Effects of systematically administered lithium on tryptophan transport and exchange in plasma-membrane vesicles isolated from rat brain

The effect of lithium on the sodium-dependent high-affinity system for tryptophan uptake was examined in plasma membrane vesicles derived from rat brain. We demonstrated that Na⁺ could be replaced by lithium in the external medium and the presence of lithium produced an increase in theV _max of the tryptophan transport whereas it had no significant effect on theK _m for the substrate. Plasma membrane vesicles derived from synaptosomes obtained from long-term lithium-treated rats are able to accumulate tryptophan to a greater extent than normal rats and maintain a more negative membrane potential than controls. Our data support the idea that the stimulation by lithium of the high-affinity uptake system for tryptophan by maintaining adequate membrane potentials across the membrane, could lead to the stabilization of serotonin production, as has been demonstrated in long termlithium treatment.     

Comparison of double layer potentials in lipid monolayers and lipid bilayer membranes

Summary It is shown that the Gouy-Chapman double layer analysis adequately describes the variation of the surface potential of monolayers of acidic natural lipids over a wide range of surface charge density and salt concentration. It is also shown that the potential which initially appears when an electrolyte gradient is rapidly imposed across a bilayer membrane is due to a difference in the double layer potentials on the two sides of the membrane. This conclusion follows from the fact that the observed bilayer potentials arise much more rapidly than can be accounted for by charge migration across the membrane and from the observation that the bilayer membrane concentration potentials, when measured immediately after establishment of a gradient, are equal to the surface potential change observed when the subphase concentration of a monolayer of the same lipid is changed by an amount equal to the gradient across the bilayer. The bilayer potential and monolayer potential changes, so measured, agree in a number of different electrolyte solutions over a wide range of electrolyte concentrations and surface charge densities. Because of this agreement and the applicability of the Gouy theory to monolayers, initial bilayer potentials may be calculated if the composition of the mixture used to form the membrane is known, provided that the pK's and areas of such components are available. In the absence of this information, membrane potentials may be calculated from electrophoretic data on the membrane lipid mixture; the conditions under which the latter approach is possible have been determined. The experimental results indicate that the composition of monolyers and bilayers spread from the same lipid mixture in decane are very similar, that the composition of the two types of film closely resembles the composition of the solution used to generate them, and that bilayer membranes are close-packed. The evidence further indicates that if any hydrocarbon solvent remains in these bilayers, it must be so situated that it contributes little, if anything, to the surface area. The steady state potential in the bilayer membrane system is frequently not identical with the initial potential which supports the hypothesis that in many cases only a fraction of the electrical conductance of unmodified membranes is caused by the ions which constitute the bulk electrolyte. An expression for the relationship between diffusion and double layer potentials has been derived which shows that, in the absence of any intrinsic selectivity of the hydrocarbon region of the membrane for hydrogen, hydroxyl, or impurity, the two potentials should be identical.     

ATP-and NADH-dependent membrane potential generation in plasmalemma enriched vesicles from parenchyma of dormant and non-dormant jerusalem artichoke tubers

Using a membrane potential probe, Oxonol VI, it was possible to demonstrate generation of ATP- and NADH-dependent membrane potential across the plasmalemma, with membrane vesicles derived from parenchyma cells of Jerusalem artichoke tubers(Helianthus tuberosus L.). It was shown that ATP- and NADH-dependent membrane potential generation was higher in dormant material than in non-dormant tissue and that the effects of ATP and NADH on membrane potential generation were additive. ATP-dependent potential generation was sensitive to vanadate, an inhibitor of plasmalemma ATPase activity. The results are discussed in relation to the properties of the different enzymes bound to the plasma membrane, the morphogenetic potentialities of tuber buds and the hypothesis that tuber dormancy could be an extreme case of nutrient deficiency induced by short-distance intercellular relationships.     

The flux-force relations across complex membranes with active transport

Equations are derived for the total material flux, and the total electric current flux, across a complex membrane system with active transport. The equations describe the fluxes as linear functions of forces across the system, and specifically of electrical potential, hydrostatic pressure, chemical potentials, and active transport rates. The equations can be simplified for experimental studies by making one or more of the forces equal to zero. The osmotic pressure difference across a membrane system is shown to be a function of the electrical potential and chemical potential differences and of the active transport rates. The transmembrane potential is shown to be the sum of a diffusion potential and an active transport potential. A simple equation is derived describing the current across a membrane as a linear function of the electrical potential and the active transport rate. Specific examples of the application of the equations to nerve membrane potentials are considered.     

Magnesium-resistant excitatory synaptic potentials in the leech retzius cell

Postsynaptic potentials (PSPs) recorded from leech Retzius cells in response to stimulation of interganglionic connective could not be reversed by soma depolarization or abolished by 40 mM Mg ion, nor could input resistance changes be detected during them. Alteration of external Cl and K over a tenfold range provided no clear evidence that the PSPs involved a conductance change to either ion. The method of extrapolation yielded an apparent PSP equilibrium potential of about ?20 mV. The steep portion of the relationship between Retzius cell action potential amplitude and membrane potential extrapolated to an apparent reversal potential of ?13 mV. It is likely that the connective-to-Retzius cell PSPs were principally electrical events. Their apparent reversal potentials could have been in the range associated with chemical synapses because they traversed an electrical synapse with a variable coupling resistance, or because the polarizing currents, passing “backwards” across electrical synapses, changed the amplitude of the presynaptic action potentials.     

Temperature dependence of the Elovich equation derived from activated electron or ion transport across a biological interface,with an application to muscle spindle adaptation

The temperature dependence of the Elovich equation, which is−dx/dt=m exp (nx) (wherex is concentration of substrate or ion,t is time, andm andn are coefficients dependent upon temperature), has been derived from the hypothesis of electron or ion conduction across an activation energy barrier at the surface of a biological particle or membrane, driven by a difference in redox or ion potentials. Using the additional hypothesis of reversible, temperature-dependent, inactivation of sites of reduction or complexing, the theory predicts that both coefficientsm andn have linear Arrhenius plots, in agreement with experimental data for gas adsorption on inorganic solid surfaces, and with two studies of muscle spindle adaptation.     

Simultaneous Measurement of the Bioelectric Potential of the Cell Wall and the Vacuoles during the Oscillatory Response to the Nitella Cell

Simultaneous measurements of bioelectric potentials of the vacuole and cell wall in cells of Nitella mucronata were made by inserting glass microelectrodes into the vacuole and cell wall respeclively. During the oscillation of the bioelectric potential of the vacuole. induced by sudden changes of the external bathing solution or by the impalement of the cell with a microelectrode. the cell wall potential also exhibited fluctuations of variable intensities in phase and concomitant with spikes of the vacuolar potential oscillation. However, the polarity of the pulses of the cell wall potential was reverse to that of the spikes of the vacuolar potential. These results suggest that the same event is registered at both sides of the plasmalemma membrane across which these phenomena are occurring. The results also support the voltage clamp and tracer flux measurements on these cells which indicate that during the generation of single action potentials, induced by current, the plasma lemma transiently increases its permeability to Cl^? and K⁺ ions expelling them from the cell. The variable intensity of the transient hyperpolarizations of the cell wall potential is explained by the distance of the microelectrode in the cell wall from the plasmalemma.     

Current Oscillations Under Voltage-Clamp Conditions: An Interplay of Series Resistance and Negative Slope Conductance

Using the patch-clamp technique, we observed profound oscillations of the whole-vacuole outward current across the tonoplast of Mesembryanthemum crystallinum L. (common ice plant). These current oscillations showed a clear voltage dependence and appeared at membrane potentials more positive than 90–100 mV. This paper describes the oscillations in terms of two separate mechanisms. First, the Mesembryanthemum vacuolar membrane shows a negative slope conductance at membrane potentials more positive than 100–120 mV. The fact that the oscillations and the negative slope conductance show a similar threshold potential suggests that (part of) the same mechanism is involved in both phenomena. The second mechanism involved is the voltage drop across the series resistance. As a result, the potential actually experienced by the vacuolar membrane deviates from the command potential defined by the patch-clamp amplifier. This deviation depends in an Ohmic manner on the current magnitude. We suggest that the interplay of the negative slope conductance and the voltage drop across the series resistance can cause a positive feedback which is responsible for the current oscillations. Received: 30 April 1999/Revised: 9 September     

Transport phenomena in a model membrane accompanying a conformational change: Transient processes in response to external stimuli

Summary A model membrane composed of a filter paper and dioleylphosphate was studied by applying various kinds of external stimuli. When the concentration in the external solution was varied successively, the physico-chemical properties of the membrane changed drastically at a certain valueC _t . The relationships between the electrical response and the external stimuli studied are as follows: (1) The membrane potential oscillates spontaneously in a spikelike fashion when the concentration of the external solution is suddenly changed. (2) The current through the membrane oscillates in spikelike fashion for a duration of about 50 msec when the constant external voltage, V larger than a certain value V _c , is applied across the membrane. (3) The electric resistance sharply decreases, and a kind of action potential similar to that observed in living tissues is produced when a short rectangular electric stimulus, whose magnitude is higher than a critical value V _p , is applied. (4) If a hydrostatic pressure difference across the membrane is applied with appropriate salt conditions, the value of the membrane potential varies with time, as in the case of (3). The observed changes in emf and electric resistance are discussed in connection with the conformational change of DOPH molecules in the membrane.     

Potassium ion accumulation in a periaxonal space and its effect on the measurement of membrane potassium ion conductance

Summary Potassium currents of various durations were obtained from squid giant axons voltage-clamped in artificial seawater solutions containing sufficient tetrodotoxin to block the sodium conductance completely. From instantaneous potassium current-voltage relations, the reversal potentials immediately at the end of these currents were determined. On the basis of these reversal potential measurements, the potassium ion concentration gradient across the membrane was shown to decrease as the potassium current duration increased. The kinetics of this change was shown to vary monotonically with the potassium ion efflux across the membrane estimated from the integral over time of the potassium current divided by the Faraday, and to be independent of both the external sodium ion concentration and the presence or absence of membrane series resistance compensation. It was assumed that during outward potassium current flow, potassium ions accumulated in a periaxonal space bounded by the membrane and an external diffusion barrier. A model system was used to describe this accumulation as a continuous function of the membrane currents. On this basis, the mean periaxonal space thickness and the permeability of the external barrier to K⁺ were found to be 357 Å and 3.21×10^–4 cm/sec, respectively. In hyperosmotic seawater, the value of the space thickness increased significantly even though the potassium currents were not changed significantly. Values of the resistance in series with the membrane were calculated from the values of the permeability of the external barrier and these values were shown to be roughly equivalent to series resistance values determined by current clamp measurements. Membrane potassium ion conductances were determined as a function of time and voltage. When these were determined from data corrected for the potassium current reversal potential changes, larger maximal potassium conductances were obtained than were obtained using a constant reversal potential. In addition, the potassium conductance turn-on with time at a variety of membrane potentials was shown to be slower when potassium conductance values were obtained using a variable reversal potential than when using a constant reversal potential.     

Ion transport and displacement currents with membrane-bound carriers

Summary The standard carrier model for ion transport by a one-to-one mechanism is developed to predict the time-dependent currents for systems that are symmetrical at zero applied potential. The complete solution for ions and carriers bearing any charge is derived by assuming that the concentration of ions in the membrane is low and either that the applied potential is small or that the applied potential affects equally all of the association and dissociation reactions between the ions and the carriers. The response to an abruptly applied potential is then given by the sum of a constant and two declining exponential terms. The time constants of these relaxations are described by the equations derived for neutral carriers by Stark, Ketterer, Benz and Läuger in 1971 (Biophys. J. 11:981). The sum of the amplitudes of the exponentials for small applied potentials obeys a relation like that first derived by Markin and Liberman in 1973 (Biofizika 18:453). For small applied potentials expressions are also provided for the voltage transients in charge-pulse experiments and for the membrane admittance.     

Reaction of ascorbic acid with cytochrome b561. Concerted electron and proton transfer.

Rate constants for reduction of cytochrome b561 by internal ascorbate (k0A) and oxidation by external ferricyanide (k1F) were determined as a function of pH from rates of steady-state electron transfer across chromaffin-vesicle membranes. The pH dependence of electron transfer from cytochrome b561 to ferricyanide (k1F) may be attributed to the pH dependence of the membrane surface potential. The rate constant for reduction by internal ascorbate (k0A), like the previously measured rate constant for reduction by external ascorbate (k-1A), is not very pH-dependent and is not consistent with reduction of cytochrome b561 by the ascorbate dianion. The rate at which ascorbate reduces cytochrome b561 is orders of magnitude faster than the rate at which it reduces cytochrome c, despite the fact that midpoint reduction potentials favor reduction of cytochrome c. Moreover, the rate constant for oxidation of cytochrome b561 by ferricyanide (k1F) is smaller than the previously measured rate constant for oxidation by semidehydroascorbate, despite the fact that ferricyanide has a higher midpoint reduction potential. These results may be reconciled by a mechanism in which electron transfer between cytochrome b561 and ascorbate/semidehydroascorbate is accelerated by concerted transfer of a proton. This may be a general property of biologically significant electron transfer reactions of ascorbic acid.     

Tentacle regulating potentials inNoctiluca miliaris: their generation sites and ionic mechanisms

Summary Membrane potential responses that regulate movement of the food-gathering tentacle ofNoctiluca miliaris (tentacle regulating potentials, TRPs) were examined electrophysiologically under various ionic conditions. These spontaneous TRPs were modified by changing the external ionic conditions. Positive spike appeared as external Ca²⁺ concentration was lowered. The peak of the spike became more positive with increasing external Na⁺ concentration. The spike could be evoked by injecting a depolarizing current when the membrane was hyperpolarized. The positive spike is assumed to be caused by regenerative activation of depolarization-sensitive Na channels. The peak of the negative spike, reported by previous workers, became more negative with increasing external Cl^– concentration. The spike was evoked by injecting a hyperpolarizing current when the membrane was depolarized. The negative spike is assumed to be caused by regenerative activation of hyperpolarization-sensitive Cl^– channels. The waveforms and amplitudes of the TRPs recorded from the nucleus were identical to those recorded from the flotation vacuole. This suggests that the TRPs are generated on the membrane facing the external solution. Possible roles of the TRPs in the control of tentacle movement are discussed.Abbreviations ASW artificial sea water - FTP flash-triggering potential - TRP tentacle regulating potential     

Ion channels in the plasma membrane of protoplasts from the halophytic angiosperm Zostera muelleri

Patch clamp studies show that there may be as many as seven different channel types in the plasma membrane of protoplasts derived from young leaves of the halophytic angiosperm Zostera muelleri. In whole-cell preparations, both outward and inward rectifying currents that activate in a timeand voltage-dependent manner are observed as the membrane is either depolarized or hyperpolarized. Current voltage plots of the tail currents indicate that both currents are carried by K⁺. The channels responsible for the outward currents have a unit conductance of approximately 70 pS and are five times more permeable to K⁺ than to Na⁺. In outside-out patches we have identified a stretch-activated channel with a conductance of 100 pS and a channel that inwardly rectifies with a conductance of 6 pS. The reversal potentials of these channels indicate a significant permeability to K⁺. In addition, the plasma membrane contains a much larger K⁺ channel with a conductance of 300 pS. Single channel recordings also indicate the existence of two Cl^– channels, with conductances of 20 and 80 pS with distinct substates. The membrane potential difference of perfused protoplasts showed rapid action potentials of up to 50 mV from the resting level. The frequency of these action potentials increased as the external osmolarity was decreased. The action potentials disappeared with the addition of Gd³⁺, an effect that is reversible upon washout.We would like to thank K. Morris and D. McKenzie for technical assistance and the Australian Research Council for financial support.     

The membrane potential in everted vesicles of Escherichia coli.

Ion-selective electrodes were used to measure the equilibration of thiocyanate across the membrane of everted (“inside-out”) vesicles of Escherichia coli W1485. Membrane potentials, vesicle interior positive, generated by the oxidation of NADH, succinate, and d-lactate, or by the hydrolysis of ATP, fell in the range of 100–150 mV depending on the carbon source for cell growth and the substrate used to energize the membranes. There was no relationship between the rate of oxidation of different substrates and the membrane potential they generated. The membrane potential generated by oxidation of NADH was relatively constant between pH 7.0 and 8.5. Somewhat lower values obtained at pH 5.5 to 6.5 were attributed to the effect of pH on substrate oxidation.     

The oxidation of exogenous NADH by mitochondria of Euglena gracilis

A novel oxidase activity of external NADH was found in mitochondria of a streptomycin-bleached mutant and the wild strain of Euglena gracilis. In contrast to higher plants the oxidation of external NADH in mitochondria of E. gracilis is sensitive to rotenone and yields the same phosphorylation efficiency as the matrix pool of NADH. Simulation of this activity by the classic complex I of the matrix side of the mitochondrial membrane, as a result of preparation-generated artefacts, is excluded. The external NADH-dehydrogenase activity is bound to the inner mitochondrial membrane with its active side facing the cytosol. State-4 enzyme activity is only slightly influenced by pH in the physiological range, whereas state-3 oxidation indicates an optimum in the physiological pH, as expected from a limitation by the ATPase. The external redox potential of NADH does not control enzyme activity. The results are discussed with respect to the metabolic status of the cells at the time of harvesting.     

Mycoplasma membrane potentials determined by potential-sensitive fluorescent dyes

The membrane potentials of mycoplasmas were investigated by using potential-sensitive cyanine dyes. The fluorescence response results from a potential-dependent partition of the dyes between the cells and the extracellular medium. Cell hyperpolarization (inside more negative), e.g., by the addition of valinomycin, results in uptake of the dyes into the cells and, by formation of dye aggregates, in quenching of the fluorescence intensity. The magnitude of the fluorescence change upon addition of valinomycin depended on the external K⁺ concentration. At a defined external K⁺ concentration, no change in fluorescence occurred. The intracellular K⁺ concentration was determined by atomic absorption spectroscopy. Mycoplasma membrane potentials were calculated according to the Nernst equation. The membrane potential of bothMycoplasma mycoides subsp.capri andMycoplasma gallisepticum was −48 mV±10%; the membrane potential ofAcholeplasma laidlawii was −28 mV±20%.     

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 tetraphenylphosphonium 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.     

Osmotic regulation of assimilate unloading from seed coats of Vicia faba. Role of turgor and identification of turgor-dependent fluxes

Osmotic regulation of assimilate efflux from excised coats of developing Vicia faba (cv. Coles Prolific) seed was examined by exposing these to bathing solutions (adjusted to –0. 02 to –0. 75 MPa with sorbitol) introduced into the cavity vacated by the embryo. ¹⁴C photosynthate efflux was found to be independent of solution osmotic potentials below –0. 63 MPa. At higher osmotic potentials, efflux was stimulated and exhibited a biphasic response to osmotic potential with apparent saturation being reached at –0. 37 MPa. Efflux could be repeatedly stimulated and slowed by exposing seed coats to solutions of high and low osmotic potentials, respectively. Manipulation of components of tissue water potential, with slowly- and rapidly-permeating osmotica, demonstrated that turgor functioned as the signal regulating ¹⁴C photosynthate efflux. Com-partmental analysis of ¹⁴C photosynthate preloaded seed coats was consistent with exchange from 4 kinetically-distinct compartments. The kinetics of turgor-dependent efflux exhibited characteristics consistent with the transport mechanism residing in the plasma membranes of the unloading cells. These characteristics included the rapidity (<2 min) of the efflux response to turgor increases, similar rate constants for efflux from the putative turgor-sensitive and cytoplasmic compartments and the apparent small pool size from which turgor-dependent efflux could repeatedly occur. In contrast, influx of [¹⁴C] sucrose across the plasma and tonoplast membranes was found to be insensitive to turgor. The plasma membrane [¹⁴C] sucrose influx was unaffected by p-chloromercuribenzenesulfonic acid and erythrosin B and exhibited a linear dependence on the external sucrose concentration. This behaviour suggested that influx across the plasma membrane occurs by passive diffusion. Preloading excised seed coats with a range of solutes demonstrated that turgor-dependent efflux exhibited partial solute selectivity. Based on these findings, it is proposed that turgor controls assimilate exchange from the seed coat by regulating an efflux mechanism located in the plasma membranes of the unloading cells.     

Spike Potentials Recorded from the Insect Photoreceptor

Slow and spike potentials were recorded from single cells in the receptor layer of the compound eye of the drone of the honeybee. From electron microscopic observation of the drone ommatidium, it was concluded that the response had been recorded from the retinula cell. The following hypothesis is suggested for the initiation of spike potentials in the drone compound eye: Photic stimulation results in a decrease in the resistance of all or part of the retinula cell membrane, giving rise to the retinal action potential. The retinal action potential causes outflow of the current through the proximal process of the cell. This depolarizing current initiates spike potentials in the proximal process or axon of the retinula cell which are recorded across the soma membrane of the retinula cell.