Detection of charge movements in ion pumps by a family of styryl dyes.
AuthorsPedersen M, Roudna M, Beutner S, Birmes M, Reifers B, Martin HD, Apell HJ
JournalJ Membr Biol
PubMed ID11891580
'A family of fluorescent styryl dyes was synthesized to apply them as probes that monitor the ion-translocating activity of the Na,K-ATPase and the SR Ca-ATPase, similar to the widely used dye RH421. All dyes had the same chromophore but they differed in the length of the spacer between chromophore and ... More
Voltage-sensitive dye RH421 increases contractility of cardiac muscle.
AuthorsCheng Y, Van Wagoner DR, Mazgalev TN, Tchou PJ, Efimov IR
JournalCan J Physiol Pharmacol
PubMed ID10326837
'Voltage-sensitive dyes and imaging techniques have proved to be indispensable tools for use in in vitro electrophysiological studies. To avoid motion artifacts in optical recordings, electromechanical uncouplers such as 2,3-butanedione monoxime (BDM) are required. In this study, we sought to determine whether the voltage-sensitive dye RH421 had an effect on ... More
Near infrared two-photon excitation cross-sections of voltage-sensitive dyes.
AuthorsFisher JA, Salzberg BM, Yodh AG
JournalJ Neurosci Methods
PubMed ID16129493
'Microscopy based on voltage-sensitive dyes has proven effective for revealing spatio-temporal patterns of neuronal activity in vivo and in vitro. Two-photon microscopy using voltage-sensitive dyes offers the possibility of wide-field visualization of membrane potential on sub-cellular length scales, hundreds of microns below the tissue surface. Very little information is available, ... More
Optical detection of membrane dipole potential: avoidance of fluidity and dye-induced effects.
AuthorsClarke RJ, Kane DJ
JournalBiochim Biophys Acta
PubMed ID9042345
'Fluorescent styrylpyridinium dyes have recently been suggested as probes of the membrane dipole potential and of the kinetics of electrogenic ion pumps. It is necessary, however, to be able to confidently attribute observed fluorescence changes to electrical effects alone and avoid interference from changes in membrane fluidity. Furthermore, the effect ... More
Entrance port for Na(+) and K(+) ions on Na(+),K(+)-ATPase in the cytoplasmic loop between trans-membrane segments M6 and M7 of the alpha subunit. Proximity Of the cytoplasmic segment of the beta subunit.
AuthorsShainskaya A, Schneeberger A, Apell HJ, Karlish SJ
JournalJ Biol Chem
PubMed ID10636905
'Based on the following observations we propose that the cytoplasmic loop between trans-membrane segments M6 and M7 (L6/7) of the alpha subunit of Na(+),K(+)-ATPase acts as an entrance port for Na(+) and K(+) ions. 1) In defined conditions chymotrypsin specifically cleaves L6/7 in the M5/M6 fragment of 19-kDa membranes, produced ... More
Kinetics of the phosphorylation of Na,K-ATPase by inorganic phosphate detected by a fluorescence method.
AuthorsApell HJ, Roudna M, Corrie JE, Trentham DR
JournalBiochemistry
PubMed ID8718885
'Phosphorylation by Pi of the Na,K-ATPase from rabbit kidney in the absence of Na+ ions but in the presence of Mg2+ ions has been studied. In the absence of K+ ions, unphosphorylated and phosphorylated states induce different fluorescence levels in the membrane-bound styryl dye RH421, and hence transitions between the ... More
Voltage-sensitive dye recording of action potentials and synaptic potentials from sympathetic microcultures.
AuthorsChien CB, Pine J
JournalBiophys J
PubMed ID1681956
'Given the appropriate multicell electrophysiological techniques, small networks of cultured neurons (microcultures) are well suited to long-term studies of synaptic plasticity. To this end, we have developed an apparatus for optical recording from cultured vertebrate neurons using voltage-sensitive fluorescent dyes (Chien, C.-B., and J. Pine. 1991. J. Neurosci. Methods. 38:93-105). ... More
Fluorescent styryl dyes applied as fast optical probes of cardiac action potential.
AuthorsMüller W, Windisch H, Tritthart HA
JournalEur Biophys J
PubMed ID3816701
'Several styryl dyes were tested as fast optical probes of membrane action potentials in mammalian heart muscle tissue. After staining, atrial specimens were superfused in physiological salt solution, and fluorescence was excited by an argon ion laser. Excitation spot size on the surface of the preparation was 60 microns in ... More
Dephosphorylation kinetics of pig kidney Na+,K+-ATPase.
AuthorsKane DJ, Grell E, Bamberg E, Clarke RJ
JournalBiochemistry
PubMed ID9521778
'The kinetics of K+-stimulated dephosphorylation of the Na+,K+-ATPase were investigated at pH 7.4, 24 degrees C, and an ATP concentration of 1.0 mM via the stopped-flow technique using the fluorescent label RH421. Two different mixing procedures were used: (a) premixing with ATP to allow phosphorylation to go to completion, followed ... More
Effect of gramicidin A on the dipole potential of phospholipid membranes.
AuthorsShapovalov VL, Kotova EA, Rokitskaya TI, Antonenko YN
JournalBiophys J
PubMed ID10388758
'The effect of channel-forming peptide gramicidin A on the dipole potential of phospholipid monolayers and bilayers has been studied. Surface pressure and surface potential isotherms of monolayers have been measured with a Langmuir trough equipped with a Wilhelmy balance and a surface potential meter (Kelvin probe). Gramicidin has been shown ... More
Charge translocation by the Na,K-pump: I. Kinetics of local field changes studied by time-resolved fluorescence measurements.
AuthorsBühler R, Stürmer W, Apell HJ, Läuger P
JournalJ Membr Biol
PubMed ID1652643
'Membrane fragments containing a high density of Na,K-ATPase can be noncovalently labeled with amphiphilic styryl dyes (e.g., RH 421). Phosphorylation of the Na,K-ATPase by ATP in the presence of Na+ and in the absence of K+ leads to a large increase of the fluorescence of RH 421 (up to 100%). ... More
Fluorescent styryl dyes of the RH series affect a potential drop on the membrane/solution boundary.
AuthorsMalkov DY, Sokolov VS
JournalBiochim Biophys Acta
PubMed ID8593277
'The effects of the adsorption of the fluorescent potential-sensitive dyes RH-421, RH-237 and RH-160 on the bilayer lipid membrane were studied. It was shown that a dipole potential drop, positive in the hydrophobic part of the membrane, arose due to the dye adsorption. The dye adsorption led to a considerable ... More
Origin of the in vivo rat piriform cortex activity recorded with voltage-sensitive dyes: comparison of the optical signals and the field potentials.
AuthorsLitaudon P, Cattarelli M
JournalBrain Res
PubMed ID1467937
'The comparison of optical recordings and evoked field potentials recorded on the rat piriform cortex pointed out that both signals were strongly correlated. As the field potentials, the two waves of the optical signals originated from the mono- (direct olfactory bulb afferents) and polysynaptic (intrinsic association fibers) excitatory postsynaptic potentials. ... More
Effective gating charge of ion channels induced by toxin syringomycin E in lipid bilayers.
'To elucidate the voltage gating of syringomycin E (SRE) ion channels in lipid bilayers, the effective gating charge q was measured under different conditions. It was shown that q and its sign are dependent on membrane surface charge, dipole potential, and the outer potential (Delta phi). The q values were ... More
Fluorescent styryl dyes as probes for Na,K-ATPase reaction. Enzyme source and fluorescence response.
AuthorsKlodos I, Fedosova NU, Cornelius F
JournalAnn N Y Acad Sci
PubMed ID9405832
Dipole potential drop due to RH-dye adsorption on the lipid bilayer and its influence on Na+/K+-ATPase activity.
AuthorsMalkov DY, Pavlov KV, Sokolov VS
JournalAnn N Y Acad Sci
PubMed ID9432911
Spatially-resolved optical imaging of membrane potentials induced by applied electric fields.
AuthorsGross D, Loew LM, Ryan TA, Webb WW
JournalProg Clin Biol Res
PubMed ID3960915
Optical measurement of membrane potential in cells, organelles, and vesicles.
AuthorsFreedman JC, Novak TS
JournalMethods Enzymol
PubMed ID2747524
Ionics and conformational transitions of Na,K-ATPase.
AuthorsGrell E, Warmuth R, Lewitzki E, Ruf H
JournalActa Physiol Scand Suppl
PubMed ID1333153
Equilibrium binding studies have been carried out by spectrofluorometric precision titrations on FITC-Na,K-ATPase and employing the styryl dye RH-421 to obtain equilibrium constants and stoichiometric coefficients together with information related to competition between different binding equilibria. A new interpretation concerning the assignment of spectral properties and cation complex formation equilibria, ... More
Cable properties of arborized Retzius cells of the leech in culture as probed by a voltage-sensitive dye.
AuthorsFromherz P, Vetter T
JournalProc Natl Acad Sci U S A
PubMed ID1549563
Retzius cells of Hirudo medicinalis were cultivated on extracellular matrix protein so that extended arborizations were formed. The propagation of voltage transients along 1-microns-thick neurites was observed at a resolution of 8 microns at 10 kHz by use of a voltage-sensitive dye. Delay and width of the fluorescence transients caused ... More
Specific RNA binding to ordered phospholipid bilayers.
AuthorsJanas T, Janas T, Yarus M,
JournalNucleic Acids Res
PubMed ID16641318
We have studied RNA binding to vesicles bounded by ordered and disordered phospholipid membranes. A positive correlation exists between bilayer order and RNA affinity. In particular, structure-dependent RNA binding appears for rafted (liquid-ordered) domains in sphingomyelin-cholesterol-1,2-dioleoyl-sn-glycero-3-phosphocholine vesicles. Binding to more highly ordered gel phase membranes is stronger, but much less ... More
Fluorescence recordings of electrical activity in goldfish optic tectum in vitro.
AuthorsManis PB, Freeman JA
JournalJ Neurosci
PubMed ID3339422
Optical methods for recording electrical activity in the goldfish optic tectum were evaluated. Tectal slices, with a short section of the optic nerve attached, were stained with a fluorescent styryl dye. Potential-dependent fluorescence changes following optic nerve stimulation were monitored with a photodiode. We found that large optical signals could ... More
Fluorescence study on cardiac glycoside binding to the Na,K-pump. Ouabain binding is associated with movement of electrical charge.
AuthorsStürmer W, Apell HJ
JournalFEBS Lett
PubMed ID1312482
Recently we have presented evidence that the fluorescence probe RH 421 can be used to detect binding and release of ions at the extracellular face of the pump since these processes are associated with translocation of electrical charge. Applying this method to experiments with cardiac glycosides we found that: (1) ... More
E2P phosphoforms of Na,K-ATPase. I. Comparison of phosphointermediates formed from ATP and Pi by their reactivity toward hydroxylamine and vanadate.
AuthorsFedosova NU, Cornelius F, Klodos I
JournalBiochemistry
PubMed ID9753450
The properties of Na,K-ATPase phosphoenzymes formed either from ATP in the presence of Mg2+ and Na+ or from Pi in the absence of alkali cations were investigated by biochemical methods and spectrofluorometry employing the styryl dye RH421. We characterized the phosphoenzyme species by their reaction to N-methyl hydroxylamine, which attacks ... More
Voltage-sensitive fluorescence of amphiphilic hemicyanine dyes in neuron membrane.
AuthorsFromherz P, Müller CO
JournalBiochim Biophys Acta
PubMed ID8347665
Fluorescent amphiphilic hemicyanine dyes were adsorbed to the plasma membrane of isolated Retzius neurons of the leech. Voltage steps were applied to the neuron by the patch-clamp technique in whole-cell configuration. The change of fluorescence was observed as induced by the voltage jump. The relative changes of the excitation spectrum ... More
Cable properties of a straight neurite of a leech neuron probed by a voltage-sensitive dye.
AuthorsFromherz P, Müller CO
JournalProc Natl Acad Sci U S A
PubMed ID8183956
We measured a time-resolved map of electrical activity in a thin straight neurite (1.5 microns thick, 500 microns long) at a resolution of 8 microns and 0.4 ms. The neurite was obtained by guided outgrowth of an identified neuron of the leech on lanes of extracellular matrix protein. The electrical ... More
Optical imaging of the heart.
AuthorsEfimov IR, Nikolski VP, Salama G
JournalCirc Res
PubMed ID15242982
Optical techniques have revolutionized the investigation of cardiac cellular physiology and advanced our understanding of basic mechanisms of electrical activity, calcium homeostasis, and metabolism. Although optical methods are widely accepted and have been at the forefront of scientific discoveries, they have been primarily applied at cellular and subcellular levels and ... More
Effect of the dipole potential of a bilayer lipid membrane on gramicidin channel dissociation kinetics.
AuthorsRokitskaya TI, Antonenko YN, Kotova EA
JournalBiophys J
PubMed ID9251801
A technique of measuring of the light-induced transients of the gramicidin-mediated electric current across a membrane in the presence of a photosensitizer has been applied for the study of the effect of agents modifying the dipole potential of a bilayer lipid membrane (phloretin, 6-ketocholestanol, and RH421) on the processes of ... More
Static and dynamic studies of the potential-sensitive membrane probe RH421 in dimyristoylphosphatidylcholine vesicles.
AuthorsZouni A, Clarke RJ, Visser AJ, Visser NV, Holzwarth JF
JournalBiochim Biophys Acta
PubMed ID8274489
The dynamics of the potential-sensitive styryl dye RH421 in dimyristoylphosphatidylcholine vesicles have been investigated above and below the main phase transition temperature using iodine-laser temperature-jump relaxation spectrophotometry and time-resolved fluorescence lifetime measurements. Equilibrium fluorescence titrations have shown that the affinity of the dye for the membrane is much higher in ... More
Optical detection of neuromodulatory effects of conditioned taste aversion in the pond snail Lymnaea stagnalis.
AuthorsKojima S, Hosono T, Fujito Y, Ito E
JournalJ Neurobiol
PubMed ID11598919
Multiple site optical recording was used to analyze the neural activity changes caused by conditioned taste aversion (CTA) training in the pond snail Lymnaea stagnalis. In response to electrical stimulation of the median lip nerve, which transmits chemosensory signals of appetitive taste to the central nervous system, we optically detected ... More
Bcl-2 on the endoplasmic reticulum regulates Bax activity by binding to BH3-only proteins.
AuthorsThomenius MJ, Wang NS, Reineks EZ, Wang Z, Distelhorst CW
JournalJ Biol Chem
PubMed ID12477729
Bcl-2 family members have been shown to be key mediators of apoptosis as either pro- or anti-apoptotic factors. It is thought that both classes of Bcl-2 family members act at the level of the mitochondria to regulate apoptosis, although the founding anti-apoptotic family member, Bcl-2 is localized to the endoplasmic ... More
Mechanism of the conformational change in sodium pump reported by eosin.
AuthorsSmirnova IN, Faller LD
JournalBiochemistry
PubMed ID7548078
The conformational change in sodium pump reported by eosin has been reinvestigated because of apparent discrepancies between results obtained in other laboratories with this reversibly-bound, fluorescent reporter group and results obtained in our laboratory with covalently bound fluorescein. The study demonstrates that both probes report the same reaction. Data obtained ... More
Spectroscopic investigations of the potential-sensitive membrane probe RH421.
AuthorsClarke RJ, Schrimpf P, Schöneich M
JournalBiochim Biophys Acta
PubMed ID1329964
The absorbance spectra, fluorescence emission and excitation spectra, and fluorescence anisotropy of the potential-sensitive styryl dye RH421 have been investigated in aqueous solution and bound to the lipid membrane. The potential-sensitive response of the dye has been studied using a preparation of membrane fragments containing a high density of Na+, ... More
Influence of anions and cations on the dipole potential of phosphatidylcholine vesicles: a basis for the Hofmeister effect.
AuthorsClarke RJ, Lüpfert C
JournalBiophys J
PubMed ID10233076
Anions and cations have long been recognized to be capable of modifying the functioning of various membrane-related physiological processes. Here, a fluorescent ratio method using the styrylpyridinium dyes, RH421 and di-8-ANEPPS, was applied to determine the effect of a range of anions and cations on the intramembrane dipole potential of ... More
Binding of sodium ions and cardiotonic steroids to native and selectively trypsinized Na,K pump, detected by charge movements.
A fluorescent dye, RH421, has been used to characterize charge movements associated with cation and cardiotonic steroid binding to Na,K-ATPase and to a specifically trypsinized preparation, so-called "19-kDa membranes." A fluorescence decrease induced by Na+ is attributed to electrogenic binding of one Na+ ion from the cytoplasm. The apparent affinity ... More
Voltage sensitivity of the fluorescent probe RH421 in a model membrane system.
AuthorsClarke RJ, Zouni A, Holzwarth JF
JournalBiophys J
PubMed ID7787026
The voltage sensitivity of the fluorescent styrylpyridinium dye RH421 has been investigated in dimyristoylphosphatidylcholine vesicles by inducing an intramembrane electric field through the binding of the hydrophobic ion tetraphenylborate (TPB). To assess the probability of electrochromic and solvatochromic mechanisms for the dye response, the ground-state dipole moment of the dye ... More
Hofmeister effects of anions on the kinetics of partial reactions of the Na+,K+-ATPase.
AuthorsGanea C, Babes A, Lüpfert C, Grell E, Fendler K, Clarke RJ
JournalBiophys J
PubMed ID10388756
The effects of lyotropic anions, particularly perchlorate, on the kinetics of partial reactions of the Na+,K+-ATPase from pig kidney were investigated by two different kinetic techniques: stopped flow in combination with the fluorescent label RH421 and a stationary electrical relaxation technique. It was found that 130 mM NaClO4 caused an ... More
Interaction of the fluorescent probe RH421 with ribulose-1,5-bisphosphate carboxylase/oxygenase and with Na+,K(+)-ATPase membrane fragments.
AuthorsFrank J, Zouni A, van Hoek A, Visser AJ, Clarke RJ
JournalBiochim Biophys Acta
PubMed ID8634316
Fluorescence titrations have shown that the voltage-sensitive probe RH421 interacts with the water-soluble protein ribulose-1,5-bisphosphate carboxylase/oxygenase and with Na+,K(+)-ATPase membrane fragments. The probe exhibits significantly different fluorescence excitation spectra in pure lipid and pure protein environments. Experiments with a range of polyamino acids showed interactions of the probe with tyrosine, ... More
Fluorescent styryl dyes as probes for Na,K-ATPase reaction mechanism: significance of the charge of the hydrophilic moiety of RH dyes.
AuthorsFedosova NU, Cornelius F, Klodos I
JournalBiochemistry
PubMed ID8527456
The fluorescence responses of a series of potential-sensitive styryl-based dyes (either zwitterionic RH160, RH421, di-4-ANEPPS, or positively charged RH795, RH414, RH461) to phosphorylation of Na,K-ATPase from ATP or inorganic phosphate, and ouabain binding to phospho- or dephosphoforms, have been characterized and compared in broken membrane preparations of the enzyme. Zwitterionic ... More
Charge translocation by the Na,K-pump: II. Ion binding and release at the extracellular face.
AuthorsStürmer W, Bühler R, Apell HJ, Läuger P
JournalJ Membr Biol
PubMed ID1652644
In the first part of the paper, evidence has been presented that electrochromic styryl dyes, such as RH 421, incorporate into Na,K-ATPase membranes isolated from mammalian kidney and respond to changes of local electric field strength. In this second part of the paper, fluorescence studies with RH-421-labeled membranes are described, ... More
Time-resolved fluorescence investigations of the interaction of the voltage-sensitive probe RH421 with lipid membranes and proteins.
AuthorsVisser NV, van Hoek A, Visser AJ, Frank J, Apell HJ, Clarke RJ
JournalBiochemistry
PubMed ID7547910
Fluorescence lifetimes and fluorescence anisotropy decays of the voltage-sensitive styryl-pyridinium dye RH421 have been measured in the presence of dimyristoylphosphatidylcholine vesicles, the water soluble enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase (rubisco), and Na+,K(+)-ATPase-containing membrane fragments. The effect of an intramembrane electric field on the photophysical properties of the dye was investigated by ... More
Rapid kinetic analyses of the Na+/K(+)-ATPase distinguish among different criteria for conformational change.
AuthorsPratap PR, Robinson JD
JournalBiochim Biophys Acta
PubMed ID8395217
The Na+/K(+)-ATPase couples the hydrolysis of ATP to the transport of Na+ and K+ via a phosphorylated intermediate and conformational changes. In order to identify these conformational changes, we have probed the sequence of steps from EP(3Na+ in) to EP + 3Na+ out with three fluorescent probes (IAF: 5-iodoacetamidofluorescein; BIPM: ... More
Partial reactions of the Na,K-ATPase: determination of rate constants.
AuthorsHeyse S, Wuddel I, Apell HJ, Stürmer W
JournalJ Gen Physiol
PubMed ID7807047
Experiments were designed to characterize several partial reactions of the Na,K-ATPase and to demonstrate that a model can be defined that reproduces most of the transport features of the pump with a single set of kientic parameters. We used the fluorescence label 5-iodoacetamidofluorescein, which is thought to be sensitive to ... More
Kinetics of Na(+)-dependent conformational changes of rabbit kidney Na+,K(+)-ATPase.
AuthorsClarke RJ, Kane DJ, Apell HJ, Roudna M, Bamberg E
JournalBiophys J
PubMed ID9726935
The kinetics of Na(+)-dependent partial reactions of the Na+,K(+)-ATPase from rabbit kidney were investigated via the stopped-flow technique, using the fluorescent labels N-(4-sulfobutyl)-4-(4-(p-(dipentylamino)phenyl)butadienyl)py ridinium inner salt (RH421) and 5-iodoacetamidofluorescein (5-IAF). When covalently labeled 5-IAF enzyme is mixed with ATP, the two labels give almost identical kinetic responses. Under the chosen ... More
Rate limitation of the Na(+),K(+)-ATPase pump cycle.
AuthorsLüpfert C, Grell E, Pintschovius V, Apell HJ, Cornelius F, Clarke RJ
JournalBiophys J
PubMed ID11566779
The kinetics of Na(+)-dependent phosphorylation of the Na(+),K(+)-ATPase by ATP were investigated via the stopped-flow technique using the fluorescent label RH421 (saturating [ATP], [Na(+)], and [Mg(2+)], pH 7.4, and 24 degrees C). The well-established effect of buffer composition on the E(2)-E(1) equilibrium was used as a tool to investigate the ... More
Stopped-flow kinetic investigations of conformational changes of pig kidney Na+,K+-ATPase.
AuthorsKane DJ, Fendler K, Grell E, Bamberg E, Taniguchi K, Froehlich JP, Clarke RJ
JournalBiochemistry
PubMed ID9341234
The kinetics of Na+-dependent partial reactions of the Na+,K+-ATPase were investigated via the stopped-flow technique using the fluorescent labels RH421 and BIPM. After the enzyme is mixed with MgATP, both labels give almost identical kinetic responses. Under the chosen experimental conditions two exponential time functions are necessary to fit the ... More
Regulation of eNOS expression in brain endothelial cells by perinuclear EP(3) receptors.
AuthorsGobeil F, Dumont I, Marrache AM, Vazquez-Tello A, Bernier SG, Abran D, Hou X, Beauchamp MH, Quiniou C, Bouayad A, Choufani S, Bhattacharya M, Molotchnikoff S, Ribeiro-Da-Silva A, Varma DR, Bkaily G, Chemtob S
JournalCirc Res
PubMed ID11934836
We reported upregulation of endothelial nitric oxide synthase (eNOS) by PGE(2) in tissues and presence of perinuclear PGE(2) receptors (EP). We presently studied mechanisms by which PGE(2) induces eNOS expression in cerebral microvessel endothelial cells (ECs). 16,16-Dimethyl PGE(2) and selective EP(3) receptor agonist M&B28767 increased eNOS expression in ECs and ... More