valinomycin and tetramethylammonium

Spermine penetrates the mitochondrial matrix at significant rates which increase sharply and non-ohmically with membrane potential. In this respect, spermine uptake is qualitatively similar to that of other cations whose electrophoretic transport has been studied in mitochondria. At 200 mV and 1 mM spermine, the observed rate of spermine uptake was about 7 nmol x mg-1 x min-1, and the rate constant was about 8 times greater than that of tetraethylammonium cation. These rates are remarkably rapid considering that spermine is largely tetravalent at the pH of the experiment. The fluxes of spermine and tetraethylammonium are log-linear with membrane potential. The slope of the tetraethylammonium plot is consistent with leakage of this ion across a sharp Eyring barrier located in the middle of the membrane. The slope of the spermine plot is half that predicted by such a leak pathway, raising the possibility that spermine may cross the inner membrane by means of a channel. Whatever its mechanism of penetration, if comparable rates of uptake obtain in vivo and if spermine is not metabolized within the mitochondrial matrix, then a separate efflux mechanism would appear to be required to prevent unlimited spermine loading.

Topics: Animals; Biological Transport; Intracellular Membranes; Membrane Potentials; Mitochondria, Liver; Nigericin; Potassium; Quaternary Ammonium Compounds; Rats; Spermine; Valinomycin

The effect of nigericin on aggregation of bovine platelets was investigated in media containing the chloride salts of various alkali metal cations of quaternary ammonium cations. In medium with K+, which has the highest permeability with the ionophore among the cations tested, nigericin slightly enhanced both ADP- and thrombin-induced aggregation. In medium with Na+, nigericin scarcely affected ADP-induced aggregation, and slightly inhibited thrombin-induced aggregation. In media with Cs+, choline and tetramethylammonium, it inhibited the aggregations induced by both ADP and thrombin. Measurement of the cytoplasmic pH with the fluorescent probe 2',7'-bis(carboxyethyl)5,6-carboxyfluorescein showed that nigericin increased the intracellular pH in K+ medium and caused its stable decrease (of about 0.6) in Cs+, choline and tetramethylammonium media, but caused only a small transient decrease in medium with Na+. These results suggest that the effects of nigericin on platelet aggregation are mainly due to its effects on the cytoplasmic pH. This conclusion is supported by the findings that the effects on platelet aggregation of other types of ionophore tested were also proportional to their effects on the cytoplasmic pH.

Topics: Adenosine Diphosphate; Animals; Anti-Bacterial Agents; Blood Platelets; Cations; Cattle; Cesium; Choline; Cytoplasm; Gramicidin; Hydrogen-Ion Concentration; Nigericin; Platelet Aggregation; Potassium; Quaternary Ammonium Compounds; Sodium; Thrombin; Valinomycin

The Na+ concentration inside vascular smooth muscle cells is an important regulator of vascular smooth muscle function, but the mechanisms that mediate Na+ influx are not known. We studied Na+ transport in a newly described vesicle preparation preferentially enriched in sarcolemma and obtained by Mg2+ aggregation and differential centrifugation of homogenized dog superior mesenteric artery. In the presence of an outwardly directed proton gradient (pHout = 7.5, pHin = 5.0), 1 mM 22Na+ uptake was stimulated over twofold relative to the absence of a pH gradient (pHin = 7.5 or 5.0). pH gradient-stimulated Na+ uptake was inhibited by 1 mM amiloride. 22Na efflux was stimulated by an inwardly directed proton gradient (pHin = 7.5, pHout = 5.7 vs. 7.5). The rate of proton efflux from acid-loaded vesicles was measured by acridine orange fluorescence and was stimulated by 100 mM Naout but not by Nain = Naout = 100 mM. H+ gradient-stimulated Na+ transport and Na+ gradient-stimulated H+ transport were not due to electrical coupling between the two cations. The pH gradient-stimulated component of Na+ transport in the final vesicles, an intermediate fraction, and microsomes were proportional to the respective enzyme marker activities for sarcolemma but not for sarcoplasmic reticulum or mitochondrial membranes. We conclude that Mg2+ aggregation and differential centrifugation of homogenized vascular smooth muscle yield a vesicle preparation preferentially enriched in sarcolemma. Furthermore, the sarcolemma of vascular smooth muscle contains an amiloride-sensitive Na+-H+ proton countertransport system.

Topics: Animals; Biological Transport, Active; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Carrier Proteins; Cell Fractionation; Dogs; Electron Transport Complex IV; Female; Hydrogen-Ion Concentration; Male; Mesenteric Arteries; Muscle, Smooth, Vascular; NADPH-Ferrihemoprotein Reductase; Ouabain; Quaternary Ammonium Compounds; Sarcolemma; Sodium; Sodium-Hydrogen Exchangers; Valinomycin

The Cl- transport properties of the luminal border of bovine tracheal epithelium have been investigated using a highly purified preparation of apical plasma membrane vesicles. Transport of Cl- into an intravesicular space was demonstrated by (1) a linear inverse correlation between Cl- uptake and medium osmolarity and (2) complete release of accumulated Cl- by treatment with detergent. The rate of Cl- uptake was highly temperature-sensitive and was enhanced by exchange diffusion, providing evidence for a carrier-mediated transport mechanism. Transport of Cl- was not affected by the 'loop' diuretic bumetanide or by the stilbene-derivative anion-exchange inhibitors SITS (4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid) and DIDS (4,4'-diisothiocyanostilbene-2,2'-disulfonic acid). In the presence of the impermeant cation, tetramethylammonium (TMA+), uptake of Cl- was minimal; transport was stimulated equally by the substitution of either K+ or Na+ for TMA+. Valinomycin in the presence of K+ enhanced further Cl- uptake, while amiloride reduced Na+-stimulated Cl- uptake towards the minimal level observed with TMA+. These results suggest the following conclusions: (1) the tracheal vesicle membrane has a finite permeability to both Na+ and K+; (2) the membrane permeability to the medium counterion determines the rate of Cl- uptake; (3) Cl- transport is not specifically coupled with either Na+ or K+; and, finally (4) Cl- crosses the tracheal luminal membrane via an electrogenic transport mechanism.

Topics: Amiloride; Animals; Biological Transport; Cattle; Cell Membrane; Chlorides; Epithelium; Kinetics; Potassium; Quaternary Ammonium Compounds; Sodium; Trachea; Valinomycin