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PubMed2005

Antidiabetic activity and toxicity of Zizyphus spina-christi leaves.

The effect of the butanol extract of Zizyphus spina-christi (L.), Willd (Rhamnaceae) leaves and its major saponin glycoside, christinin-A, on the serum glucose and insulin levels was studied in non-diabetic control, type-I (insulin-dependent) and type-II (non-insulin-dependent) diabetic rats. Pretreatment either with 100 mg/kg butanol extract or christinin-A potentiated glucose-induced insulin release in non-diabetic control rats. In type-II but not in type-I diabetic rats pretreatment with the butanol extract or christinin-A improved the oral glucose tolerance and potentiated glucose-induced insulin release. Treatment either with 100 mg/kg butanol extract or christinin-A reduced the serum glucose level and increased the serum insulin level of non-diabetic control and type-II diabetic rats but not of type-I diabetic rats. Effects of the butanol extract and christinin-A were similar. Pretreatment of non-diabetic control and type-II diabetic rats either with 100 mg/kg butanol extract or christinin-A enhanced the glucose lowering and insulinotropic effects of 5 g/kg glibenclamide. The hyperglycemic and hypoinsulinemic effects of 30 mg/kg diazoxide in non-diabetic control and type-II diabetic rats were inhibited and antagonized, respectively by pretreatment with the butanol extract or christinin-A. The relaxant effects of different concentrations of diazoxide on the isolated norepinephrine-contracted aortic strips were inhibited by 100 micromol/l christinin-A or 10 micromol/l glibenclamide. The combination of glibenclamide and christinin-A led to complete inhibition of the relaxant effects of different concentrations of diazoxide. At a dose level much higher than that required to produce satisfactory insulinotropic and hypoglycemic effects, the butanol extract of Zizyphus spina-christi leaves produced a depressant effect on the central nervous system in rats. Treatment of rats with 100mg/kg butanol extract for 3 months produced no functional or structural disturbances in liver and kidney and no haematological changes. In addition, the oral LD50 of the butanol extract in mice was 3820 mg/kg, while that of glibenclamide was 3160 mg/kg. Thus, Zizyphusspina-christi leaves appears to be a safe alternative to lower blood glucose. The safe insulinotropic and subsequent hypoglycemic effects of Zizyphus spina-christi leaves may be due to a sulfonylurea-like activity.

Animals↗
PubMed1999

Pharmacological and molecular characterization of ATP-sensitive K+ channels in the TE671 human medulloblastoma cell line.

ATP-sensitive K+ (K(ATP)) channels in the human medulloblastoma TE671 cell line were characterized by membrane potential assays utilizing a potentiometric fluorescent probe, bis-(1,3-dibutylbarbituric acid)trimethine oxonol (DiBAC4(3)), and by mRNA analysis. Membrane potential assays showed concentration-dependent and glyburide-sensitive changes in fluorescence upon addition of (-)-cromakalim, pinacidil, diazoxide and P1075. The rank order of potency for these openers was P1075 > (-)-cromakalim approximately = pinacidil > diazoxide. Additionally, glyburide and glipizide inhibited P1075-evoked responses in TE671 cells with half-maximal inhibitory concentrations of 0.22 and 14 microM, respectively. The rank order potencies of both openers and inhibitors were similar to those observed in the rat smooth muscle A-10 cell line. In contrast, in the rat pancreatic insulinoma RIN-m5F cell line, only diazoxide was effective as an opener. Reverse transcription-polymerase chain reaction (RT-PCR) studies detected sulfonylurea receptors SUR2B and SUR1 mRNA in TE671 cells whereas only SUR2B and SUR1 mRNA were, respectively, detected in A-10 and RIN-m5F cells. The inward rectifier Kir6.2 mRNA was detected in all three cell types whereas Kir6.1 was detected only in A-10 cells. Collectively, the molecular and pharmacologic studies suggest that K(ATP) channels endogenously expressed in TE671 medulloblastoma resemble those present in the smooth muscle.

Adenosine Triphosphate↗
PubMedDate not supplied

Synthesis and pharmacological evaluation of 2, 3-dihydro-3-oxo-4H-thieno[3,4-e][1,2,4]thiadiazine 1,1-dioxides as voltage-dependent calcium channel blockers.

The synthesis of a novel series of 2,3-dihydro-3-oxo-4H-thieno[3, 4-e][1,2,4]thiadiazine 1,1-dioxides and their pharmacological evaluation as drugs with effects on the rat cardiovascular system are described. The compounds under study were synthesized via Curtius rearrangement of appropriate sulfamoylacylazides which, in turn, were prepared from known starting materials. In isolated rat portal vein, these thienothiadiazines, like verapamil and diazoxide, inhibited the spontaneous motility produced by KCl (20 mM). In addition, the new compounds, like verapamil and unlike diazoxide, also exhibited inhibitory effects in the same preparation when the cell membrane was depolarized by an increased extracellular KCl concentration (80 mM) and, consequently, the membrane potential approached a level close to the K(+) equilibrium potential. Further characterization of this inhibitory activity led to the identification of a selective inhibitory effect of the new compounds on KCl (80 mM)-induced 45Ca(2+) uptake in the same vascular tissue. When tested in vivo (anaesthetized normotensive rats), acute administration of verapamil, diazoxide and some of the most in vitro potent compounds in 45Ca(2+) uptake experiments produced a gradual, dose-dependent and sustained decrease in diastolic arterial blood pressure, devoid of cardiac effects. These results suggest that, like verapamil, the cardiovascular effects produced by the new thienothiadiazines seem to be due, at least in part, to a blockade of transmembrane voltage-dependent calcium channels present in vascular smooth muscle cells and not to an activation of ATP-sensitive K(+) channels. Compounds 5b, 5e and 5i have been selected for further studies as antihypertensive agents.

Animals↗
PubMed2000

Roles of mitochondrial ATP-sensitive K channels and PKC in anti-infarct tolerance afforded by adenosine A1 receptor activation.

OBJECTIVES: This study intended to assess the role of mitochondrial ATP-sensitive potassium (mitoK ATP) channels and the sequence of signal transduction with protein kinase C (PKC) and adenosine A1 receptors in rabbits. BACKGROUND: To our knowledge, the link between trigger receptors of preconditioning, PKC and mitoK ATP channels has not been examined in a whole heart model of infarction. METHODS: In the first series of experiments, myocardial infarction was induced in isolated buffer-perfused rabbit hearts by 30-min global ischemia and 2-h reperfusion. Infarct size in the left ventricle was determined by tetrazolium staining and expressed as a percentage of area at risk (i.e., the whole left ventricle) (%IS/AR). In the second series of experiments, mitochondria were isolated from the heart, and their respiratory function was examined using glutamate as a substrate. RESULTS: Pretreatment with R-phenylisopropyladenosine (R-PIA, 1 micromol/liter), an A1-receptor agonist, reduced %IS/AR from 49.8 +/- 6.5% to 13.4 +/- 2.9%. This protection was abolished by calphostin C, a PKC inhibitor, and by 5-hydroxydecanoate (5-HD), a selective inhibitor of mitoK ATP channels. A selective mitoK ATP channel opener, diazoxide (100 micromol/liter), mimicked the effect of R-PIA on infarct size (%IS/AR = 11.6 +/- 4.0%), and this protective effect was also abolished by 5-HD. However, calphostin C failed to block the infarct size-limiting effect of diazoxide. Neither calphostin C nor 5-HD alone modified %IS/AR. State III respiration (QO2) and respiratory control index (RCI) were reduced after 30 min of ischemia (QO2 = 147.3 +/- 5.3 vs. 108.5 +/- 12.3, RCI = 22.3 +/- 1.1 vs. 12.1 +/- 1.8, p < 0.05). This mitochondrial dysfunction was persistent after 10 min of reperfusion (QO2 = 96.1 +/- 15.5, RCI = 9.5 +/- 1.9). Diazoxide significantly attenuated the respiratory dysfunction after 30 min of ischemia (QO2 = 142.8 +/- 9.7, RCI = 16.2 +/- 0.8) and subsequent 10-min reperfusion (QO2 = 135.3 +/- 7.2, RCI = 19.1 +/- 0.8). CONCLUSIONS: These results suggest that mitoK ATP channels are downstream of PKC in the mechanism of infarct-size limitation by A1-receptor activation and that the anti-infarct tolerance afforded by opening of mitoK ATP channels is associated with preservation of mitochondrial function during ischemia/reperfusion.

Adenosine Triphosphate↗
PubMed1997

KATP-channel on the somata of spiny neurones in rat caudate nucleus: regulation by drugs and nucleotides.

1. The aim of the present study was to characterize the pharmacological properties of the adenosine 5'-triphosphate(ATP)-sensitive K+ channel (KATP-channel) on the somata of spiny neurones in rat caudate nucleus and to compare them with those of beta-cells. For that purpose we tested the effects of several KATP-channel-inhibiting and -activating drugs on the opening activity of the KATP-channel in caudate nucleus by use of the patch-clamp technique. In addition, the modulation of drug responses by cytosolic nucleotides was examined. 2. When KATP-channels in caudate nucleus were activated in cell-attached patches by inhibition of mitochondrial energy production, meglitinide (a benzoic acid derivative), Hoe36320 (a sulphonylurea of low lipophilicity) and glipizide reduced KATP-channel activity half-maximally at 0.4 microM, 0.4 microM and about 0.5 nM, respectively. 3. In inside-out patches (presence of 0.7 mM free Mg2+ at the cytoplasmic membrane side), tolbutamide (0.1 mM) caused only partial inhibition of KATP-channels in the absence of cytosolic nucleotides but complete inhibition in the simultaneous presence of the channel-activating nucleotide guanosine 5'-diphosphate (GDP; 1 mM) and the channel-inhibiting nucleotide adenylyl-imidodiphosphate (AMP-PNP; 0.2 mM). 4. Diazoxide (0.3 mM) strongly increased channel activity in the presence of ATP (0.1 mM) or GDP (0.03 mM), but was ineffective in the presence of AMP-PNP (0.1 mM). In the absence of cytosolic nucleotides diazoxide even decreased channel activity. 5. In the presence of 0.1 mM ATP, diazoxide activated KATP-channels half-maximally at 38 microM. 6. When KATP-channel activity was inhibited by 0.1 mM ATP, (-)-pinacidil (0.5 mM) elicited a slight activation of KATP-channels in caudate nucleus, whereas (+)-pinacidil (0.5 mM) and lemakalim (0.3 mM) were ineffective. 7. Since our data indicate similar control by drugs and nucleotides of KATP-channels in the somata of spiny neurones and pancreatic beta-cells, we conclude that the high affinity sulphonylurea receptors of these tissues are probably closely related.

Adenosine Triphosphate↗
PubMed2003

Binding and effect of K ATP channel openers in the absence of Mg2+.

1 Openers of ATP-sensitive K(+) channels (K(ATP) channels) are thought to act by enhancing the ATPase activity of sulphonylurea receptors (SURs), the regulatory channel subunits. At higher concentrations, some openers activate K(ATP) channels also in the absence of MgATP. Here, we describe binding and effect of structurally diverse openers in the absence of Mg(2+) and presence of EDTA. 2 Binding of openers to SUR2B was measured using a mutant with high affinity for [(3)H]glibenclamide ([(3)H]GBC). In the absence of Mg(2+), 'typical' openers (benzopyrans, cyanoguanidines and aprikalim) inhibited [(3)H]GBC binding with K(i) values approximately 200 x higher than in the presence of MgATP. Minoxidil sulphate and nicorandil were inactive, whereas binding of diazoxide was unaffected by MgATP. 3 In the absence/presence of MgATP, N-cyano-N'-(1,1-dimethylpropyl)-N"-3-pyridylguanidine (P1075) activated the Kir6.2/SUR2B channel in inside-out patches with EC(50)=2000/67nM and E(max)=32/134%. In the absence of Mg(2+), responses were variable with only a small part of the variability being explained by a decrease in channel responsiveness with time after patch excision and to differences in the ATP sensitivity between patches. 4 The rank order of efficacy of the openers was P1075>rilmakalim approximately nicorandil>diazoxide>minoxidil sulphate. 5 The data show that structurally diverse openers are able to bind to, and to activate the Kir6.2/SUR2B channel by a pathway independent of ATP hydrolysis. These effects are observed at concentrations used to define the biochemical mechanism of the openers in the presence of MgATP and allow the openers to be classified into 'typical' and 'atypical' KCOs with diazoxide standing apart.

ATP-Binding Cassette Transporters↗
PubMed2003

Functional involvement of sulphonylurea receptor (SUR) type 1 and 2B in the activity of pig urethral ATP-sensitive K+ channels.

(1) We have investigated the possible roles of sulphonylurea receptor (SUR) type 1 and 2B in the activity of pig urethral ATP-sensitive K(+) channels (K(ATP) channels) by use of patch-clamp and reverse transcriptase-polymerase chain reaction (RT-PCR) techniques. (2) In voltage-clamp experiments, not only diazoxide, a SUR1 and weak SUR2B activator, but also pinacidil, a selective SUR2 activator, caused an inward current at a holding potential of -50 mV in symmetrical 140 mM K(+) conditions. (3) Gliclazide (</=1 micro M), a selective SUR1 blocker, inhibited the 10 micro M pinacidil-induced currents (K(i)=177 micro M) and the 500 micro M diazoxide-induced currents (high-affinity site, K(i1)=5 nM; low-affinity site, K(i2)=108 micro M) at -50 mV. (4) Application of tolbutamide (</=100 micro M) reversibly caused an inhibition of the 500 micro M diazoxide-induced current at -50 mV. (5) MCC-134, a SUR type-specific K(ATP) channel regulator (1-100 micro M), produced a concentration-dependent inward K(+) current, which was suppressed by the application of glibenclamide at -50 mV. The amplitude of the MCC-134 (100 micro M)-induced current was approximately 50% of that of the 100 micro M pinacidil-induced currents. (6) Using cell-attached configuration, MCC-134 activated a glibenclamide-sensitive K(ATP) channel which was also activated by pinacidil. (7) RT-PCR analysis demonstrated the presence of SUR1 and SUR2B transcripts in pig urethra. 8 These results indicate that both SUR1 and SUR2B subunits play a functional role in regulating the activity of pig urethral K(ATP) channels and that SUR1 contributes less than 25% to total K(ATP) currents.

ATP-Binding Cassette Transporters↗
PubMed1997

Activation and inhibition of K-ATP currents by guanine nucleotides is mediated by different channel subunits.

The ATP-sensitive potassium channel (K-ATP channel) plays a key role in insulin secretion from pancreatic beta-cells. It is closed by glucose metabolism, which stimulates secretion, and opened by the drug diazoxide, which inhibits insulin release. Metabolic regulation is mediated by changes in ATP and MgADP concentration, which inhibit and potentiate channel activity, respectively. The beta-cell K-ATP channel consists of a pore-forming subunit, Kir6.2, and a regulatory subunit, SUR1. The site at which ATP mediates channel inhibition lies on Kir6.2, while the potentiatory action of MgADP involves the nucleotide-binding domains of SUR1. K-ATP channels are also activated by MgGTP and MgGDP. Furthermore, both nucleotides support the stimulatory actions of diazoxide. It is not known, however, whether guanine nucleotides mediate their effects by direct interaction with one or more of the K-ATP channel subunits or indirectly via a GTP-binding protein. We used a truncated form of Kir6.2, which expresses independently of SUR1, to show that GTP blocks K-ATP currents by interaction with Kir6.2 and that the potentiatory effects of GTP are endowed by SUR1. We also showed that mutation of the lysine residue in the Walker A motif of either the first (K719A) or second (K1384M) nucleotide-binding domain of SUR1 abolished both the potentiatory effects of GTP and GDP on K-ATP currents and their ability to support stimulation by diazoxide. This argues that the stimulatory effects of guanine nucleotides require the presence of both Walker A lysines.

Adenosine Triphosphate↗
PubMed1992

Effect of ATP-sensitive K+ channel regulators on cystic fibrosis transmembrane conductance regulator chloride currents.

The cystic fibrosis transmembrane conductance regulator (CFTR) is a Cl- channel that is regulated by cAMP-dependent phosphorylation and by intracellular ATP. Intracellular ATP also regulates a class of K+ channels that have a distinct pharmacology: they are inhibited by sulfonylureas and activated by a novel class of drugs called K+ channel openers. In search of modulators of CFTR Cl- channels, we examined the effect of sulfonylureas and K+ channel openers on CFTR Cl- currents in cells expressing recombinant CFTR. The sulfonylureas, tolbutamide and glibenclamide, inhibited whole-cell CFTR Cl- currents at half-maximal concentrations of approximately 150 and 20 microM, respectively. Inhibition by both agents showed little voltage dependence and developed slowly; > 90% inhibition occurred 3 min after adding 1 mM tolbutamide or 100 microM glibenclamide. The effect of tolbutamide was reversible, while that of glibenclamide was not. In contrast to their activating effect on K+ channels, the K+ channel openers, diazoxide, BRL 38227, and minoxidil sulfate inhibited CFTR Cl- currents. Half-maximal inhibition was observed at approximately 250 microM diazoxide, 50 microM BRL 38227, and 40 microM minoxidil sulfate. The rank order of potency for inhibition of CFTR Cl- currents was: glibenclamide < BRL 38227 approximately equal to minoxidil sulfate > tolbutamide > diazoxide. Site-directed mutations of CFTR in the first membrane-spanning domain and second nucleotide-binding domain did not affect glibenclamide inhibition of CFTR Cl- currents. However, when part of the R domain was deleted, glibenclamide inhibition showed significant voltage dependence. These agents, especially glibenclamide, which was the most potent, may be of value in identifying CFTR Cl- channels. They or related analogues might also prove to be of value in treating diseases such as diarrhea, which may involve increased activity of the CFTR Cl- channel.

Adenosine Triphosphate↗
PubMed2003

ATP-sensitive potassium channels induced in liver cells after transfection with insulin cDNA and the GLUT 2 transporter regulate glucose-stimulated insulin secretion.

As part of our research into the liver-directed gene therapy of Type I diabetes, we have engineered a human hepatoma cell line (HEPG2ins/g cells) to store and secrete insulin to a glucose stimulus. The aim of the present study was to determine whether HEPG2ins/g cells respond to glucose via signaling pathways that depend on ATP-sensitive potassium channels (KATP). Using patch-clamp electrophysiology with symmetrical KCl solutions, the single-channel conductance of KATP was 61pS. KATP was inhibited by ATP (1 mM) or cAMP (50 microM) applied to the cytosolic side of the membrane. Single KATP channels and macroscopic whole-cell currents were inhibited by glucose (20 mM) and glibenclamide (20 microM) and were activated by diazoxide (150 microM). Immunoprecipitation and Western blot analysis confirmed the presence of Kir6.2 KATP channel subunit protein in HEPG2ins/g and HEPG2ins cells. Using radioimmunoassay techniques, we report that exposure of the cells to tolbutamide (100 microM) resulted in an increase in insulin secretion from 0.3 +/- 0.05 to 1.8 +/- 0.2 pmol insulin/10(6) cells and glibenclamide (20 microM) from 0.4 +/- 0.06 to 2.1 +/- 0.3 (n=4), similar to what is seen on glucose (20 mM) stimulation. Diazoxide (150 microM) completely inhibited glucose-stimulated insulin release. Glucose 20 mM and glibenclamide 100 microM increased intracellular Ca2+ level in the HEPG2ins/g cells. However, glucose 20 mM did not stimulate a rise in intracellular Ca2+ in the un-transfected parent cell-line HEPG2. We used confocal microscopy to confirm that glucose (20 mM) stimulated the release of insulin from the fluorescently labeled secretion granules in the cells. Furthermore, glibenclamide (20 microM) also stimulated the release of insulin from fluorescently labeled secretion granules, and diazoxide (150 microM) blocked that stimulated release of insulin. Our results suggest that HEPG2ins/g cells respond to glucose via signaling pathways that depend on KATP, similar to a normal pancreatic beta cell.

Adenosine Triphosphate↗
PubMed1997

Emptying of intracellular Ca2+ stores stimulates Ca2+ entry in mouse pancreatic beta-cells by both direct and indirect mechanisms.

1. In non-excitable cells, the depletion of intracellular Ca2+ stores triggers Ca2+ influx by a process called capacitative Ca2+ entry. In the present study, we have investigated how the emptying of these stores by thapsigargin (1 microM) influences Ca2+ influx in electrically excitable pancreatic beta-cells. The cytoplasmic Ca2+ concentration ([Ca2+]i) was monitored in clusters of mouse beta-cells or in whole islets loaded with fura-2. 2. The membrane was first held hyperpolarized by diazoxide, an opener of ATP-sensitive K+ (KATP) channels, in the presence of 4.8 mM K+. Alternating between Ca(2+)-free medium and medium containing 2.5 mM Ca2+ caused a minor rise in [Ca2+]i (approximately 14 nM) in clusters of beta-cells. A larger rise (approximately 65 nM), resistant to the blockade of voltage-dependent Ca2+ channels by D600, occurred when extracellular Ca2+ was readmitted after emptying intracellular Ca2+ stores with thapsigargin or acetylcholine. Thus there exists a small capacitative Ca2+ entry in beta-cells. 3. When the membrane potential was clamped at depolarized levels with 10, 20 or 45 mM K+ in the presence of diazoxide, [Ca2+]i increased to different plateau levels ranging between 100 and 900 nM. Thapsigargin consistently caused a further transient rise in [Ca2+]i, but had little (at 10 mM K+) or no effect on the plateau level. This confirms that the capacitative Ca2+ entry is small. 4. In clusters of cells whose membrane potential was not clamped with diazoxide, 15 mM glucose (in 4.8 mM K+) induced [Ca2+]i oscillations by promoting Ca2+ influx through voltage-dependent Ca2+ channels. The application of thapsigargin accelerated these oscillations and increased their amplitude, sometimes causing a sustained elevation of [Ca2+]i. Similar results were obtained from whole islets perifused with a medium containing > or = 6 mM glucose. The effect of thapsigargin was always much larger than expected from the capacitative Ca2+ entry, probably because of a potentiation of Ca2+ influx through voltage-dependent Ca2+ channels. 5. This potentiating effect of thapsigargin did not result from an acceleration of cell metabolism since the drug did not affect glucose-induced changes in NAD(P)H fluorescence. It is also unlikely to involve the inhibition of KATP channels because thapsigargin steadily elevated [Ca2+]i in cells in which [Ca2+]i oscillations persisted in the presence of a maximally effective concentration of tolbutamide. 6. In conclusion, the emptying of intracellular Ca2+ stores in beta-cells induces a small capacitative Ca2+ entry and activates a depolarizing current which potentiates glucose-induced Ca2+ influx through voltage-dependent Ca2+ channels.

Acetylcholine↗
PubMed1995

KATP channel mediation of anoxia-induced outward current in rat dorsal vagal neurons in vitro.

1. Thin brainstem slices (150 microns thickness) were taken from mature rats, and membrane potentials (Em) and currents (Im) in the dorsal vagal neurons (DVN) were analysed with whole-cell patch clamp techniques during anoxia. 2. At a holding potential (Vh) of -50 mV, a sustained anoxia-induced outward current (AOC) of 92 +/- 44 pA (reversal potential (Erev), -78 +/- 12 mV) and a concomitant increase of membrane conductance (gm) from 2.2 +/- 0.45 to 5.9 +/- 2.4 nS were revealed in 40% of 142 DVN analysed. The AOC led to a hyperpolarization of the cells by 14.4 +/- 6.1 mV from a mean resting Em of -51 +/- 6 mV, and to blockade of spontaneous action potential discharges. In the remaining DVN, anoxia had almost no effect on Em, Im or gm and did not block spontaneous action potential discharges. 3. The AOC was not affected by 0.5 microM tetrodotoxin (TTX), 2 mM Mn2+, 50 microM cyanonitroquinoxaline dione (CNQX) or 100 microM bicuculline. 4. Elevation of the extracellular [K+] from 3 to 10 mM resulted in a positive shift of Erev of the AOC by 23 mV, whereas an increase in the [Cl-] of the patch pipette solution from 5 to 144 mM had no effect on Erev. 5. In DVN responding with an AOC, addition of 200 microM diazoxide, an activator of ATP-sensitive K+ (KATP) channels, to oxygenated solutions elicited a similar outward current (Erev = -79 +/- 5.5 mV, n = 12) and increase in gm. Diazoxide did not affect Em, Im or gm in cells which did not show an AOC. 6. In a subpopulation of DVN (n = 26), spontaneous activation of a KATP current with an Erev of -80 +/- 6 mV was observed. As analysed in four of these cells, an AOC was revealed during the initial phase of development of the spontaneous outward current but not under steady-state conditions. 7. The AOC, the diazoxide-induced current, and the spontaneous outward current were completely blocked upon bath application of the KATP channel blocker tolbutamide (100-200 microM). 8. The results indicate that the sustained anoxia-induced outward current of dorsal vagal neurons is due to activation of KATP channels. A possible physiological role of functional inactivation of these cells during metabolic disturbances is discussed.

Adenosine Triphosphate↗
PubMed1996

ATP-sensitive K+ channels are functional in expiratory neurones of normoxic cats.

1. We analysed spontaneously active expiratory neurones (n = 48) of anaesthetized cats for the presence of ATP-sensitive K+ (KATP) channels. 2. Intracellular injection of ATP reversibly depolarized neurones during all phases of the respiratory cycle. During expiration, membrane potential depolarized by an average of 1.5 +/- 0.1 mV leading to a 25% increase of discharge frequency. During inspiration, ATP induced a 1.8 +/- 0.2 mV depolarization, which was accompanied by a maximum of 20% increase of input resistance (Rn). 3. Extracellular application of diazoxide, an agonist of KATP channels, resulted in reversible membrane hyperpolarization in 68% of neurones (n = 19). This hyperpolarization (2.5 mV during expiration and 3.1 mV during inspiration) was accompanied by a 22% decrease in Rn. 4. Extracellular application of tolbutamide and glibenclamide, two antagonists of KATP channels, evoked reversible depolarizations in 76% of neurones (n = 21). The depolarization was relatively constant throughout the respiratory cycle (1.4 mV during expiration and 2.3 mV during inspiration). Rn increased by 22%. 5. The same sulphonylureas also changed the steepness of membrane depolarization when neurones escaped spontaneous synaptic inhibition during postinspiration. Extracellularly applied tolbutamide and glibenclamide increased the steepness of depolarization by 21%, while diazoxide reduced it by 20%. 6. Antagonism of drugs was verified by simultaneous extra- and intracellular application of diazoxide and glibenclamide, respectively. 7. During voltage clamp at holding potential at -60 to -67 mV, intracellular or extracellular application of tolbutamide and glibenclamide blocked a persistent outward current. 8. We conclude that KATP channels are functional in expiratory neurones of adult cats and contribute to the control of excitability even during normoxia.

Adenosine Triphosphate↗
PubMed2002

Pre- and postsynaptic ATP-sensitive potassium channels during metabolic inhibition of rat hippocampal CA1 neurons.

Presynaptic and postsynaptic membrane activities during experimental metabolic inhibition were analysed in mechanically dissociated rat hippocampal neurons using nystatin-perforated and conventional whole-cell patch clamp recordings. NaCN, an inhibitor of mitochondrial ATP synthesis, induced an outward current across the postsynaptic soma membrane. This current was blocked by tolbutamide, a sulfonylurea, which blocks ATP-sensitive K+ (KATP) channels. The presynaptic effect of metabolic inhibitors such as NaCN, NaN3, or glucose-free solution was to increase the frequency of GABAergic miniature inhibitory postsynaptic currents (mIPSCs). Tolbutamide had no effect on this increase in mIPSC frequency induced by metabolic inhibition. Diazoxide, a KATP channel opener, evoked a similar somatic outward current in a dose-dependent manner. In addition, diazoxide decreased the frequency of mIPSCs in a dose-dependent fashion. Both these pre- and postsynaptic effects of diazoxide were reversed by tolbutamide, suggesting the existence of KATP channels on both pre- and postsynaptic membranes. These results confirm the presence of KATP channels on both the pre- and postsynaptic membranes but indicate that the channels have significantly different sensitivities to metabolic inhibition.

ATP-Binding Cassette Transporters↗
PubMed2000

Selective pharmacological agents implicate mitochondrial but not sarcolemmal K(ATP) channels in ischemic cardioprotection.

BACKGROUND: Pharmacological evidence has implicated ATP-sensitive K(+) (K(ATP)) channels as the effectors of cardioprotection, but the relative roles of mitochondrial (mitoK(ATP)) and sarcolemmal (surfaceK(ATP)) channels remain controversial. METHODS AND RESULTS: We examined the effects of the K(ATP) channel blocker HMR1098 and the K(ATP) channel opener P-1075 on surfaceK(ATP) and mitoK(ATP) channels in rabbit ventricular myocytes. HMR1098 (30 micromol/L) inhibited the surfaceK(ATP) current activated by metabolic inhibition, whereas the drug did not blunt diazoxide (100 micromol/L)-induced flavoprotein oxidation, an index of mitoK(ATP) channel activity. P-1075 (30 micromol/L) did not increase flavoprotein oxidation but did elicit a robust surfaceK(ATP) current that was completely inhibited by HMR1098. These results indicate that HMR1098 selectively inhibits surfaceK(ATP) channels, whereas P-1075 selectively activates surface K(ATP) channels. In a cellular model of simulated ischemia, the mitoK(ATP) channel opener diazoxide (100 micromol/L), but not P-1075, blunted cellular injury. The cardioprotection afforded by diazoxide or by preconditioning was prevented by the mitoK(ATP) channel blocker 5-hydroxydecanoate (500 micromol/L) but not by the surfaceK(ATP) channel blocker HMR1098 (30 micromol/L). CONCLUSIONS: The cellular effects of mitochondria- or surface-selective agents provide further support for the emerging consensus that mitoK(ATP) channels rather than surfaceK(ATP) channels are the likely effectors of cardioprotection.

Adenosine Triphosphate↗
PubMed1997

Octreotide in hyperinsulinism.

In a limited number of case reports in infants, octreotide raised the blood glucose concentrations and decreased glucose requirements sufficiently to avoid pancreatectomy. This response occurs in the presence of frequent feedings and diazoxide therapy, and lasts from 1 month to greater than 5 years. As expected, octreotide reduces growth indices such as growth factors and growth rate in short-term assessment. However, an insufficient sample size, a lack of follow-up, and poor study design provide inconclusive data. Among the few case reports in adults with benign or malignant insulinoma, octreotide can significantly raise blood glucose concentrations. In long-term follow-up, octreotide has alleviated or reduced the frequency of hypoglycemic episodes for periods of 5 months to 2.5 years. Octreotide was administered subcutaneously in regimens of 100-1500 micrograms in three to four divided doses or as a continuous infusion. Continuous subcutaneous infusion may be attempted in patients intolerant to intermittent administration. Octreotide may worsen existing hypoglycemia as result of suppressing glucagon and growth hormone in the presence of unresponsive pancreatic hyperinsulinism. While the long-term effects of growth remain undetermined, current findings suggest octreotide may provide a reasonable addition or alternative to diazoxide in controlling symptoms of congenital hyperinsulinism. Octreotide may be useful in management of hypoglycemic symptoms in adult patients requiring medical treatment for insulinoma who are refractory or intolerant of diazoxide. Additional long-term studies are needed to address the cost effectiveness of octreotide therapy, identify patients most likely to respond, and determine the impact of octreotide on height.

Adult↗
PubMed2002

BPDZ 154 activates adenosine 5'-triphosphate-sensitive potassium channels: in vitro studies using rodent insulin-secreting cells and islets isolated from patients with hyperinsulinism.

A novel ATP-sensitive potassium channel (K(ATP)) channel agonist, BPDZ 154 (6,7-dichloro-3-isopropylamino-4H-1,2,4-benzothiadiazine 1,1-dioxide), was synthesized, and its effects on insulin-secreting cells were evaluated using electrophysiology, (86)Rb(+) and (45)Ca(2+) efflux, and RIA determinations of insulin secretion. BPDZ 154, an analog of diazoxide, inhibited both glucose-induced insulin secretion from isolated perifused islets and the secretion of insulin induced by glucose and tolbutamide. These effects were mediated by the activation of ATP-sensitive potassium channels because BPDZ 154 induced a concentration-dependent increase in channel activity that was inhibited by the sulfonylurea tolbutamide and the imidazoline efaroxan. In beta-cells isolated from patients with either nontypical hyperinsulinism (preserved K(ATP) channel function) or from the control areas of the pancreas of patients with focal hyperinsulinism, BPDZ 154 activated K(ATP) channels and was found to be more effective and less readily reversible than diazoxide. By contrast, it was not possible to activate K(ATP) channels by either diazoxide or BPDZ 154 in beta-cells from patients with hyperinsulinism as a consequence of defects in K(ATP) channel function. In beta-cells isolated from a patient with pancreatic insulinoma, K(ATP) channels were readily recorded and modulated by BPDZ 154. These data suggest that BPDZ 154 or BPDZ 154-like compounds may have therapeutic potential in the treatment of certain forms of hyperinsulinism.

Adenosine Triphosphate↗
PubMed2006

Local anesthetic-induced protection against lipopolysaccharide-induced injury in endothelial cells: the role of mitochondrial adenosine triphosphate-sensitive potassium channels.

Lidocaine attenuates cell injury induced by ischemic-reperfusion and inflammation, although the protective mechanisms are not understood. We hypothesized that lidocaine and other amide local anesthetics protect against endothelial cell injury through activation of the mitochondrial adenosine triphosphate-sensitive potassium (mitoK(ATP)) channels. We determined the effects of amide local anesthetics (lidocaine, ropivacaine, and bupivacaine), ester local anesthetics (tetracaine and procaine), one amide analog (YWI), and two non-amide local anesthetic analogs (JDA and ICM) on viability of human microvascular endothelial cells after exposure to lipopolysaccharide (LPS) in the absence or presence of the mitoK(ATP) channel antagonist 5-hydroxydecaonate. Flavoprotein fluorescence was used to investigate the effects of local anesthetics on diazoxide-induced activation of mitoK(ATP) channels. Lidocaine, ropivacaine, bupivicaine, YWI, JDA, and ICM attenuated by 60% to 70% the decrease in cell viability caused by LPS. Amide local anesthetics and YWI protection was inhibited by 5-hydroxydecaonate, whereas the protection induced by JDA and ICM was not. Tetracaine and procaine did not protect against LPS-induced injury. The amide local anesthetics and the amide analog (YWI) enhanced diazoxide-induced flavoprotein fluorescence by 5% to 20%, whereas ester local anesthetics decreased diazoxide-induced flavoprotein fluorescence by 5% to 60% and the non-amide local anesthetic analogs had no effect. In conclusion, amide local anesthetics and the amide analog (YWI) attenuate LPS-induced cell injury, in part, through activation of mitoK(ATP) channels. In contrast, tetracaine and procaine had no protective effects and inhibited activation of mitoK(ATP) channels. The non-amide local anesthetic analogs induced protection but through mechanisms independent of mitoK(ATP) channels.

Adenosine Triphosphate↗