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Biochimica Et Biophysica Acta Nov 1997The effects of two ionizable cryptands, the Na-selective (221)C10 and the K-selective (222)C10, and of valinomycin, FCCP and nystatin on K+ fluxes in opossum kidney (OK)...
The effects of two ionizable cryptands, the Na-selective (221)C10 and the K-selective (222)C10, and of valinomycin, FCCP and nystatin on K+ fluxes in opossum kidney (OK) cells have been quantified. The Na,K-ATPase (ouabain-sensitive 86Rb influx) was stimulated by nystatin (> or = 20%), and inhibited by the other ionophores (50-80%), by barium (K-channel blocker) (61%) and by amiloride (Na entry blocker) (34%). The Vmax of the Na,K-ATPase phosphatase activity was unmodified by the ionophores, indicating the absence of direct interaction with the enzyme. The ATPi content was unmodified by the inhibitors and nystatin, but was lowered by (221)C10 (47%), (222)C10 (75%), valinomycin (72%) and FCCP (88%). Amiloride was found to partially remove the inhibition caused by (222)C10 (51%) and valinomycin (49%). Rb efflux was stimulated by nystatin (32%), unmodified by valinomycin, and was inhibited by (221)C10 (19%), (222)C10 (19%) and FCCP (10%). Barium (39%) and amiloride (32%) inhibited this efflux and, in their presence, the nystatin effect persisted, whereas that of the other ionophores vanished. At pH 6.4, the Rb efflux decreased by 14% of its value at pH 7.4, with no additional inhibition by cryptands. Cryptands are shown to inhibit the pH-sensitive K+-conductance, probably by inducing a K+-H+ exchange at the plasma membrane, and by uncoupling oxidative phosphorylation by inducing the entry of K+ and H+ (and possibly Ca2+) ions into the mitochondria.
Topics: Adenosine Triphosphate; Animals; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cell Line; Ion Transport; Kidney; Nystatin; Opossums; Potassium; Rubidium; Sodium; Sodium-Potassium-Exchanging ATPase; Valinomycin
PubMed: 9375811
DOI: 10.1016/s0005-2736(97)00140-5 -
The Journal of Physiology Feb 19691. The average rate constant for loss of (45)Ca from an unpoisoned squid axon was 1.8 x 10(-3) min(-1), corresponding to an efflux of 0.2 p-mole/cm(2) sec.2. The Ca...
1. The average rate constant for loss of (45)Ca from an unpoisoned squid axon was 1.8 x 10(-3) min(-1), corresponding to an efflux of 0.2 p-mole/cm(2) sec.2. The Ca efflux from unpoisoned axons was reduced if external calcium was replaced with magnesium, or external sodium with lithium, choline or dextrose. Replacing both sodium and calcium reduced the efflux to about 40%.3. Cyanide caused little immediate change in Ca efflux but after 1(1/2)-2(1/2) hr the efflux increased to 5-15 times its normal value. The effect was rapidly reversed when cyanide was removed.4. The large Ca efflux into cyanide was reduced by a factor of three when external calcium was replaced with magnesium and by a further factor of about six when external sodium was replaced with lithium.5. The Ca efflux from both poisoned and unpoisoned axons had a Q(10) of 2-3, was not affected by ouabain and was greatly reduced by injecting ethyleneglycol bis (aminoethylether)-N,N'-tetra-acetic acid (EGTA).6. After injecting (45)Ca along the axis, the efflux of calcium reached its maximum much more rapidly in a cyanide-treated axon than in an unpoisoned axon.7. Pre-treatment with cyanide greatly increased the rate at which calcium was lost from axoplasm extruded into flattened dialysis bags. A similar effect was observed when cyanide was applied after extrusion.8. Replacing external sodium glutamate with potassium glutamate greatly reduced the loss of (45)Ca from intact axons poisoned with cyanide but had little effect on the loss from extruded axoplasm.9. The rate constant for loss of the Ca EGTA complex was about 3 x 10(-5) min(-1) for intact axons and 2 x 10(-2) min(-1) for extruded axoplasm.10. A possible explanation of the cyanide effect is that, after poisoning, calcium ions are released from a store and can then exchange at a higher rate with external sodium or calcium.11. The experiments suggest that part of the calcium efflux may be coupled to sodium entry.12. Theoretical equations for ;diffusion and chemical reaction in a cylinder' are described in the Appendix.
Topics: Animals; Axons; Biological Transport, Active; Calcium; Calcium Isotopes; Cyanides; Cytoplasm; Dialysis; Ethers; Imides; In Vitro Techniques; Kinetics; Mollusca; Sodium; Time Factors
PubMed: 5764408
DOI: 10.1113/jphysiol.1969.sp008704 -
Journal of Bacteriology Mar 1977Cell of Mycoplasma pneumoniae FH gliding on a glass surface in liquid medium were examined by microscopic observation and quantitatively by microcinematography (30...
Cell of Mycoplasma pneumoniae FH gliding on a glass surface in liquid medium were examined by microscopic observation and quantitatively by microcinematography (30 frames per min). Comparisons were made only within the individual experiments. The cells moved in an irregular pattern with numerous narrow bends and circles. They never changed their leading end. The average speed (without pauses) was relatively constant between o.2 and 0.5 mum/s. The maximum speed was about 1.5 to 2.0 mum/s. The movements were interrupted by resting periods of different lengths and frequency. Temperature, viscosity, pH, and the presence of yeast extract in the medium influenced the motility significantly; changes in glucose, calcium ions, and serum content were less effective. The movements were affected by iodoacetate, p-mercuribenzoate, and mitomycin C at inhibitory or subinhibitory concentrations. Sodium fluoride, sodium cyanide, dinitrophenol, chloramphenicol, puromycin, cholchicin, and cytochalasin B at minimal inhibitory concentrations did not affect motility. The movements were effectively inhibited by anti-M. pneumoniae antiserum. Studies with absorbed antiserum suggested that the surface components involved in motility are heat labile. The gliding of M. pneumoniae cells required an intact energy metabolism and the proteins involved seemed to have a low turnover.
Topics: Anti-Bacterial Agents; Antibodies, Bacterial; Chloromercuribenzoates; Concanavalin A; Culture Media; Cyanides; Cytochalasin B; Dinitrophenols; Fluorides; Glucose; Hydrogen-Ion Concentration; Motion Pictures; Movement; Mycoplasma; Temperature; Ultraviolet Rays; Viscosity
PubMed: 14925
DOI: 10.1128/jb.129.3.1495-1501.1977 -
EBioMedicine Sep 2015Global or local ischemia contributes to the pathogenesis of acute kidney injury (AKI). Currently there are no specific therapies to prevent AKI. Potentiation of...
Global or local ischemia contributes to the pathogenesis of acute kidney injury (AKI). Currently there are no specific therapies to prevent AKI. Potentiation of glycolytic metabolism and attenuation of mitochondrial respiration may decrease cell injury and reduce reactive oxygen species generation from the mitochondria. Meclizine, an over-the-counter anti-nausea and -dizziness drug, was identified in a 'nutrient-sensitized' chemical screen. Pretreatment with 100 mg/kg of meclizine, 17 h prior to ischemia protected mice from IRI. Serum creatinine levels at 24 h after IRI were 0.13 ± 0.06 mg/dl (sham, n = 3), 1.59 ± 0.10 mg/dl (vehicle, n = 8) and 0.89 ± 0.11 mg/dl (meclizine, n = 8). Kidney injury was significantly decreased in meclizine treated mice compared with vehicle group (p < 0.001). Protection was also seen when meclizine was administered 24 h prior to ischemia. Meclizine reduced inflammation, mitochondrial oxygen consumption, oxidative stress, mitochondrial fragmentation, and tubular injury. Meclizine preconditioned kidney tubular epithelial cells, exposed to blockade of glycolytic and oxidative metabolism with 2-deoxyglucose and NaCN, had reduced LDH and cytochrome c release. Meclizine upregulated glycolysis in glucose-containing media and reduced cellular ATP levels in galactose-containing media. Meclizine inhibited the Kennedy pathway and caused rapid accumulation of phosphoethanolamine. Phosphoethanolamine recapitulated meclizine-induced protection both in vitro and in vivo.
Topics: Acute Kidney Injury; Adenosine Triphosphate; Animals; Cell Respiration; Cytochromes c; Deoxyglucose; Disease Models, Animal; Epithelial Cells; Ethanolamines; Galactose; Glycolysis; Humans; Inflammation; Ischemic Preconditioning; Kidney; Kidney Tubules; L-Lactate Dehydrogenase; LLC-PK1 Cells; Male; Meclizine; Mice, Inbred C57BL; Mitochondria; Protective Agents; Reperfusion Injury; Sodium Cyanide; Swine; Up-Regulation
PubMed: 26501107
DOI: 10.1016/j.ebiom.2015.07.035 -
British Journal of Anaesthesia Jul 1982
Topics: Animals; Dogs; Ferricyanides; Humans; Hydrogen Cyanide; Nitroprusside
PubMed: 6285944
DOI: 10.1093/bja/54.7.791-b -
Frontiers in Physiology 2011Metabolic inhibition causes a decline in mechanical performance and, if prolonged, myocardial contracture and cell death. The decline in mechanical performance is mainly...
OBJECTIVE
Metabolic inhibition causes a decline in mechanical performance and, if prolonged, myocardial contracture and cell death. The decline in mechanical performance is mainly due to altered intracellular calcium handling, which is under control of the Na(+)/Ca(2+)-exchanger (NCX) The driving force of the NCX (ΔG(ncx)) determines the activity of NCX. The aim of this study was to describe the relation between ΔG(ncx) and calcium homeostasis during metabolic inhibition.
METHODS
In left ventricular rabbit myocytes, during metabolic inhibition (2 mmol/L sodium cyanide), sodium ([Na(+)](i)), calcium ([Ca(2+);](i)), and action potentials were determined with SBFI, indo-1, and the patch clamp technique. Changes of ΔG(ncx) were calculated.
RESULTS
During metabolic inhibition: The first 8 min [Na(+)](i) remained constant, systolic calcium decreased from 532 ± 28 to 82 ± 13 nM, diastolic calcium decreased from 121 ± 12 to 36 ± 10 nM and the sarcoplasmic reticulum (SR) calcium content was depleted for 85 ± 3%. After 8 min [Na(+);](i) and diastolic calcium started to increase to 30 ± 1.3 mmol/L and 500 ± 31 nM after 30 min respectively. The action potential duration shortened biphasically. In the first 5 min it shortened from 225 ± 12 to 153 ± 11 ms and remained almost constant until it shortened again after 10 min. After 14 min action potential and calcium transients disappeared due to unexcitability of the myocytes. This resulted in an increased of the time average of ΔG(ncx) from 6.2 ± 0.2 to 7.7 ± 0.3 kJ/mol during the first 3 min, where after it decreased and became negative after about 15 min.
CONCLUSION
Metabolic inhibition caused an early increase of ΔG(ncx) caused by shortening of the action potential. The increase of ΔG(ncx) contributed to decrease of diastolic calcium, calcium transient amplitude, SR calcium content, and contractility. The increase of diastolic calcium started after ΔG(ncx) became lower than under aerobic conditions.
PubMed: 21483726
DOI: 10.3389/fphys.2011.00010 -
Medicina (Kaunas, Lithuania) Apr 2022: The commissural nucleus of the tractus solitarius (cNTS) not only responds to glucose levels directly, but also receives afferent signals from the liver, and from the...
Leptin in the Commissural Nucleus of the Tractus Solitarius (cNTS) and Anoxic Stimulus in the Carotid Body Chemoreceptors Increases cNTS Leptin Signaling Receptor and Brain Glucose Retention in Rats.
: The commissural nucleus of the tractus solitarius (cNTS) not only responds to glucose levels directly, but also receives afferent signals from the liver, and from the carotid chemoreceptors (CChR). In addition, leptin, through its receptors in the cNTS, regulates food intake, body weight, blood glucose levels, and brain glucose retention (BGR). These leptin effects on cNTS are thought to be mediated through the sympathetic-adrenal system. How these different sources of information converging in the NTS regulate blood glucose levels and brain glucose retention remains largely unknown. The goal of the present study was to determine whether the local administration of leptin in cNTS alone, or after local anoxic stimulation using sodium cyanide (NaCN) in the carotid sinus, modifies the expression of leptin and of mRNA. We also investigated how leptin, alone, or in combination with carotid sinus stimulation, affected brain glucose retention. : The experiments were carried out in anesthetized male Wistar rats artificially ventilated to maintain homeostatic values for pO, pCO, and pH. We had four groups: (a) experimental 1, leptin infusion in cNTS and NaCN in the isolated carotid sinus (ICS; = 10); (b) experimental 2, leptin infusion in cNTS and saline in the ICS ( = 10); (c) control 1, artificial cerebrospinal fluid (aCSF) in cNTS and NaCN in the ICS ( = 10); (d) control 2, aCSF in cNTS and saline in the ICS ( = 10). : Leptin in cNTS, preceded by NaCN in the ICS increased BGR and leptin mRNA receptor expression, with no significant increases in mRNA in the NTSc. : Leptin in the cNTS enhances brain glucose retention induced by an anoxic stimulus in the carotid chemoreceptors, through an increase in receptors, without persistent changes in neuronal activation.
Topics: Animals; Blood Glucose; Carotid Body; Glucose; Hypoxia; Leptin; Male; RNA, Messenger; Rats; Rats, Wistar; Receptors, Leptin; Solitary Nucleus
PubMed: 35454388
DOI: 10.3390/medicina58040550 -
Journal of Molecular and Cellular... Sep 2013ATP-sensitive potassium channel (KATP) activation can drastically shorten action potential duration (APD) in metabolically compromised myocytes. We showed previously...
ATP-sensitive potassium channel (KATP) activation can drastically shorten action potential duration (APD) in metabolically compromised myocytes. We showed previously that SUR1 with Kir6.2 forms the functional channel in mouse atria while Kir6.2 and SUR2A predominate in ventricles. SUR1 is more sensitive to metabolic stress than SUR2A, raising the possibility that KATP in atria and ventricles may respond differently to metabolic stress. Action potential duration (APD) and calcium transient duration (CaTD) were measured simultaneously in both atria and ventricles by optical mapping of the posterior surface of Langendorff-perfused hearts from C57BL wild-type (WT; n=11), Kir6.2(-/-) (n=5), and SUR1(-/-) (n=6) mice during metabolic inhibition (MI, 0mM glucose+2mM sodium cyanide). After variable delay, MI led to significant shortening of APD in WT hearts. On average, atrial APD shortened by 60.5 ± 2.7% at 13.1 ± 2.1 min (n=6, p<0.01) after onset of MI. Ventricular APD shortening (56.4 ± 10.0% shortening at 18.2 ± 1.8 min) followed atrial APD shortening. In SUR1(-/-) hearts (n=6), atrial APD shortening was abolished, but ventricular shortening (65.0 ± 15.4% at 25.33 ± 4.48 min, p<0.01) was unaffected. In Kir6.2(-/-) hearts, two disparate responses to MI were observed; 3 of 5 hearts displayed slight shortening of APD in the ventricles (24 ± 3%, p<0.05) and atria (39.0 ± 1.9%, p<0.05) but this shortening occurred later and to much less extent than in WT (p<0.05). Marked prolongation of ventricular APD was observed in the remaining hearts (327% and 489% prolongation) and was associated with occurrence of ventricular tachyarrhythmias. The results confirm that Kir6.2 contributes to APD shortening in both atria and ventricle during metabolic stress, and that SUR1 is required for atrial APD shortening while SUR2A is required for ventricular APD shortening. Importantly, the results show that the presence of SUR1-dependent KATP in the atria results in the action potential being more susceptible to metabolically driven shortening than the ventricle.
Topics: Action Potentials; Animals; Heart Atria; Heart Ventricles; In Vitro Techniques; KATP Channels; Mice; Mice, Inbred C57BL; Mice, Knockout; Sulfonylurea Receptors
PubMed: 23624089
DOI: 10.1016/j.yjmcc.2013.04.016 -
British Journal of Pharmacology 19801 The effects of intracarotid injections of methionine-enkephalin (Met-enkephalin) and morphine on chemoreceptor activity recorded from the peripheral end of a sectioned...
1 The effects of intracarotid injections of methionine-enkephalin (Met-enkephalin) and morphine on chemoreceptor activity recorded from the peripheral end of a sectioned carotid sinus nerve have been studied in cats anaesthetized with pentobarbitone. 2 Met-enkephalin caused a rapid, powerful, inhibition of spontaneous chemoreceptor discharge, the intensity and duration of which was dose-dependent. 3 Morphine was a less potent inhibitor of spontaneous chemoreceptor discharge, and the inhibition it evoked was rather variable and tended to be biphasic. Low doses of morphine caused a slight increase in discharge. 4 Naloxone (0.2 mg i.c.) slightly increased spontaneous discharge, greatly reduced the chemo-inhibition caused by morphine, and reduced the inhibitory effect of Met-enkephalin. A higher dose of naloxone (0.8 mg) caused a substantial reduction of the Met-enkephalin effect. 5 Chemo-excitation evoked by intracarotid injections of acetylcholine, CO2-saturated Locke solution, and sodium cyanide were only slightly and somewhat variably reduced following injections of Met-enkephalin, whereas the inhibitory effect of dopamine was potentiated. Following morphine administration, response to acetylcholine and sodium cyanide were reduced slightly, whereas those to CO2 and dopamine were potentiated. 6 Responses to acetylcholine and CO2 were slightly potentiated during infusion of Met-enkephalin (50 micrograms/min, i.c.) and the response to sodium cyanide was slightly reduced. 7 It is concluded that naloxone-sensitive opiate receptors are present in the cat carotid body; when activated they cause inhibition of spontaneous chemoreceptor discharge. The physiological role of these receptors and the identity of any endogenous ligand remains to be established.
Topics: Acetylcholine; Animals; Carbon Dioxide; Carotid Sinus; Cats; Chemoreceptor Cells; Cyanides; Dopamine; Endorphins; Enkephalin, Methionine; Enkephalins; In Vitro Techniques; Morphine; Naloxone
PubMed: 6781573
DOI: 10.1111/j.1476-5381.1980.tb10939.x -
Infection and Immunity Apr 1975Ammonium chloride (4 times 10-3 M) rendered HEp-2 monolayers completely insensitive to the action of diphtheria toxin, as measured by de novo protein synthesis. Total...
Ammonium chloride (4 times 10-3 M) rendered HEp-2 monolayers completely insensitive to the action of diphtheria toxin, as measured by de novo protein synthesis. Total protection was observed even with large amounts of toxin (400 minimum lethal doses/ml). Ammonium chloride did not reduce toxicity by direct action on the protein, nor did it prevent the adsorption of toxin to the cell membrane. Although the ammonium salt did not block the initial interaction between cell and toxin, it did maintain the toxin at a site amenable to neutralization with antitoxin. Surface-adsorbed toxin was inactivated by cellular enzymes or alternatively was desorbed from the membrane during a 12-h incubation in the presence of ammonium chloride. In addition, ammonium chloride provided protection to both toxin-sensitive guinea pig peritoneal macrophages and a partially toxin-resistant strain of HEp-2 cells. Sodium arsenite was effective in protecting cell monolayers from the action of diphtheria toxin; unlike ammonium chloride, its action was not dependent upon continued incubation with cells during exposure to toxin. Inhibitors of energy metabolism abolished toxin action either totally (sodium fluoride) or partially (dinitrophenol and sodium cyanide). Inhibitors of cellular proteases, on the other hand, did not modify toxin activity. The ability of several modifiers of membrane function to alter expression of toxicity for HEp-2 cells was also examined. One compound known to enhance endocytic activity, Tuftsin, had no effect, whereas poly-L-ornithine provided partial protection. Of the two compounds known to alter membrane fluidity, cytochalasin B provided partial protection for HEp-2 cell cultures, whereas colchicine had no effect. Agents that bind to sulfhydryl groups on the cell surface had no apparent effect on toxicity, suggesting that the initial toxin-cell interaction does not involve sulfhydryl groups. Those compounds that provide virtually full protection against the action of diphtheria toxic on cell monolayers (i.e., ammonium chloride, sodium fluoride, and sodium arsenite) had no inhibitory effect on the in vitro enzyme activity associated with fragment A of the toxin.
Topics: Ammonium Chloride; Antimetabolites; Arsenicals; Cell Line; Cell Membrane; Culture Techniques; Cytochalasin B; Diphtheria Antitoxin; Diphtheria Toxin; Dithiothreitol; Drug Resistance; Enzyme Inhibitors; Epithelial Cells; Glutathione; Humans; Laryngeal Neoplasms; Leucine; Macrophages; NADP; Neutralization Tests; Peptide Hydrolases; Pyridines; Tritium
PubMed: 235491
DOI: 10.1128/iai.11.4.665-674.1975