The plasma membrane and mitochondrial membrane potentials of Plasmodium yoelii

1. The plasma membrane potential and the mitochondrial membrane potential of P. yoellii was examined by fluorescence microscopy using rhodamine 123 and by transmembrane distribution of tetraphenylphosphonium. 2. The mitochondrion of P. yoelii, free of gametocyte stage, maintained a high negative inside membrane potential. 3. Deprivation of glucose in incubation medium largely abolished the plasma membrane potential but not the mitochondrial membrane potential. 4. Studies with metabolic inhibitors showed that the mitochondrial membrane potential constituted a marginal portion as compared with the plasma membrane potential in intact infected erythrocytes.     

Control of membrane potential by external H+ concentration in Bacillus subtilis as determined by an ion-selective electrode

The membrane potential of intact bacteria was monitored by measuring the tetraphenylphosphonium ion distribution across the membrane using poly--(vinyl chloride) matrix-type electrode selective to tetraphenylphosphonimum ion. It was found that the tetraphenylphosphonium ion was not countertransported against H+ movement. The membrane potential of Bacillus subtilis was estimated to be 80-120 mV inside-negative at external pH 7. The effect of the external pH on the membrane potential was studied. It varied from 30 to 40 mV/decade change in the external [H+] in the pH region of greater than 6.5, increasing pH making it more inside-negative. The addition of carbonyl cyanide m-chlorophenylhydrazone depolarized the membrane, and the membrane potential approached the H+ equilibrium potential. The addition of N,N'-dicyclohexylcarbodiimide did not abolish the pH dependence of the membrane potential. Increasing the external [K+] did not affect the pH dependence. CN- partially depolarized the membrane. A parallel conductance model for membrane potential could explain the results qualitatively.     

The use of the tip potential of glass microelectrodes in the determination of low cell membrane potentials

Summary The tip potential of Ling-Gerard glass microelectrodes changes upon insertion into cells and thus impedes the determination of the actual membrane potential. The lower the membrane potential of a cell, the larger will be the error due to this tip potential. However, as is demonstrated, a relationship exists between the tip potential of the electrode and the measured potential difference, which allows the determination of the membrane potential of a particular cell type by linear regression. This method showed that resting lymphocytes had no membrane potential, whereas for the slime mouldDictyostelitim discoideum a membrane potential of about –9 mV could be calculated.Dedicated to Prof. Dr. Dr. h.c. mult. B. Rajewsky on the occasion of his 80th birthday.     

The membrane potential of the cellular slime mold Dictyostelium discoideum is mainly generated by an electrogenic proton pump

Trans membrane potential or ionic current changes may play a role in signal transduction and differentiation in the cellular slime mold dictyostelium discoideum. Therefore, the contribution of electrogenic ion pumps to the membrane potential of D. discoideum cells was investigated. the (negative) peak-value of the rapid potential transient, seen upon microelectrode impalement, was used to detect membrane potential changes upon changes in the external pH in the range of 5.5 to 8.0. The membrane potential was close to the Nernstian potential for protons over the pH range 5.5 to 7.5. The acid-induced changes in membrane potential were consistent with outward-proton pumping. The maximal membrane potential was at pH 7.5. Furthermore, the proton pump inhibitors diethylstilbestrol, miconazole and zearalenone directly depolarize the membrane. Cyanide and temperature decrease cause membrane depolarization as well. During recovery from cyanide poisoning a H+ efflux is present. From these measurements we conclude that the membrane potential of d. discoideum cells is mainly generated by an electrogenic proton pump. Measurements in cells with different extracellular potassium and H+ concentrations suggest a role for potassium in the function of the electrogenic proton pump. These results provide a framework for future research towards a possible role for the proton pump in signal transduction and differentiation.     

An electrophysiological study of the non-junctional membrane of epidermal cells in Tenebrio molitor

The contribution of K and Cl to the membrane potential of the epidermal cells of the recently-ecdysed larva of the mealworm was examined. The ionic basis for the membrane potential is complex. Although increasing the external K level depolarized the cell membrane, the relationship obtained suggests that ions other than K contribute largely to the recorded membrane potential. In particular, exposing the cells to K concentrations below the normal level of 40 mM has only slight effects on membrane potential, irrespective of whether K is lowered by direct substitution with Na or under conditions in which Na and Cl levels are held constant. Increasing the external Cl levels from 4 mM to 154 mM while holding K and Na levels constant resulted in a 10 mV hyperpolarization. The slight hyperpolarizing effects of high external Cl could be mimicked by citrate, but not by acetate, the latter drastically hyperpolarizing the cell membrane at levels of K that normally maintain a reduced membrane potential. External Na has little effect on the membrane potential at normal physiological levels of K, but may depolarize the cell at low K levels. The results suggest that several inorganic ions, and possibly organic acids, participate in generating the membrane potential of the epidermal cell. The passive ionic properties of non-junctional epidermal membrane and muscle membrane appear to the similar in this insect.The electrical resistance on the non-junctional membrane is highly dependent on the external K level, and can be reduced by three orders of magnitude by increasing external K from 1 mM to 120 mM. The resistance of the junctional membrane remains constant over this range of external K concentrations.     

The motile and tactic behaviour of Pseudomonas aeruginosa in anaerobic environments

ATP generated by the anaerobic metabolism of L-arginine in Pseudomonas aeruginosa was used to maintain the membrane potential. Although both the ATP concentration and membrane potential were lower than in aerobically incubated bacteria, motility and chemotaxis were almost normal. Venturicidin stopped anaerobic motility by abolishing the membrane potential. The addition of venturicidin to aerobic bacteria caused an increase in the membrane potential, but a decrease in internal ATP concentration, resulting in bacteria which were motile but non-chemotactic. The membrane potential was the only requirement for continued motility but ATP was required in addition for chemotaxis.     

Effect of bovine serum albumin on membrane potential in plant mitochondria

The membrane potential in highly coupled potato ( Solanum tuberosum L.) mitochondria, as measured by changes in safranine absorbance, was significantly increased by addition of bovine serum albumin. Purification of potato mitochondria on Percoll, in removing 50% of free unsaturated fatty acids, decreased the BSA-de-pendent membrane potential. The effect of added linoleic acid and of the natural accumulation of fatty acids during aging was studied. The response of membrane potential to addition of bovine serum albumin appeared to be directly correlated to the amount of free unsaturated fatty acids. Aging in vitro, in releasing free fatty acids, decreased respiratory control and ADP:O ratios and collapsed the membrane potential. During 2–3 h of incubation, addition of BSA completely restored membrane potential and oxidative phosphorylation.
It is concluded that both in fresh and in aged potato mitochondria the effect of bovine serum albumin on oxidative phosphorylation can be ascribed to an effect on membrane permeability to ions. BSA, in binding free unsaturated fatty acids, restored maximal membrane potential. The bovine serum albumin-dependent membrane potential appears to be a sensitive criterion of the functional integrity of the inner mitochondrial membrane.     

Platelet membrane potential as a modulator of aggregating mechanisms

The membrane potential of platelets suspended in physiological medium and membrane potential changes induced by high potassium concentrations, ouabain and cooling have been measured using a cyanine fluorescent dye (3,3'-dipropylthiodicarbocyanine) [corrected]. The membrane potential of platelets suspended in physiological medium was -63.8 mV. High potassium concentrations, ouabain and cooling induced depolarization of platelet membrane. Depolarization using the above procedures enhanced platelet aggregation induced by ADP, adrenaline and collagen. These results suggest that the membrane potential could modulate platelet activity.     

Model for the dynamic responses of taste receptor cells to salty stimuli. I. Function of lipid bilayer membranes.

The dynamic response of the lipid bilayer membrane is studied theoretically using a microscopic model of the membrane. The time courses of membrane potential variations due to monovalent salt stimulation are calculated explicitly under various conditions. A set of equations describing the time evolution of membrane surface potential and diffusion potential is derived and solved numerically. It is shown that a rather simple membrane such as lipid bilayer has functions capable of reproducing the following properties of dynamic response observed in gustatory receptor potential. Initial transient depolarization does not occur under Ringer adaptation but does under water. It appears only for comparatively rapid flows of stimuli, the peak height of transient response is expressed by a power function of the flow rate, and the membrane potential gradually decreases after reaching its peak under long and strong stimulation. The dynamic responses in the present model arise from the differences between the time dependences in the surface potential phi s and the diffusion potential phi d across a membrane. Under salt stimulation phi d cannot immediately follow the variation in phi s because of the delay due to the charging up of membrane capacitance. It is suggested that lipid bilayer in the apical membrane is the most probable agency producing the initial phasic response to the stimulation.     

Current-voltage relationships for proton flow through the F0 sector of the ATP-synthase, carbonylcyanide-p-trifluoromethoxyphenylhydrazone or leak pathways in submitochondrial particles

Respiring submitochondrial particles from which the F1 sector of ATP-synthase was displaced generated a membrane potential in the range of 115-140 mV. Addition of oligomycin raised the membrane potential by approximately 40 mV. The lower membrane potential in particles with F1 displaced is attributed to partial dissipation of the proton electrochemical gradient as a consequence of proton flow through the open proton channels provided by the F0 sectors of the ATP-synthase. The characteristics of proton flow through the open F0 channels were studied by varying the rate of electron transport-driven proton translocation which permitted the establishment of a range of steady-state membrane potentials. Open F0 channels appeared to have a gated response to the membrane potential such that they were inoperative when the potential fell below approximately 110 mV. The membrane potential was measured as a function of respiratory rate in intact Mg-ATP submitochondrial particles that had been treated with low concentrations of the protonophore carbonylcyanide-p-trifluoromethoxyphenylhydrazone. In general a linear dependence of membrane potential upon respiratory rate was observed except at the lowest concentrations of protonophore and highest respiratory rates, presumably because the effect of the protonophore was then offset by an increased rate of proton translocation driven by the respiratory chain. The effect of increasing concentrations of carbonylcyanide-p-trifluoromethoxyphenylhydrazone on the membrane potential of respiring submitochondrial particles was studied. It was found that equal amounts of the protonophore lowered the membrane potential to a lesser extent at lower values of the membrane potential. Treatment of Mg-ATP submitochondrial particles with oligomycin slightly increased (by approximately 10 mV) the size of the respiration-dependent membrane potential, but did not alter the profile of membrane potential as a function of succinate oxidation rate. The latter was controlled by titration with malonate. This result indicates that the F0 sector of the ATP-synthase does not significantly contribute to leak pathways in intact submitochondrial particles.     

Membrane protein topology: effects of delta mu H+ on the translocation of charged residues explain the ''positive inside'' rule.

The membrane electrochemical potential is critical for the export of most periplasmic proteins in Escherichia coli. Its exact role during insertion of integral inner membrane proteins, however, remains obscure. Using derivatives of the inner membrane protein leader peptidase (Lep), we now show that the membrane potential appears to stimulate the membrane translocation of chain segments containing negatively charged residues, that positively charged regions appear to be more easily translocated in the absence of a potential, and that certain Lep constructs insert with different topologies in the presence and absence of a membrane potential, suggesting that the electrochemical potential introduces an asymmetry between the topological effects of positively and negatively charged amino acids during the process of membrane protein insertion in E. coli.     

Mechanism of neurotensin depolarization of rabbit colonic smooth muscle

The aims of this study were (1) to measure the effect of neurotensin on the membrane potential of circular muscle of the distal colon of the rabbit and (2) to determine the mechanism by which neurotensin affects the membrane potential of this tissue. The membrane potential was measured with microelectrodes placed intracellularly and the double sucrose gap. Neurotensin (10(-11) M to 10(-7) M) dose-dependently decreased the membrane potential. The maximum decrease in membrane potential occurred with 10(-9) M neurotensin. The ED50 of neurotensin depolarization of the membrane potential was 0.87 +/- 0.33 X 10(-10) M. The frequency of the slow waves was unchanged after neurotensin. The voltage response to a constant current pulse decreased as the concentration of neurotensin increased. The amplitude of the voltage response after a 0.6 microA current pulse decreased by 6 +/- 0.5 mV after neurotensin (10(-7) M) compared to the Krebs control (P less than 0.05). Decreasing the [Na+]o to 0-23 mM did not affect the decrease in membrane potential after neurotensin. However, perfusion with a test solution containing no added Ca2+ or verapamil (10(-5) M) inhibited neurotensin depolarization of the tissue. Evidence was found that neurotensin depolarizes colonic circular smooth muscle, and the decrease in membrane potential is associated with an increase in conductance which is dependent on influx of Ca2+.     

Membrane Potential Fluctuations Determine the Precision of Spike Timing and Synchronous Activity: A Model Study

It is much debated on what time scale information is encoded by neuronal spike activity. With a phenomenological model that transforms time-dependent membrane potential fluctuations into spike trains, we investigate constraints for the timing of spikes and for synchronous activity of neurons with common input. The model of spike generation has a variable threshold that depends on the time elapsed since the previous action potential and on the preceding membrane potential changes. To ensure that the model operates in a biologically meaningful range, the model was adjusted to fit the responses of a fly visual interneuron to motion stimuli. The dependence of spike timing on the membrane potential dynamics was analyzed. Fast membrane potential fluctuations are needed to trigger spikes with a high temporal precision. Slow fluctuations lead to spike activity with a rate about proportional to the membrane potential. Thus, for a given level of stochastic input, the frequency range of membrane potential fluctuations induced by a stimulus determines whether a neuron can use a rate code or a temporal code. The relationship between the steepness of membrane potential fluctuations and the timing of spikes has also implications for synchronous activity in neurons with common input. Fast membrane potential changes must be shared by the neurons to produce synchronous activity.     

Characterization of the plasma and mitochondrial membrane potentials of alveolar type II cells by the use of ionic probes

The lipophilic cation triphenylmethylphosphonium (TPMP+) and the potassium analog Rb+, were used to monitor the membrane potential (delta psi) of freshly isolated rabbit type II alveolar epithelial cells. Type II cells were found to accumulate TPMP+ rapidly at 37 degrees C in Hanks' balanced-salt solution with 5 microM tetraphenyl boron, but this accumulation was partially due to non-membrane potential dependent binding of TPMP+ to the cell. Lysophosphatidylcholine (lysoPC) was found to abolish delta psi and permitted correction for bound TPMP+ or Rb+. TPMP+ remaining in the cell following correction for binding represents the sum of mitochondrial and plasma membrane potential dependent accumulation. The accumulation of Rb+ by the type II cell was found to be independent of the mitochondrial membrane potential and indicated a trans-plasma membrane Rb+ distribution potential of -62.9 +/- 4 mV. A similar value was obtained by estimating the plasma membrane potential dependent accumulation of TPMP+ in type II cells whose mitochondria were depolarized with carbonylcyanide m-chlorophenylhydrazone (CCCP). The release of TPMP+ due to CCCP treatment also permitted an estimation for the trans-mitochondrial membrane potential of -141.8 +/- 10 mV. These techniques of membrane potential measurements were found to be sensitive to changes in delta psi induced by a number of inhibitors and ionophores. The ability to measure the membrane potential of the type II pneumocyte, and the changes caused by various agents, should be useful in characterizing the functional responses of this pulmonary surfactant producing cell.     

Inner field compensation as a tool for the characterization of asymmetric membranes and Peptide-membrane interactions

Symmetric and asymmetric planar lipid bilayers prepared according to the Montal-Mueller method are a powerful tool to characterize peptide-membrane interactions. Several electrical properties of lipid bilayers such as membrane current, membrane capacitance, and the inner membrane potential differences and their changes can be deduced. The time-resolved determination of peptide-induced changes in membrane capacitance and inner membrane potential difference are of high importance for the characterization of peptide-membrane interactions. Intercalation and accumulation of peptides lead to changes in membrane capacitance, and membrane interaction of charged peptides induces changes in the charge distribution within the membrane and with that to changes in the membrane potential profile. In this study, we establish time-resolved measurements of the capacitance minimization potential DeltaPsi on various asymmetric planar lipid bilayers using the inner field compensation method. The results are compared to the respective ones of inner membrane potential differences DeltaPhi determined from ion carrier transport measurements. Finally, the time courses of membrane capacitances and of DeltaPsi have been used to characterize the interaction of cathelicidins with reconstituted lipid matrices of various Gram-negative bacteria.     

Mechanism of action of the peptide antibiotic nisin in liposomes and cytochrome c oxidase-containing proteoliposomes.

The interaction of the peptide antibiotic nisin with liposomes has been studied. The effect of this interaction was analyzed on the membrane potential (inside negative) and the pH gradient (inside alkaline) in liposomes made from Escherichia coli phosphatidylethanolamine and egg phosphatidylcholine (9:1, wt/wt). The membrane potential and pH gradient were generated by artificial ion gradients or by the oxidation of ascorbate, N,N,N',N'-tetramethyl-p-phenylenediamine, and cytochrome c by the beef heart cytochrome c oxidase incorporated in the liposomal membranes. Nisin dissipated the membrane potential and the pH gradient in both types of liposomes and inhibited oxygen consumption by cytochrome c oxidase in proteoliposomes. The dissipation of the proton motive force in proteoliposomes was only to a minor extent due to a decrease of the oxidase activity by nisin. The results in these model systems show that a membrane potential and/or a pH gradient across the membrane enhances the activity of nisin. Nisin incorporates into the membrane and makes the membrane permeable for ions. As a result, both the membrane potential and pH gradient are dissipated. The activity of nisin was found to be influenced by the phospholipid composition of the liposomal membrane.     

Generation of membrane potential beyond the conceptual range of Donnan theory and Goldman-Hodgkin-Katz equation

Donnan theory and Goldman-Hodgkin-Katz equation (GHK eq.) state that the nonzero membrane potential is generated by the asymmetric ion distribution between two solutions separated by a semipermeable membrane and/or by the continuous ion transport across the semipermeable membrane. However, there have been a number of reports of the membrane potential generation behaviors in conflict with those theories. The authors of this paper performed the experimental and theoretical investigation of membrane potential and found that (1) Donnan theory is valid only when the macroscopic electroneutrality is sufficed and (2) Potential behavior across a certain type of membrane appears to be inexplicable on the concept of GHK eq. Consequently, the authors derived a conclusion that the existing theories have some limitations for predicting the membrane potential behavior and we need to find a theory to overcome those limitations. The authors suggest that the ion adsorption theory named Ling’s adsorption theory, which attributes the membrane potential generation to the mobile ion adsorption onto the adsorption sites, could overcome those problems.     

栀子提取物ZG对副流感病毒1型感染后宿主细胞膜的影响

为了探讨栀子提取物ZG抗病毒作用的生物学机制,观察了栀子提取物ZG对副流感病毒1型(PIV-1)感染后宿主细胞膜电位、膜Na -K -ATP酶活性和膜流动性的影响。以氯化乙酰胆碱为阳性对照,采用荧光探针Di-BAC4(3)标记Hep-2细胞膜电位,借助流式细胞仪检测膜电位;定磷法,分光光度计检测Na -K -ATP酶活性;荧光探针NBD-C6-HPC标记细胞膜磷脂,以荧光漂白恢复法和激光扫描共聚焦显微镜检测膜流动性。结果显示:PIV-1感染后宿主细胞膜电位下降,处于超极化状态;膜Na -K -ATP酶活性显著增加,膜流动性显著降低。栀子提取物ZG作用后,对宿主细胞膜的超极化状态没有明显影响;对膜Na -K -ATP酶活性没有明显影响;而对膜流动性则有明显的恢复作用。阳性对照药乙酰胆碱能明显改善病毒感染后膜电位的超极化状态。PIV-1感染后膜电位、Na -K -ATP酶活性和膜流动性等细胞膜能态和功能的改变,可能为病毒感染的生物学机制之一;栀子提取物ZG可能是通过改善细胞膜流动性,维持细胞膜的正常功能来发挥抗病毒感染的作用,而与膜电位和膜Na -K -ATP酶活性等能态来源的环节可能无关。     

Energy transduction in intact synaptosomes. Influence of plasma-membrane depolarization on the respiration and membrane potential of internal mitochondria determined in situ.

A method is described, based on the differential accumulation of Rb+ and methyltriphenylphosphonium, for the simultaneous estimation of the membrane potentials across the plasma membrane of isolated nerve endings (synaptosomes), and across the inner membrane of mitochondria within the synaptosomal cytoplasm. These determinations, together with measurements of respiratory rates, and ATP and phosphocreatine concentrations, are used to define the bioenergetic behaviour of isolated synaptosomes under a variety of conditions. Under control conditions, in the presence of glucose, the plasma and mitochondrial membrane potentials are respectively 45 and 148mV. Addition of a proton translocator induces a 5-fold increase in respiration, and abolishes the mitochondrial membrane potential. The addition of rotenone to inhibit respiration does not affect the plasma membrane potential, and only lowers the mitochondrial membrane potential to 128mV. Evidence is presented that ATP synthesis by anaerobic glycolysis is sufficient under these conditions to maintain ATP-dependent processes, including the reversal of the mitochondrial ATP synthetase. Addition of oligomycin under non-respiring conditions leads to a complete collapse of the mitochondrial potential. Even under control conditions the plasma membrane (Na+ + K+)-dependent ATPase is responsible for a significant proportion of the synaptosomal ATP turnover. Veratridine greatly increases respiration, and depolarizes the plasma membrane, but only slightly lowers the mitochondrial membrane potential. High K+ and ouabain also lower the plasma membrane potential without decreasing the mitochondrial membrane potential. In non-respiring synaptosomes, anaerobic glycolysis is incapable of maintaining cytosolic ATP during the increased turnover induced by veratridine, and the mitochondrial membrane potential collapses. It is concluded that the internal mitochondria must be considered in any study of synaptosomal transport.     

Modelling of pattern formation and oscillations in pH and transmembrane potential near the cell membrane of Chara corallina

A mathematical model of potencial-dependent proton transfer across the membrane of Chara corallina cells is considered. To construct the model, partial differential equations describing the system dynamics in time and in space were used. The variables of the model are the proton concentration and membrane potential. The model describes the experimentally observed inhomogeneous distribution of transmembrane potential and pH along the membrane and oscillations of the potential and pH in time. A mechanism of the distribution of pH and membrane potential along the Chara corallina cell is suggested.