Non-mediated zero voltage conductance of hydrophobic ions through bilayer lipid membranes.

A charge pulse technique applied to the study of charge transfer at metal-solution interfaces has been used to determine the capacity and the conductance of a membrane bilayer at both zero time and zero voltage. The transport of hydrophobic ions across a glycerol-monooleate bilayer (tetraphenyl borate, picrate, dipicrylamine and tetraphenyl arsonium) has been investigated by this method. A theoretical approach to the problem has been proposed based on one analogous to that used for the compact double layer at metallic electrodes.     

Electric field-driven transformations of a supported model biological membrane--an electrochemical and neutron reflectivity study

A mixed bilayer of cholesterol and dimyristoylphosphatidylcholine has been formed on a gold-coated block of quartz by fusion of small unilamellar vesicles. The formation of this bilayer lipid membrane on a conductive surface allowed us to study the influence of the support's surface charge on the structure and hydration of the bilayer lipid membrane. We have employed electrochemical measurements and the specular reflection of neutrons to measure the thickness and water content in the bilayer lipid membrane as a function of the charge on the support's surface. When the surface charge density is close to zero, the lipid vesicles fuse directly on the surface to form a bilayer with a small number of defects and hence small water content. When the support's surface is negatively charged the film swells and incorporates water. When the charge density is more negative than −8 μC cm⁻², the bilayer starts to detach from the metal surface. However, it remains in a close proximity to the metal electrode, being suspended on a thin cushion of the electrolyte. The field-driven transformations of the bilayer lead to significant changes in the film thicknesses. At charge densities more negative than −20 μC cm⁻², the bilayer is ~37 Å thick and this number is comparable to the thickness determined for hydrated multilayers of dimyristoylphosphatidylcholine from x-ray diffraction experiments. The thickness of the bilayer decreases at smaller charge densities to become equal to ~26 Å at zero charge. This result indicates that the tilt of the acyl chains with respect to the bilayer normal changes from ~35° to 59° by moving from high negative charges (and potentials) to zero charge on the metal.     

Generation of capacitance current harmonics during electrostriction of inhomogeneous bilayers

The mechanisms of generation of capacitance current harmonics arising from bilayer electrostriction are investigated theoretically in the case when fixed charges or dipoles are located inside the bilayer. If the elastic properties of the bilayer are inhomogeneous in a transversal direction, these charges or dipoles affect substantially the amplitude A2 of the second current harmonic and accordingly the magnitude of compensatory constant voltage VC, which shold be applied to the membrane to reduce A2 to zero. In membranes whose Young's modulus is minimum at the center of bilayer, the dependence of VC on depth of immersion of the charge xA into the bilayer represents a function with alternating signs, which is reduced to zero at the center of bilayer. In the case of a dipole source immersed into the bilayer, the appropriate dependence is oscillating as well, and its shape is a derivative of the corresponding dependence of VC for fixed charges with respect to a coordinate. Possible applications of the results are discussed.     

Non-isothermal potential of phospholipid bilayer films

The thermal electrical potential generated in isochemical conditions by a temperature gradient at the two sides of lipid bilayer leaflets is measured. The experimental results agree rather well with the theoretical predictions. The cephaline (from sheep brain) bilayer behaves like a film with zero charge while the phosphatidylcholine (from egg yolk) film performs like a charged membrane. The results presented suggest that the measurement of electrical thermal potential is an interesting method to investigate the electrical behaviour of bilayer membranes.     

Soluble amyloid oligomers increase bilayer conductance by altering dielectric structure

The amyloid hypothesis of Alzheimer's toxicity has undergone a resurgence with increasing evidence that it is not amyloid fibrils but a smaller oligomeric species that produces the deleterious results. In this paper we address the mechanism of this toxicity. Only oligomers increase the conductance of lipid bilayers and patch-clamped mammalian cells, producing almost identical current-voltage curves in both preparations. Oligomers increase the conductance of the bare bilayer, the cation conductance induced by nonactin, and the anion conductance induced by tetraphenyl borate. Negative charge reduces the sensitivity of the membrane to amyloid, but cholesterol has little effect. In contrast, the area compressibility of the lipid has a very large effect. Membranes with a large area compressibility modulus are almost insensitive to amyloid oligomers, but membranes formed from soft, highly compressible lipids are highly susceptible to amyloid oligomer-induced conductance changes. Furthermore, membranes formed using the solvent decane (instead of squalane) are completely insensitive to the presence of oligomers. One simple explanation for these effects on bilayer conductance is that amyloid oligomers increase the area per molecule of the membrane-forming lipids, thus thinning the membrane, lowering the dielectric barrier, and increasing the conductance of any mechanism sensitive to the dielectric barrier.     

Photogating of ionic currents across lipid bilayers. Hydrophobic ion conductance by an ion chain mechanism.

The photogating of hydrophobic ion currents across the lipid bilayer membrane allows the direct study of their kinetics by symmetrically forming charge within the membrane and across each interface, rather than across the membrane. We find that the photoinduced conductance continues to increase beyond the region where the tetraphenylboride charge density in the membrane exceeds the estimated porphyrin cation density. This photoconductance is proportional to the tetraphenylboride charge density raised to the second to third power. The risetime of the photogating effect increases with increasing concentration of tetraphenyl boride. The porphyrin cation mobility is increased when the tetraphenylboride anion is present, and low concentrations of tetraphenylphosphonium cation increase the dark conductivity while inhibiting the photoconductivity. The activation energy for both the porphyrin and phosphonium cation induced conductance is more positive than that of the tetraphenylboride conductance. From these results we conclude that in addition to some cancellation of space charge within the membrane, the mechanism of increased conductance involves the transport of these hydrophobic anions via an alternating anion-cation chain, analogous to the Grotthuss mechanism for excess proton conduction in water. This ion chain conductance can be viewed as an evolutionary prototype of an ion channel across the membrane. It also underscores the importance of the counter ion in the transport of large ions such as peptides across the lipid bilayer.     

Photogating of ionic currents across lipid bilayers. Electrostatics of ions and dipoles inside the membrane.

The conductances of the lipophilic ions tetraphenylboride and tetraphenylphosphonium across a lipid bilayer can be increased or decreased, i.e., gated, by the photoformation of closed-shell metalloporphyrin cations within the bilayer. The gating can be effected by pulsed or continuous light or by chemical oxidants. At high concentrations of lipophilic anions where the dark conductance is saturated due to space charge in the bilayer, the photogated conductance can increase 15-fold. The formation of porphyrin cations allows the conductance to increase to its nonspace charge limited value. Conversely, the decrease of conductance in the light of phosphonium cations diminishes toward zero as the dark conductance becomes space charge limited. We present electrostatic models of the space charge limited conductance that accurately fit the data. One model includes an exponentially varying dielectric constant for the polar regions of the bilayer that allows an analytical solution to the electrostatic problem. The exponential variation of the dielectric constant effectively screens the potential and implies that the inside and outside of real dielectric interfaces can be electrically isolated from one another. The charge density, the distance into the membrane of the ions, about one-quarter of its thickness, and the dielectric constant at that position are determined by these models. These calculations indicate that there is insufficient porphyrin charge density to cancel the boride ion space charge and the following article proposes a novel ion chain mechanism to explain these effects. These models indicate that the positive potential arising from oriented carbonyl ester groups, previously used to explain the 10(3)-fold larger conductance of hydrophobic anions over cations, is smaller than previously estimated. However, the synergistic movement of the positive choline group into the membrane can account for the large positive potential.     

Interaction of cationic bilayer fragments with a model oligonucleotide

The interaction between cationic bilayer fragments and a model oligonucleotide was investigated by differential scanning calorimetry, turbidimetry, determination of excimer to monomer ratio of 2-(10-(1-pyrene)-decanoyl)-phosphatidyl-choline in bilayer fragment dispersions and dynamic light scattering for sizing and zeta-potential analysis. Salt (Na?HPO?), mononucleotide (2'-deoxyadenosine-5'-monophosphate) or poly (dA) oligonucleotide (3'-AAA AAA AAA A-5') affected structure and stability of dioctadecyldimethylammonium bromide bilayer fragments. Oligonucleotide and salt increased bilayer packing due to bilayer fragment fusion. Mononucleotide did not reduce colloid stability or did not cause bilayer fragment fusion. Charge neutralization of bilayer fragments by poly (dA) at 1:10 poly (dA):dioctadecyldimethylammonium bromide molar ratio caused extensive aggregation, maximal size and zero of zeta-potential for the assemblies. Above charge neutralization, assemblies recovered colloid stability due to charge overcompensation. For bilayer fragments/poly (dA), the nonmonotonic behavior of colloid stability as a function of poly (dA) concentration was unique for the oligonucleotide and was not observed for Na?HPO? or 2'-deoxyadenosine-5'-monophosphate. For the first time, such interactions between cationic bilayer fragments and mono- or oligonucleotide were described in the literature. Bilayer fragments/oligonucleotide assemblies may find interesting applications in drug delivery.     

Charge pulse studies of transport phenomena in bilayer membranes

The Journal of Membrane Biology - A charge pulse technique has been applied to studies of transport phenomena in bilayer membranes. The membrane capacitance can be rapidly charged (in less than a...     

AFM study on the electric-field effects on supported bilayer lipid membranes

Electric-field induced changes in structure and conductivity of supported bilayer lipid membranes (SLM) have been studied at submicroscopic resolution using atomic force microscopy and electrochemical impedance spectroscopy. The SLMs are formed on gold surfaces modified with mixed self-assembled monolayers of a cholesterol-tether and 6-mercaptohexanol. At applied potentials of ≤−0.25 V versus standard hydrogen electrode, the conductance of the SLM increases and membrane areas of <150 nm in size are found to elevate from the surface up to 15 nm in height. To estimate the electric field experienced by the lipid membrane, electrowetting has been used to determine the point of zero charge of a 6-mercaptohexanol-modified surface (0.19 ± 0.13 V versus standard hydrogen electrode). The effects of electric fields on the structure and conductance of supported membranes are discussed.     

The effect of a myelin apoprotein on lipid structure: a spin probe study.

Spin probes, stable free radical derivatives of stearic acid and cholestanone, were used to observe the effects of the "Folch-Lees" protein isolated from the white matter of bovine brain on the organization and motion of lipid molecules. The incorporation of the organic solvent soluble form of this protein decreased the tendency of a variety of lipid molecules with zero, positive or negative net charges to arrange themselves close to the normal to the lipid bilayer. The aqueous form of the protein also had a profound chaotropic effect on the molecular geometry of the lipid, but only if the lipids had a net negative charge (the protein has a net positive charge in the pH range investigated). Examination of the ESR spectra indicated that this protein altered the geometry of the lipid structure without causing major changes in the mobility of the individual lipid molecules.     

Antagonism of membrane compression effects by high pressure gas mixtures in a phospholipid bilayer system

Binary mixtures of helium with nitrogen, xenon or nitrous oxide were applied to suspensions of phosphatidylcholine-cholesterol vesicles to determine those mixtures of lipid soluble gases which would exactly antagonize the membrane rigidifying effect of 100 ATA compression. A previous study has shown that the initial application of 100 ATA compression by gas produces a significant reduction in the fluidity of the phospholipid bilayer. However, as the high pressure gas dissolves into the lipid region it creates disorder and increases fluidity. Fluidity of the bilayer at equilibrium represents the sum of the compression-ordering and dissolved-gas disordering effects and is dependent on the gas/lipid partition coefficient of the particular gas. The beneficial effect of a narcotic gas added to Trimix mixtures to ameliorate HPNS in deep divers may be due to a balance of compression-ordering and solubility-disordering effects achieved within the nerve membrane. It is therefore valuable to determine those gas mixtures which achieve balance of these two effects and result in zero net change in phospholipid bilayer fluidity at an established pressure of 100 ATA. Binary mixtures of helium with 88% nitrogen, 3.8% xenon or 2.8% nitrous oxide resulted in zero net change in bilayer fluidity with our model system at 100 ATA. A graph of the percent of narcotic gas needed to produce zero net effect as a function of pressure, however, was nonlinear. This would suggest the ratio of gases in Trimix must be varied as a function of pressure. While the phosphatidylcholine-cholesterol bilayer is a good model for certain components of the nerve membrane, it does not allow for study of protein-lipid or gas-protein interactions. The data presented thus aid in our understanding of HPNS but are yet incomplete for precise use in predicting diving mixtures.     

Nucleotide conformational change induced by cationic bilayers

The effects of cetyltrimethylammonium bromide (CTAB) micelles and dioctadecyldimethylammonium bromide (DODAB) bilayers on molecular conformation of 2'-deoxyadenosine 5'-monophosphate (dAMP) were evaluated from circular dichroism spectroscopy (CD) and molecular modeling of dAMP conformations of minimal energy upon varying torsion angles for the glycosidic bond (t(1)) for four different conditions of dielectric constant of the medium (E) and negative charge on the phosphate moiety (C), namely, E80_C2, E80_C0, E1_C2, and E1_C0. Upon decreasing medium polarity, a decreased intensity of the negative band over the 190-210 nm region for the dAMP CD spectrum was observed. Upon increasing relative proportion dAMP: DODAB, an increased intensity of the positive band over the 210-230 nm region plus a red shift were obtained that could be attributed to an increased nitrogenous base stacking, similar to A stacking in poly(A). Concomitant base stacking and insertion in the cationic aggregates were observed for DODAB bilayers but not for CTAB micelles. Thereby, the nucleotide extended, anti conformation in pure water typical for nucleotides in DNA was forced by the cationic bilayer to become syn. dAMP conformational modeling upon simultaneous changes in the nucleotide environment (from water to a hydrocarbon phase) and in the charge on phosphate moiety (-2 to zero) allowed to simulate dAMP conformation in the cationic bilayer/dAMP complex. Modeling confirmed the dAMP anti-to-syn conformational change experimentally characterized from CD spectroscopy. This nucleotide conformational change would possibly be at the root of DNA denaturation upon complexation with cationic lipids.     

Repulsive interactions between uncharged bilayers. Hydration and fluctuation pressures for monoglycerides.

Pressure versus distance relations have been obtained for solid (gel) and neat (liquid-crystalline) phase uncharged lipid bilayers by the use of x-ray diffraction analysis of osmotically stressed monoglyceride aqueous dispersions and multilayers. For solid phase monoelaidin bilayers, the interbilayer repulsive pressure decays exponentially from a bilayer separation of approximately 7 A at an applied pressure of 3 x 10(7) dyn/cm2 to a separation of approximately 11 A at zero applied pressure, where an excess water phase forms. The decay length is approximately 1.3 A, which is similar to the value previously measured for gel phase phosphatidylcholine bilayers. This implies that the decay length of the hydration pressure does not depend critically on the presence of zwitterionic head groups in the bilayer surface. For liquid-crystalline monocaprylin, the repulsive pressure versus distance curve has two distinct regions. In the first region, for bilayer separations of approximately 3-8 A and applied pressures of 3 x 10(8) to 4 x 10(6) dyn/cm2, the pressure decays exponentially with a decay length of approximately 1.3 A. In the second region, for bilayer separations of approximately 8-22 A and applied pressures of 4 x 10(6) to 1 x 10(5) dyn/cm2, the pressure decays much more gradually and is inversely proportional to the cube of the distance between bilayers. These data imply that two repulsive pressures operate between liquid-crystalline monocaprylin bilayers, the hydration pressure, which dominates at small (3-8 A) bilayer separations, and the fluctuation pressure, which dominates at larger bilayer separations (greater than 8 A) and strongly influences the hydration properties of the liquid-crystalline bilayers.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hexane dissolved in dioleoyllecithin bilayers has a partial molar volume of approximately zero

Neutron diffraction has been used to measure the amount and distribution of hexane incorporated from the vapor phase into oriented dioleoylphosphatidylcholine bilayers at 66% relative humidity. We reported earlier that hexane at low concentrations is located largely in a zone 10 A wide at the center of the bilayer [White, S. H., King, G. I., & Cain, J. E. (1981) Nature (London) 290, 161-163]. Extending these studies to high hexane concentrations, we find no readily apparent change in the volume of the hydrocarbon region of the bilayer even though more than one hexane molecule per lipid enters the region. The hexane partial molar volume in the bilayer hydrocarbon region is thus approximately zero. Within our statistical confidence limits, the partial molar volume is certainly no greater than one-third the molecular volume of the hexane. Further, analysis of the data suggests that the mass density of the bilayer is considerably less than 1 in the absence of hexane. These findings are in conflict with the assumptions usually made about lipid bilayers and their interaction with nonpolar hydrophobic molecules. In the course of these experiments, we found that standard methods of interpreting diffraction results were not suitable for our purposes. We thus developed several new methods which are summarized in the text and two appendixes. One of these methods allows us to define with precision the width of the hydrocarbon core of the bilayer. The other provides a means of calculating the effects of changes in the absolute scaling of the bilayers with changes in composition without placing the structures on an absolute scattering length density scale.     

Characterization of the interaction of phospholipase A(2) with phosphatidylcholine-phosphatidylglycerol mixed lipids

The first requirement in the hydrolysis of phospholipid bilayers by phospholipase A(2) is the interaction of the enzyme with the bilayer surface. The catalytic ability of phospholipase A(2) has been shown to be extremely sensitive to the topology of the bilayer to which it binds and hydrolyzes. Phospholipid bilayer properties and composition such as unsaturation, charge, and the presence of reaction products are known regulators of the catalytic activity of phospholipase A(2) toward the phospholipids and influences the binding of enzyme to the membrane. We show in this paper that the effect of increased anionic lipid results in enhanced binding that can be described quantitatively in terms of a simple phenomenological model. However, the interaction with anionic lipid does not singularly dominate the thermodynamics of binding, nor can the lag phase observed in the time course of hydrolysis of large unilamellar vesicles simply be the result of limited interaction between the enzyme and the bilayer. Furthermore, we show that phospholipase A(2) from Akgistrodon piscivorus piscivorus can exist in at least two bilayer-bound states and that the absence of a fluorescence change upon mixing the enzyme with lipid bilayers does not necessarily indicate the absence of an interaction.     

Surface potential determination in planar lipid bilayers: a simplification of the conductance-ratio method

One of the methods available for the measurement of surface potentials of planar lipid bilayers uses the conductance ratio between a charged and a neutral bilayer doped with ionophores to calculate the surface potential of the charged bilayer. We have devised a simplification of that method which does not require the use of an electrically neutral bilayer as control. The conductance of the charged bilayer is measured before and after the addition of divalent cations (Ba(2+)) to the bathing solution. Ba(2+) ions screen fixed surface charges, decreasing the surface potential. If the membrane is negatively charged the screening has the effect of decreasing the membrane conductance to cations. The resulting conductance ratio is used to calculate the surface potential change, which is fed into an iterative computer program. The program generates pairs of surface potential values and calculates the surface charge density for the two conditions. Since the surface charge density remains constant during this procedure, there is only one pair of surface potentials that satisfies the condition of constant charge density. Applying this method to experimental data from McLaughlin et al. [McLaughlin, S.G.A., Szabo, G. and Eisenman, G., Divalent ions and the surface potential of charged phospholipid membranes, J. Gen. Physiol., 58 (1971) 667-687.] we have found very similar results. We have also successfully used this method to determine the effect of palmitic acid on the surface potential of asolectin membranes.     

On the theory of action potential propagation in plant cells

The distribution of an electric field in plant cells and zooblasts has been investigated during propagation of the action potential. The behavior of ions in the cytoplasm and in the extracellular fluid has been described with the equations of electric charge motion in electrolytes. It has been shown that the action potential causes an electric potential change not only in the depth of the cytoplasm but also in the extracellular area far from the lipid bilayer. The biomembrane resistance has been expressed by physical parameters of a cell, such as ionic diffusion coefficient in fluid, Debye-Hückel radius, dielectric constant etc. The presence of breaks in the action potential diagrams has been explained as a result of insufficient resolving power of the measuring devices at the instant the sodium ion channels of the bilayer open.     

Interaction of charged lipid vesicles with planar bilayer lipid membranes: Detection by antibiotic membrane probes

A technique has been developed for monitoring the interaction of charged phospholipid vesicles with planar bilayer lipid membranes (BLM) by use of the antibiotics Valinomycin, Nonactin, and Monazomycin as surface-charge probes. Anionic phosphatidylserine vesicles, when added to one aqueous compartment of a BLM, are shown to impart negative surface charge to zwitterionic phosphatidylocholine and phosphatidylethanolamine bilayers. The surface charge is distributed asymmertically, mainly on the vesicular side of the BLM, and is not removed by exchange of the vesicular aqueous solution. Possible mechanisms for the vesicle-BLM interactions are discussed.