valinomycin and merocyanine-dye

The addition of oxygen to anaerobic rat liver mitochondria incubated at 15 degrees C in the absence of permeant cations produced negligible rapid H+ ejection, monitored spectroscopically with phenol red, which corresponded kinetically to the rise in delta psi, as monitored by merocyanine 540. Slow H+ translocation was observed under these conditions during the aerobic phase, the extent of which was proportional to the amount of oxygen added and the rate dependent on the rate of counter-ion movement. Measurement of H+ disappearance in the mitochondrial matrix, as monitored by neutral red, likewise showed little or no rapid H+ change in the absence of counter-ion movements. In the presence of permeant cations, the H+ disappearance in the matrix was readily measured. This observation argues against the importance of the mitochondrial outer membrane and intermembrane space in masking H+ movements. The H+ translocation required in the generation of maximal or static head delta psi was determined by following the spectral response of merocyanine to increasing oxygen additions. The amount of oxygen giving maximal Delta psi corresponds to an extrusion of 2-3 ng ions of H+ . mg of protein-1. The absence of H+ movement of near this magnitude during the development of the Delta psi argues against the Delta psi-driven backflow of H+ ions as the sole explanation of these observations.

Topics: Animals; Cytochromes; Depression, Chemical; Hydrogen; Membrane Potentials; Mitochondria, Liver; Oxygen; Phenolsulfonphthalein; Potassium; Protons; Pyrimidinones; Rats; Staining and Labeling; Valinomycin

The orientation and morphology of the endothelium lining the cardiovascular system may result from hemodynamic forces acting on the endothelial cells. To investigate the flow effects at the membrane level, we have examined the variations of the fluorescence intensity of two membrane-sensitive dyes, merocyanine 540 and bis(1,3-diethylthiobarbiturate)trimethineoxonol, (i) as a function of flow shear stress and (ii) with the onset or cessation of the flow. We found a time-dependent decrease in fluorescence intensity with the onset of the flow with an exponential approach to steady state of the order of 1 min. The process is reversible; when the flow is stopped the fluorescence intensity returns to its original value. The polarization of the endothelial cell membranes or, more precisely, the amplitude of the fluorescence intensity responses is an increasing function of the shear stress (up to 120 dynes/cm2). Assuming the equilibrium potential for K+ is more hyperpolarized than the resting potential and using valinomycin, we have deduced from the sign of the ionophore effects that the flow hyperpolarizes the endothelial cell membrane.

Topics: Animals; Cattle; Endothelium, Vascular; Fluorescence Polarization; Hemodynamics; Kinetics; Membrane Potentials; Potassium; Pulmonary Artery; Pyrimidinones; Thiobarbiturates; Time Factors; Valinomycin

1. Electrogenic steps in photosynthetic cyclic electron transport in chromatophore membrane of Chromatium vinosum were studied by measuring absorption changes of added merocyanin dye and of intrinsic carotenoid. 2. The change in dye absorbance was linear with the membrane potential change induced either by light excitation or by application of diffusion potential by adding valinomycin in the presence of K+ concentration gradient. 3. It was estimated that chromatophore membrane became 40--60 mV and 110--170 mV inside positive upon single and multiple excitations with single-turnover flashes, respectively, from the responses of the dye and the carotenoid. 4. Electron transfers between cytochrome c-555 or c-552 and reaction center bacteriochlorophyll dimer (BChl2) and between BChl2 and the primary electron acceptor were concluded to be electrogenic from the redox titration of the dye response. 5. No dye response which corresponded to the change of redox level of cytochrome b was observed in the titration curve. Addition of antimycin A slightly decreased the dye response. 6. The dye response was decreased under phosphorylating conditions. 7. From the results obtained localization of the electron transfer components in chromatophore membrane is discussed.

Topics: Antimycin A; Bacterial Chromatophores; Bacteriochlorophylls; Benzoxazoles; Carotenoids; Chromatium; Cytochrome b Group; Cytochromes; Electron Transport; Intracellular Membranes; Membrane Potentials; Oxidation-Reduction; Photosynthesis; Pyrimidinones; Spectrophotometry; Valinomycin

Topics: Benzoxazoles; Circular Dichroism; Erythrocytes; Humans; Male; Potassium; Pyrimidinones; Spectrometry, Fluorescence; Spectrophotometry; Valinomycin

Topics: Animals; Benzoxazoles; Biological Transport, Active; Calcium; Fluorescent Dyes; In Vitro Techniques; Membrane Potentials; Muscles; Pyrimidinones; Rabbits; Sarcoplasmic Reticulum; Valinomycin