monensin and arsenic-acid

Vitreoscilla is a Gram-negative bacterium with unique respiratory physiology in which Na+ was implicated as a coupling cation for the generation of a transmembrane electrical gradient (delta psi). Thus, cells respiring in the presence of 110 mM Na+ generated a delta psi of -142 mV compared to only -42 and -56 mV for Li+ and choline, respectively, and even the -42 and -56 mV were insensitive to the protonophore 3,5-di-tert-butyl-4-hydroxybenzaldehyde (DTHB). The kinetics of delta psi formation and collapse correlated well with the kinetics of Na+ fluxes but not with those of H+ fluxes. Cyanide inhibited respiration, Na+ extrusion, and delta psi formation 81% or more, indicating that delta psi formation and Na+ extrusion were coupled to respiration. Experiments were performed to distinguish among three possible transport systems for this coupling: (1) a Na(+)-transporting ATPase; (2) an electrogenic Na+/H+ antiport system; (3) a primary Na+ pump directly driven by the free energy of electron transport. DCCD and arsenate decreased cellular ATP up to 86% but had no effect on delta psi, evidence against a Na(+)-transporting ATPase. Low concentrations of DTHB had no effect on delta psi; high concentrations transiently collapsed delta psi, but led to a stimulation of Na+ extrusion, the opposite of that expected for a Na+/H+ antiport system. Potassium ion, which collapses delta psi, also stimulated Na+ extrusion. The experimental evidence is against Na+ extrusion by mechanisms 1 and 2 and supports the existence of a respiratory-driven primary Na+ pump for generating delta psi in Vitreoscilla.

Topics: Arsenates; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Energy Metabolism; Hydrogen-Ion Concentration; Kinetics; Membrane Potentials; Monensin; Oxygen Consumption; Sodium; Thiotrichaceae

We have examined the ion transport properties and the inhibition of rat liver mitochondrial substrate oxidation by the antibiotic W341C. W341C was able to transport 22Na+ and 42K+ across a bulk carbon tetrachloride layer. A preference was shown for K+ transport. With equal molar antibiotic concentrations, W341C transported 42K+ at a greater rate than the K+-selective ionophore nigericin, but transported 22Na+ at a lesser rate than the Na+-selective ionophore monensin. Like nigericin, W341C was able to deplete mitochondrial K+, but not Mg2+ nor Ca2+. The inhibition of mitochondrial substrate oxidation by W341C paralleled the patterns obtained with nigericin. These data indicate that W341C is a K+-selective ionophore that inhibits mitochondrial substrate oxidation by a mechanism analogous to that of nigericin.

Topics: Animals; Anti-Bacterial Agents; Arsenates; Calcium; Carbon Tetrachloride; Cations; Ethers, Cyclic; Magnesium; Mitochondria, Liver; Monensin; Nigericin; Oxidation-Reduction; Oxygen Consumption; Potassium; Rats; Sodium

Respiration, membrane potential generation and motility of the marine alkalotolerant Vibrio alginolyticus were studied. Subbacterial vesicles competent in NADH oxidation and delta psi generation were obtained. The rate of NADH oxidation by the vesicles was stimulated by Na+ in a fashion specifically sensitive to submicromolar HQNO (2-heptyl-4-hydroxyquinoline N-oxide) concentrations. The same amounts of HQNO completely suppressed the delta psi generation. Delta psi was also inhibited by cyanide, gramicidin D and by CCCP + monensin. CCCP (carbonyl cyanide m-chlorophenylhydrazone) added without monensin exerted a much weaker effect on delta psi. Na+ was required to couple NADH oxidation with delta psi generation. These findings are in agreement with the data of Tokuda and Unemoto on Na+-motive NADH oxidase in V. alginolyticus. Motility of V. alginolyticus cells was shown to be (i) Na+-dependent, (ii) sensitive to CCCP + monensin combination, whereas CCCP and monensin, added separately, failed to paralyze the cells, (iii) sensitive to combined treatment by HQNO, cyanide or anaerobiosis and arsenate, whereas inhibition of respiration without arsenate resulted only in a partial suppression of motility. Artificially imposed delta pNa, i.e., addition of NaCl to the K+ -loaded cells paralyzed by HQNO + arsenate, was shown to initiate motility which persisted for several minutes. Monensin completely abolished the NaCl effect. Under the same conditions, respiration-supported motility was only slightly lowered by monensin. The artificially-imposed delta pH, i.e., acidification of the medium from pH 8.6 to 6.5 failed to activate motility. It is concluded that delta mu Na+ produced by (i) the respiratory chain and (ii) an arsenate-sensitive anaerobic mechanism (presumably by glycolysis + Na+ ATPase) can be consumed by an Na+ -motor responsible for motility of V. alginolyticus.

Topics: Arsenates; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Cyanides; Electrophysiology; Gramicidin; Hydrogen-Ion Concentration; Hydroxyquinolines; Monensin; Movement; NAD; Oxidation-Reduction; Potassium; Sodium; Vibrio

Monensin, a monovalent ionophore, caused swelling of mature cisternae of plant Golgi apparatus. The appearance of swollen cisternae was time-dependent and linear over a period of 1 h with an estimated maximum rate of production of one swollen cisterna every 3 to 4 min. Implicit in these observations was a need for the uptake of osmotically active monovalent cations to have occurred accompanied by a concomitant efflux of H+ and the entry of water. Furthermore, to sustain the H+ efflux, a source of H+ influx also would be required. To test for the latter, cisternal swelling, as visualized by electron microscopy, was monitored by treatment of wild carrot cells in suspension culture with drugs and inhibitors known to interfere with proton gradients. Swelling was inhibited by the protonophore, FCCP, by the inhibitor of lysosomal acidification, quercetin, and by the lysosomotropic amines, chloroquine and ammonia. While antimycin A, an inhibitor of mitochondrial oxidative phosphorylation, was ineffective, cyanide dramatically decreased swelling. The numbers of swollen cisternae produced could be reduced by prolonged treatment with arsenate, such that an ATP requirement is indicated, at least, for cisternal formation. Swelling was promoted by citrate, representative of a permeant organic anion. Reductions in numbers of monensin-induced swollen cisternae in the presence of quercetin, vanadate, and chloroquine could be compensated for by the addition of citrate. We conclude that the monensin-induced swelling of Golgi apparatus cisternae may involve a mechanism generating a proton gradient at or near the mature Golgi apparatus face.

Topics: Arsenates; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cells, Cultured; Furans; Golgi Apparatus; Hydrogen-Ion Concentration; Microscopy, Electron; Models, Biological; Monensin; Plants