout (PPKO) mice, it failed to significantly inhibit food intake in vivo. POMC neurons of PPKO mice showed a marked hyperpolarization and a reduction in basal firing rate due to increased ATP-sensitive potassium (KATP) channel activity. Leptin was not able to elicit electrical activity in PPKO POMC neurons, but application of the PI3K inhibitor LY294002 and the KATP blocker tolbutamide restored electrical activity and leptin-evoked firing of POMC neurons in these mice. Moreover, icv administration of tolbutamide abolished hyperphagia in PPKO mice. These data indicate that PIP3-mediated signals are critical regulators of the melanocortin system via modulation of KATP channels.

Animals↗
PubMed1993

Adenosine A1 receptors, KATP channels, and ischemic preconditioning in dogs.

The objective of the present study was to characterize the role of adenosine in myocardial ischemic preconditioning in the canine heart. Preconditioning with 5 min of ischemia resulted in a marked reduction in infarct size after 60 min of left circumflex coronary artery occlusion and 5 h of reperfusion in barbital-anesthetized dogs compared with dogs that were not preconditioned (4.8 +/- 1.9 vs. 27.9 +/- 4.5%; P < 0.05). Pretreatment with either the nonselective adenosine receptor antagonist PD 115199 or the selective adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine blocked this protective effect, although in the absence of preconditioning neither of the antagonists affected infarct size. Intracoronary infusion of two different doses of adenosine or dipyridamole over a 5-min period before a prolonged 60-min occlusion period did not mimic preconditioning; however, intracoronary infusion of a combination of adenosine and dipyridamole produced a significant reduction in infarct size (13.6 +/- 4.1%), which was abolished by pretreatment with the ATP-dependent potassium (KATP) channel antagonist glibenclamide. These results suggest that activation of adenosine A1 receptors produces myocardial preconditioning in the canine heart by opening KATP channels.

Adenosine↗
PubMed2003

Antiangiogenic effect of KR-31372 by apoptosis via mediation of mitochondrial KATP channel opening and the phosphatase and tensin homolog deleted from chromosome 10 phosphorylation.

Antiangiogenic action of (2R,3R,4S)-N"-cyano-N-(6-nitro-3,4-dihydro-hydroxy-2-methyl-2-dimethoxymethyl-2H-1-benzopyran-4yl)-N'-benzyl guanidine (KR-31372) was examined with its proapoptotic action in human umbilical vein endothelial cells (HUVECs) compared with diazoxide. KR-31372 as well as diazoxide significantly suppressed the neovascularization in mice induced by the Matrigel-containing recombinant human vascular endothelial growth factor (VEGF)165 in vivo and the basal tube formation of HUVECs in vitro with suppression of proliferation of HUVECs stimulated by VEGF165. KR-31372 and diazoxide enhanced DNA fragmentation associated with increase in phosphatase and tensin homolog deleted from chromosome 10 (PTEN) and decrease in serine/threonine kinase phosphorylation, which were accompanied by augmented Bax and cytochrome c release, and suppressed Bcl-2 in HUVECs. In the U87-MG cells, when transfected with expression vectors for sense PTEN, KR-31372 enhanced DNA fragmentation, but not in naive U87-MG cells. The suppression by KR-31372 and diazoxide of these variables was significantly antagonized by 5-hydroxydecanoic acid, a mitochondrial KATP channel blocker. Taken together, KR-31372 strongly inhibited angiogenesis in HUVECs by proapoptotic mechanism via mediation of 5-hydroxydecanoic acid-inhibitable mitochondrial KATP channel opening and PTEN phosphorylation.

Angiogenesis Inhibitors↗
PubMed2000

Norepinephrine acts on the KATP channel and produces different effects on [Ca2+]i in oscillating and non-oscillating HIT-T15 cells.

Norepinephrine (NE) is an inhibitor of insulin secretion that acts, in part, by decreasing intracellular free calcium ([Ca2+]i). We examined the effects of NE on [Ca2+]i in individual HIT-T15 cells loaded with indo 1. Cells were categorized as oscillators or non-oscillators on the basis of the pattern of the calcium response to glucose and the effect of NE on [Ca2+]i was subsequently measured in each cell. NE caused a simple decrease in [Ca2+]i in nonoscillators. In oscillators, NE decreased the amplitude and frequency of the oscillations. Furthermore, the duration of the NE effect in oscillators was longer than in non-oscillators. NE did not affect the rise in [Ca2+]i elicited by depolarizing concentrations of 20 mM or 35 mM KCl alone, or in the presence of 20 mM KCl, 100 microM diazoxide, and 10 mM glucose. In other experiments, NE had no effect on [Ca2+]i when the KATP channels were fully clamped with diazoxide or tolbutamide. We conclude that the action of NE to decrease [Ca2+]i in both oscillators and non-oscillators is mediated via activation of the KATP channel. Despite this common mechanism, NE exerts different effects on oscillating and non-oscillating cells.

Adenosine Triphosphate↗
PubMed2003

Quinaprilat reduces myocardial infarct size involving nitric oxide production and mitochondrial KATP channel in rabbits.

This study examined whether quinaprilat, an angiotensin-converting enzyme inhibitor, reduces the infarct size, and investigated the mechanisms for its infarct size-reducing effect, in rabbits. Japanese white rabbits underwent 30 min of ischemia and 48 h of reperfusion. Quinaprilat (100 microg/kg/h or 300 microg/kg/h for 70 min, IV) was administered 20 min before ischemia with or without pretreatment with Nomega-nitro-l-arginine methyl ester (l-NAME) (10 mg/kg, IV, a nitric oxide synthase inhibitor), 5-hydroxydecanoic acid sodium salt (5-HD) or posttreatment with 5-HD (5 mg/kg, IV, a mitochondrial KATP channel blocker). The area at risk as a percentage of the left ventricle was determined by Evans blue dye and the infarct size was determined as a percent of the area at risk by triphenyl tetrazolium chloride staining. Using a microdialysis technique, myocardial interstitial levels of 2,5-dihydroxybenzoic acid (2,5-DHBA), an indicator of hydroxyl radicals, and NOx, an indicator of nitric oxide, were measured before, during, and after 30 min of ischemia. Quinaprilat significantly reduced the infarct size in a dose-dependent manner (30.1 +/- 3%, n = 10, and 27.6 +/- 2%, n = 7, respectively) compared with the control (46.5 +/- 4%, n = 10). The infarct size-reducing effect of quinaprilat was completely blocked by pretreatment with l-NAME (43.8 +/- 2%, n = 8) and 5-HD (50.1 +/- 3%, n = 8) and posttreatment with 5-HD (50.3 +/- 2%, n = 8), respectively. Quinaprilat did not affect the myocardial interstitial 2,5-DHBA level but significantly increased the NOx level during ischemia and reperfusion. Quinaprilat reduces myocardial infarct size involving NO production and mitochondrial KATP channels in rabbits without collateral circulation.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2004

Sevoflurane-induced preconditioning of rat brain in vitro and the role of KATP channels.

In the present study we tested the ability of the inhalation anesthetic sevoflurane to induce preconditioning against hypoxia in vitro. Rat hippocampal slices were prepared using established procedures. After 90 min of incubation, slices were exposed for 30 min to 0, 1, 2 or 3 minimum alveolar concentration (MAC) of sevoflurane under normoxic conditions (95% O2/5% CO2). Fifteen minutes later, slices were exposed to 13-min hypoxia (95% N2/5% CO2) followed by 30-min reoxygenation. The amplitude of extracellularly recorded, orthodromically evoked, CA1 population spikes (neuronal function) at the end of the reoxygenation period was measured and used to quantify the degree of recovery of neuronal function posthypoxia. To assess the role that the mitochondrial KATP channel plays in preconditioning, its blocker, 5-hydroxydecanoic acid (5-HD), was added during sevoflurane exposure. Sevoflurane-preconditioning with 1, 2 and 3 MAC increased the degree of recovery of neuronal function after 13-min hypoxia and 30-min reoxygenation from 51 +/- 1% (0 MAC), to 55 +/- 3%, 63 +/- 3%, and 72 +/- 2%, respectively. The effect of 3 MAC sevoflurane was blocked by 5-HD (53 +/- 3%), whereas 5-HD alone had no effect (48 +/- 3%) on the recovery of neuronal function from hypoxia. It is concluded that sevoflurane is capable of inducing preconditioning in vitro in a dose-dependent fashion and involves activation of mitochondrial KATP channels.

Adenosine Triphosphate↗
PubMed1999

ATP inhibition of KATP channels: control of nucleotide sensitivity by the N-terminal domain of the Kir6.2 subunit.

1. To gain insight into the role of the cytoplasmic regions of the Kir6.2 subunit in regulating channel activity, we have expressed the sulphonylurea receptor SUR1 with Kir6.2 subunits containing systematic truncations of the N- and C-termini. Up to 30 amino acids could be truncated from the N-terminus, and up to 36 amino acids from the C-terminus without loss of functional channels in co-expression with SUR1. Furthermore, Kir6.2DeltaC25 and Kir6. 2DeltaC36 subunits expressed functional channels in the absence of SUR1. 2. In co-expression with SUR1, N-terminal truncations increased Ki,ATP ([ATP] causing half-maximal inhibition of channel activity) by as much as 10-fold, accompanied by an increase in the ATP-insensitive open probability, whereas the C-terminal truncations did not affect the ATP sensitivity of co-expressed channels. 3. A mutation in the near C-terminal region, K185Q, reduced ATP sensitivity of co-expressed channels by approximately 30-fold, and on the Kir6.2DeltaN2-30 background, this mutation decreased ATP sensitivity of co-expressed channels by approximately 400-fold. 4. Each of these mutations also reduced the sensitivity to inhibition by ADP, AMP and adenosine tetraphosphate. 5. The results can be quantitatively explained by assuming that the N-terminal deletions stabilize the ATP-independent open state, whereas the Kir6.2K185Q mutation may alter the stability of ATP binding. These two effects are energetically additive, causing the large reduction of ATP sensitivity in the double mutant channels.

ATP-Binding Cassette Transporters↗
PubMed1999

Inward rectification in KATP channels: a pH switch in the pore.

Inward-rectifier potassium channels (Kir channels) stabilize the resting membrane potential and set a threshold for excitation in many types of cell. This function arises from voltage-dependent rectification of these channels due to blockage by intracellular polyamines. In all Kir channels studied to date, the voltage-dependence of rectification is either strong or weak. Here we show that in cardiac as well as in cloned KATP channels (Kir6.2 + sulfonylurea receptor) polyamine-mediated rectification is not fixed but changes with intracellular pH in the physiological range: inward-rectification is prominent at basic pH, while at acidic pH rectification is very weak. The pH-dependence of polyamine block is specific for KATP as shown in experiments with other Kir channels. Systematic mutagenesis revealed a titratable C-terminal histidine residue (H216) in Kir6.2 to be the structural determinant, and electrostatic interaction between this residue and polyamines was shown to be the molecular mechanism underlying pH-dependent rectification. This pH-dependent block of KATP channels may represent a novel and direct link between excitation and intracellular pH.

ATP-Binding Cassette Transporters↗
PubMed1995

Isolation and chromosomal localization of a human ATP-regulated potassium channel.

KATP channels are K+ channels whose activity is inhibited by the presence of and enhanced by the absence of cytosolic ATP. This property allows KATP channels to sense cellular intermediary metabolism and directly couple this information to the modulation of membrane excitability. Indeed, recent studies from our laboratory and others have suggested that activation of KATP channels during anoxia is important in the response and adaptation of central neurons to hypoxia. In order to identify KATP channels from human brain, we performed a polymerase chain reaction (PCR) using human cerebral cortex mRNA and primers derived from the ROMK1 sequence, a cDNA clone encoding an ATP-regulated potassium channel, recently isolated from rat kidney. We thus identified a novel 308-bp PCR product, pKCNJ1, whose expression was found to be restricted to a 3.0-kb band in the kidney by probing a human multiple tissue northern blot, pKCNJ1 was then used to isolate genomic clones and, using fluorescence in situ hybridization (FISH) to human metaphase chromosomes, was mapped to chromosome 11q.

Adenosine Triphosphate↗
PubMed1999

Myocardial protection by insulin is dependent on phospatidylinositol 3-kinase but not protein kinase C or KATP channels in the isolated rabbit heart.

Because tyrosine kinase blockade prevents protection by ischemic preconditioning (p.c.) in several species, activation of tyrosine kinase appears to be critical for cardioprotection. The tyrosine kinase's identity, however, is unknown. The present study tested whether activation of a receptor tyrosine kinase, the insulin receptor, could mimic p.c. and if the mechanism of protection was similar to that of p.c. Isolated rabbit hearts were subjected to 30 min of regional ischemia and 2 h of reperfusion. Infarct size was determined by triphenyltetrazolium staining and expressed as a percentage of the area at risk. Infarct size in control hearts was 32.6 +/- 2.3%. A 5-min infusion of insulin (5 mU/ml) followed by a 10-min washout period prior to ischemia significantly reduced infarction to 14.7 +/- 2.1% (P < 0.05). The tyrosine kinase inhibitor genistein (50 microM) given around the insulin infusion blocked protection (28.9 +/- 2.8%). However, when present during the onset of ischemia, genistein had no effect on protection triggered by insulin (14.0 +/- 2.4%; P < 0.05). Inhibition of either PKC by polymyxin B (50 microM) or KATP channels by 5-hydroxydecanoate (100 microM) also failed to prevent protection by insulin (17.5 +/- 3.2% and 17.6 +/- 3.0%, respectively). However, the reduction in infarct size by insulin was significantly attenuated by wortmannin (100 nM), a selective inhibitor of phosphatidylinositol 3-kinase (PI3K, 28.3 +/- 2.2%). Insulin was still able to protect the heart when given only during the reperfusion period (13.2 +/- 3.4%). P.c. reduced infarction to 12.8 +/- 2.0% (P < 0.05) and still offered significant protection in the presence of wortmannin (22.1 +/- 2.4%; P < 0.05). In conclusion, activation of the insulin receptor reduces infarct size in the rabbit heart even when instituted upon reperfusion. However, the mechanism of protection is quite different from that of p.c. and involves activation of PI3K but not PKC or KATP channels.

Animals↗
PubMed2003

Combined pharmacological preconditioning with a G-protein-coupled receptor agonist, a mitochondrial KATP channel opener and a nitric oxide donor mimics ischaemic preconditioning.

1. Although pharmacological preconditioning (PPC) has emerged as an alternative to ischaemic preconditioning (IPC) in cardioprotection, the efficacy of PPC compared with IPC has not been investigated. Because IPC is mediated by complex signalling cascades arising from multiple triggers, we have hypothesized that combined PPC is necessary to mimic IPC. 2. Isolated and perfused rat hearts underwent IPC by three cycles of 5 min ischaemia and 5 min reperfusion before 30 min global ischaemia followed by 120 min reperfusion. Adenosine (30 micromol/L), diazoxide (50 micromol/L) and s-nitroso-N-acetylpenicillamine (SNAP; 50 micromol/L) were added for 25 min just before (pretreatment modality) or 45 min before (PPC modality) the index ischaemia. 3. Ischaemic preconditioning significantly improved isovolumic left ventricular (LV) function and reduced infarct size. Although pretreatment with adenosine, diazoxide or SNAP alone was capable of reducing infarct size, PPC with each drug alone or in a combination of two drugs except for diazoxide plus SNAP failed to reduce infarct size. In contrast, PPC in combination with adenosine, diazoxide and SNAP (triple combination PPC) conferred significant improvement of LV function and reduction of infarct size that was as effective as IPC. 4. Cardioprotection afforded by triple combination PPC was abolished by the Gi/o-protein inhibitor pertussis toxin, the mitochondiral KATP channel inhibitor 5-hydroxydecanoate or the nitric oxide (NO) scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethyl imidazoline-1-oxyl 3-oxide (carboxy-PTIO). 5. Protein kinase C (PKC)-epsilon in the particulate fraction was activated throughout preconditioning ischaemia and reperfusion. Although PKC-epsilon was activated during treatment with adenosine, diazoxide or SNAP alone, it was inactivated after washout. In contrast, PKC-epsilon remained activated after triple combination PPC. The PKC inhibitor chelerythrine abolished activation of PKC-epsilon and cardioprotection afforded by IPC and triple combination PPC. 6. These results demonstrate that combined PPC with a G-protein-coupled receptor agonist, a mitochondrial KATP channel opener and an NO donor is necessary to mimic IPC and such synergistic cardioprotection is associated with enhanced and sustained activation of PKC-epsilon.

Adenosine Triphosphate↗
PubMed1991

ATP dependence of KATP channel kinetics in isolated membrane patches from rat ventricle.

The dependence of KATP channel activity on [ATP] has been examined in isolated membrane patches from rat ventricular myocytes. The steady-state [ATP] dependence of channel open probability could be described by a sigmoidal relationship with the ki ([ATP] causing half-maximal inhibition of open probability) = 25 microM and Hill coefficient of 2. Description of channel open- and closed-time distributions required at least 2, and 3, time constants, respectively. Long open-channel lifetimes decreased with [ATP]; unconditional mean channel closed-times increased with [ATP]. Step decrease (jump) in bathing [ATP] resulted in a delay (of up to hundreds of milliseconds) followed by a pseudo-exponential rise of current (with a time constant of up to hundreds of milliseconds). The time course of channel current after changes of [ATP] (or the ATP-analogue AMP-PNP) was shown to be predominantly determined by the time course of diffusion into the tip of the electrode and to the membrane. This time course of diffusion of ATP into the pipette tip had to be taken into account when analyzing the current response to [ATP] steps. Several possible kinetic models of the ATP-dependent regulation of channel activity were considered. Adequate explanation of the data required a model with sequential ATP-binding sites. The model can account for the time course of channel opening after steps of [ATP], as well as for the steady-state dependence of P0 on [ATP]. The model predicts [ATP]-dependent closed and open lifetimes as were observed experimentally.

Adenosine Triphosphate↗
PubMed1998

Activation of KATP channels by Na/K pump in isolated cardiac myocytes and giant membrane patches.

Strophanthidin inhibits KATP channels in 2,4-dinitrophenol-poisoned heart cells (). The current study shows that the Na/K pump interacts with KATP current (IK-ATP) via submembrane ATP depletion in isolated giant membrane patches and in nonpoisoned guinea pig cardiac cells in whole-cell configuration. IK-ATP was inhibited by ATP, glibenclamide, or intracellular Cs+. Na/K pump inactivation by substitution of cytoplasmic Na+ for Li+ or N-methylglucamine decreased both IK-ATP by 1/3 (1 mM ATP, zero calcium), and IC50 of ATP for IK-ATP (0.3 +/- 0.1 mM) by 2/5. The Na+/Li+ replacement had no effect on IK-ATP at low pump activity ([ATP] </= 0.1 mM or 100 microM ouabain) or when IK-ATP was completely inhibited by 10 mM ATP. In whole-cell configuration, ouabain inhibited up to 60% of inwardly rectifying IK-ATP at 1 mM ATP in the pipette but not at 10 mM ATP and 10 mM phosphocreatine when IK-ATP was always blocked. However, mathematical simulation of giant-patch experiments revealed that only 20% of ATP depletion may be attributed to the ATP concentration gradient in the bulk solution, and the remaining 80% probably occurs in the submembrane space.

Adenosine Triphosphate↗
PubMed1999

A KATP-channel opener as a potential treatment modality for erectile dysfunction.

OBJECTIVES: To assess the effects of pinacidil (a KATP-channel opener) for the treatment of penile erectile dysfunction and to examine the role of the K+-channel in cavernosal smooth muscle contractility. MATERIALS AND METHODS: Using a feline model, the magnitude of penile erection caused by pinacidil was compared with that caused by erectogenic drugs, e.g. acetylcholine, prostaglandin E1 (PGE1) and L-arginine. The effects of K+-channel blockers (4-aminopyridine, glibenclamide and tetraethylammonium) and pinacidil on penile erections induced by the drugs were investigated. RESULTS: The intra-arterial injection of pinacidil caused a dose-dependent increase in intracavernosal pressure (ICP) and the increase in ICP induced by pinacidil with acetylcholine, PGE1 or L-arginine was more pronounced than with the compounds alone. Furthermore, pinacidil (1 mmol/L) effectively reversed the inhibitory effects of the K+-channel blockers on the cavernosal relaxation induced by acetylcholine, PGE1 or L-arginine (P<0.01). Notably, pinacidil induced cavernosal relaxation after injecting the drugs even in cases refractory to higher concentrations (0.1 mol/L) of the drugs (n=11, P<0.01). CONCLUSIONS: These results suggest that pinacidil is effective in relaxing feline erectile tissue in vivo, probably via increased K+ permeability and subsequent hyperpolarization. Further comparative studies with erectogenic compounds on human erectile tissue and clinical testing are required to determine whether K+-channel openers can be used in the diagnosis and treatment of erectile dysfunction. However, pinacidil seems promising as an intracavernosal agent combined with PGE1 to produce synergistic effects.

Animals↗
PubMed2006

KATP channel openers of the benzopyran type reach their binding site via the cytosol.

BACKGROUND AND PURPOSE: ATP-sensitive K+ (KATP) channels are composed of pore-forming subunits (Kir6.x) and of sulphonylurea receptors (SUR). Both sulphonylureas and K(ATP) channel openers act by binding to SUR. Sulphonylureas reach their binding site from the cytosol but it remains unknown whether this holds for openers too. EXPERIMENTAL APPROACH: A poorly membrane-permeant sulphonic acid derivative of the benzopyran-type opener, bimakalim, was synthesized, descyano-bimakalim-6-sulphonic acid (BMSA). Binding of BMSA and bimakalim was compared in membranes and intact cells expressing the Kir6.2/SUR2B channel and channel opening was compared in inside-out patches and whole cells. KEY RESULTS: In membranes, bimakalim and BMSA bound to Kir6.2/SUR2B with Ki values of 61 nM and 4.3 microM, showing that the negative charge decreased affinity 69-fold. In intact cells, however, binding of BMSA was much weaker than in membranes (75-fold) whereas that of bimakalim was unchanged. The Ki value of BMSA decreased with increasing incubation time. In inside-out patches, bimakalim (1 microM) and BMSA (100 microM) opened the Kir6.2/SUR2B channel closed by MgATP to a similar degree whereas in whole-cell experiments, only bimakalim was effective. CONCLUSIONS AND IMPLICATIONS: Despite its negative charge, BMSA is an effective channel opener. The fact that BMSA binds and acts more effectively when applied to the inner side of the cell membrane shows that benzopyran openers reach their binding site at SUR from the cytosol. This suggests that the binding pocket of SUR is only open on the cytoplasmic side.

Benzopyrans↗
PubMed2004

Diazoxide causes early activation of cardiac sarcolemmal KATP channels during metabolic inhibition by an indirect mechanism.

OBJECTIVE: We have used isolated myocytes to investigate the effects of diazoxide on sarcolemmal KATP channel (sarcoKATP) activity and action potential failure during metabolic inhibition, and the role of these channels in protection of functional recovery on reperfusion. MATERIALS AND METHODS: Isolated adult rat ventricular myocytes were exposed to metabolic inhibition (NaCN and iodoacetate) and reperfusion. Functional recovery was assessed from the ability of cells to contract on electrical stimulation and to recover calcium homeostasis, measured with fura-2. Action potentials and KATP currents were measured using patch clamp. RESULTS: Pretreatment with diazoxide (100 microM, 5 min) increased the proportion of cells that recovered contractile function after MI and reperfusion from 16.8 +/- 2.4% to 65.0 +/- 2.2% (p<0.001) and the proportion of cells in which [Ca2+]i recovered to <250 nM. Pretreatment also accelerated action potential and contractile failure during MI. In cell-attached patches, MI activated sarcoKATP channels after 224 +/- 11 s, and diazoxide pretreatment decreased this to 145 +/- 24 s (p<0.01). However, diazoxide present in the patch pipette did not accelerate sarcoKATP channel activation. Intracellular Mg2+ rose earlier in diazoxide-pretreated cells. The sarcoKATP blocker HMR 1883 delayed action potential failure and reduced diazoxide protection. CONCLUSIONS: Diazoxide pretreatment increases recovery of function and [Ca2+]i following reperfusion. Protection is coupled with early action potential failure, due to early activation of sarcoKATP channels during metabolic inhibition (MI), which is likely to involve an indirect effect of diazoxide.

Action Potentials↗
PubMed1999

Eukaliuric diuresis and natriuresis in response to the KATP channel blocker U37883A: micropuncture studies on the tubular site of action.

1. Systemic application of U37883A, a blocker of ATP sensitive potassium (KATP) channels, elicits diuresis and natriuresis without significantly altering urinary potassium excretion. 2. To elucidate tubular sites of action upstream to the distal nephron, micropuncture experiments were performed in nephrons with superficial glomeruli of anaesthetized Munich-Wistar-Frömter rats during systemic application of U37883A (1, 5 or 15 mg kg-1 i.v.). 3. The observed eukaliuric diuresis and natriuresis in response to U37883A at 15 mg kg-1 was accompanied by an increase in early distal tubular flow rate (VED) from 10 - 18 nl min(-1) reflecting a reduction in fractional reabsorption of fluid up to this site (FR-fluid) of 13%. The latter proposed an effect on water-permeable segments such as the proximal tubule which could fully account for the observed reduction in fractional reabsorption of Na+ up to the early distal tubule (FR-Na+) of 8% and the increase in early distal tubular Na+ concentration ([Na+]ED) from 35 - 51 mM whereas [K+]ED was left unaltered. 4. In comparison, furosemide (3 mg kg-1 i.v.), which acts in the water-impermeable thick ascending limb, elicited diuresis, natriuresis and kaliuresis which were associated with a fall in FR-Na+ of 10% with no change in FR-fluid, and a rise in [Na+]ED from 42 - 117 mM and [K+]ED from 1.2 - 5.7 mM with no change in VED. 5. Direct late proximal tubular fluid collections confirmed a significant inhibition of fluid reabsorption in proximal convoluted tubule in response to systemic application of U37883A. 6. These findings suggest that the diuretic and natriuretic effect upstream to the distal tubule in response to systemic application of U37883A involves actions on water-permeable segments such as the proximal convoluted tubule.

Adamantane↗