Two-dimensional nanopores and nanoporous membranes for ion and molecule transport

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Electrostatic coupling across a membrane with titratable surface groups

The potential drop across a membrane is calculated for the case of ionizable groups on both membrane surfaces. The presence of both acid and amine groups on the membrane surfaces is considered. The membrane surface potential is obtained from the non-linear Poisson-Boltzmann equation by treating the fraction of dissociated ionizable surface groups as a self-consistent functional of the electrostatic potential. A discussion of the error due to ignoring the electrostatic coupling of the potential across the membrane is presented. The error turns out to be quite small for most membrane problems of biological interest. Finally, the conductance data of Mozhayeva & Naumov (1970) for the frog node are reanalyzed within the context of the diffuse double layer theory. It is shown to be unnecessary to invoke a specific binding of divalent cations to the membrane.     

Theoretical approach to the description of microvesicular transport in the endothelium

Theoretical consideration of mechanisms on curvature of the cell membrane, resulting in formation of microvesicles in the cells has been carried out. These mechanisms are presented as: 1) local changes of membrane surface tension at the border membranes--external medium; 2) local changes of the membrane potential; 3) hydrostatic pressure; 4) crystallization of membrane domains; 5) enzymatic changes of the lipid molecule configurations. Energy of the curved membrane and the energy of the external effects for the vesicle are estimated. Spontaneous formation of vesicles at the expense of the oscillatory mechanism is impossible. The most probable mechanism, from the energetic point of view, for vesicular formation is adsorption of surface-active substances or ions on the membrane. Thus, the processes of microvesicular mass transfer, playing an important role in transcapillary mass exchange are determined by physicochemical mechanisms. From the physico-chemical point of view adsorption of substances can be the most probable mechanism in the vesicular formation and determine intensity of vesicular transport.     

Ferrous ion transport across chloroplast inner envelope membranes

The initial rate of Fe(2+) movement across the inner envelope membrane of pea (Pisum sativum) chloroplasts was directly measured by stopped-flow spectrofluorometry using membrane vesicles loaded with the Fe(2+)-sensitive fluorophore, Phen Green SK. The rate of Fe(2+) transport was rapid, coming to equilibrium within 3s. The maximal rate and concentration dependence of Fe(2+) transport in predominantly right-side-out vesicles were nearly equivalent to those measured in largely inside-out vesicles. Fe(2+) transport was stimulated by an inwardly directed electrochemical proton gradient across right-side-out vesicles, an effect that was diminished by the addition of valinomycin in the presence of K(+). Fe(2+) transport was inhibited by Zn(2+), in a competitive manner, as well as by Cu(2+) and Mn(2+). These results indicate that inward-directed Fe(2+) transport across the chloroplast inner envelope occurs by a potential-stimulated uniport mechanism.     

Cell signaling and ion transport across the fish gill epithelium

A large array of circulating and local signaling agents modulate transport of ions across the gill epithelium of fishes by either affecting transport directly or by altering the size and distribution of transporting cells in the epithelium. In some cases, these transport effects are in addition to cardiovascular effects of the same agents, which may affect the perfusion pathways in the gill vasculature and, in turn, affect epithelial transport indirectly. Prolactin is generally considered to function in freshwater, because it is the only agent that allows survival of some hypophysectomized fish species in freshwater. It appears to function by either reducing branchial permeability, Na,K-activated ATPase activity, or reducing the density of chloride cells. Cortisol was initially considered to produce virtually opposite effects (e.g., stimulation of Na,K-activated ATPase and of chloride cell size and density), but more recent studies have found that this steroid stimulates ionic uptake in freshwater fishes, as well as the activity of H-ATPase, an enzyme thought to be central to ionic uptake. Thus, cortisol may function in both high and low salinities. Growth hormone and insulin-like growth factor appear to act synergistically to affect ion regulation in seawater fishes, stimulating both Na,K-activated ATPase and Na-K-2Cl co-transporter activity, and chloride cell size, independent of their effects on growth. Some of the effects of the GH-IGF axis may be via stimulation of the number of cortisol receptors. Thyroid hormones appear to affect seawater ion regulation indirectly, by stimulating the GH-IGF axis. Natriuretic peptides were initially thought to stimulate gill ionic extrusion, but recent studies have not corroborated this finding, so it appears that the major mode of action of these peptides may be reduction of salt loading by inhibition of oral ingestion and intestinal ionic uptake. Receptors for both arginine vasotocin and angiotensin have been described in the gill epithelium, but their respective roles and importance in fish ion regulation remains unknown. The gill epithelium may be affected by both circulating and local adrenergic agents, and a variety of studies have demonstrated that stimulation of alpha-adrenergic versus beta-adrenergic receptors produces inhibition or stimulation of active salt extrusion, respectively. Local effectors, such as prostaglandins, nitric oxide, and endothelin, may affect active salt extrusion as well as gill perfusion. Recent studies have suggested that the endothelin inhibition of salt extrusion is actually mediated by the release of both NO and prostaglandins. It is hoped that modern molecular techniques, combined with physiological measurements, will allow the dissection of the relative roles in ion transport across the fish gill epithelium of this surprisingly large array of putative signaling agents.     

Paths of ion transport across canine fetal tracheal epithelium

Fluid secretion by the fetal sheep lung is thought to be driven by secretion of Cl- by the pulmonary epithelium. We previously demonstrated Cl- secretion by tracheal epithelium excised from fetal dogs and sheep. In this study we characterized the ion transport pathways across fetal canine tracheal epithelium. The transport of Na+ and Cl- across trachea excised from fetal dogs was evaluated from transepithelial electrical properties and isotope fluxes. Under basal conditions the tissues were characterized by a lumen-negative potential difference (PD) of 11 mV and conductance of 5.2 mS/cm2. The short-circuit current (Isc) was 43 microA/cm2 (1.6 mueq.cm-2.h-1). Basal Na+ flows were symmetrical, but net Na+ absorption (1.1 mueq.cm-2.h-1) could be induced by exposure of the luminal surface to amphotericin B (10(-6) M). Bilateral replacement of Na+ reduced Isc by 85%. Replacement of submucosal Na+ or exposure to submucosal furosemide (10(-4) M) reduced net Cl- secretion by 60-70%. Luminal exposure to indomethacin (10(-6) M) induced a 50% decrease in Isc, whereas isoproterenol (10(-6) M) increased Isc by 120%. The properties of the Cl- secretory pathway across fetal dog trachea are consistent with the model proposed for Cl- secretion across adult dog trachea and other Cl- -secreting tissues (e.g., bullfrog cornea and shark rectal gland). The absence of basal Na+ absorption by fetal dog trachea probably reflects limited apical membrane Na+ permeability.     

Kinetics of macrotetrolide-induced ion transport across lipid bilayer membranes

Ion transport across lipid bilayer membranes in the presence of macrotetrolide antibiotics has been studied by stationary conductance and nonstationary relaxation methods. The results are discussed on the basis of a carrier model which has already been successfully applied to valinomycin induced ion transport. Again a kinetic analysis has been performed from which the single rate constants of the carrier model could be derived. In addition the equilibrium constant of complex formation in the aqueous phase could be determined. Measurements have been made for 4 macrotetrolides, for several ions and for various chain lengths of the lipids molecules composing the membrane.     

Bile salt alteration of ion transport across jejunal mucosa

The effect of conjugated dihydroxy and trihydroxy bile salts on electrolyte transport across isolated rabbit jejunal mucosa was studied. Both taurochenodeoxycholic acid and taurocholic acid increased the short-circuit current (I_sc) in bicarbonate-Ringer solution but not in a bicarbonate-free, chloride-free solution. Taurochenodeoxycholic acid was significantly more effective than taurocholic acid in increasing I_sc. The presence of theophylline prevented the taurochenodeoxycholic acid-and taurocholic acid-induced increase in I_sc. Transmural ion fluxes across jejunal mucosa demonstrated that 2 mM taurochenodeoxycholic acid decreased net Na⁺ absorption, increased net Cl⁻ secretion and increased the residual flux (which probably represents HCO₃⁻ secretion). These studies support the hypothesis that cyclic AMP may be a mediator of intestinal electrolyte secretion.     

A radioactive uptake assay to measure ion transport across ion channel-containing liposomes

Here we describe a procedure for incorporating ion channels into lipid vesicles (liposomes) and functional characterization of the channel population by assaying radioactive isotope uptake into these proteoliposomes. The technique as described will work only for potassium channels but can be easily modified, as suggested in the text, for other ion channels and transporters. Purified ion channel proteins in detergent micelles are combined with solubilized lipids. Detergent is subsequently removed from protein-lipid complexes by gel filtration or dialysis into high potassium (high [K+]) buffer. After freezing-thawing and sonication, the resultant larger liposomes are passed over another gel-filtration column to exchange an extraliposomal high [K+] to a low [K+] buffer, thus establishing a large K+ gradient across the liposomal membrane. Trace 86Rb is then added to the extraliposomal space and the reaction begins. If the ion channel is permeable to K+, the K+ inside exits the liposomes down its concentration gradient and the 86Rb outside accumulates in the intraliposomal space until equilibrium is reached. The reaction time course is monitored by measurement of accumulated 86Rb after removal of external 86Rb over an ion-exchange column. The 86Rb flux assay takes 2-5 hours depending on the reaction rate and the number of desired time points.     

Shear stress-facilitated calcium ion transport across lipid bilayers.

Small unilamellar liposomes were used in this study of shear stress effects on the trans-bilayer flux of calcium ions (Ca2+). Liposome suspensions were prepared from 99% egg phosphatidylcholine by a microporous filter extrusion technique. The inner aqueous phase of the unilamellar liposomes contained indo-1(5-), a fluorescent indicator of free Ca2+. The external aqueous phase was composed of Hepes-buffered saline containing normal physiological levels of common ionic species. Calcium ion levels were set at 100 nM and 1 mM in the inner and outer aqueous phases, respectively. Liposome suspensions were exposed to graded levels of uniform shear stress in an optically modified rotational viscometer. Intraliposome Ca2+ concentration was estimated from continuous measurement of indo-1(5-) fluorescence. Electronically measured particle size distribution was used to determine liposome surface area for estimation of trans-bilayer Ca2+ flux. Trans-bilayer Ca2+ flux increased linearly with applied shear rate from 27 s-1 to 2700 s-1. Diffusional resistance of the lipid bilayer, not the convective resistance of the surrounding fluid, was the limiting step in the transport of Ca2+. Liposome permeability to Ca2+ increased by nearly two orders of magnitude over the physiologically relevant shear rate range studied. Solute transport in injectable liposome preparations may be dramatically influenced by cardiovascular fluid stress. Solute delivery rates determined in liposomes exposed to static conditions may not accurately predict in vivo, cardiovascular solute transport.     

Phloretin-induced changes in ion transport across lipid bilayer membranes

Phloretin, the aglucone derivative of phlorizin, increases cation conductance and decreases anion conductance in lipid bilayer membranes. In this paper we present evidence that phloretin acts almost exclusively by altering the permeability of the membrane interior and not by modifying the partition of the permanent species between the membrane and the bulk aqueous phases. We base our conclusion on an analysis of the current responses to a senylborate, and the cation complex, peptide PV-K+. These results are consistent with the hypothesis that phloretin decreases the intrinsic positive internal membrane potential but does not modify to a great extent the potential energy minima at the membrane interfaces. Phloretin increases the conductance for the nonactin-K+ complex, but above 10(-5) M the steady- state nonactin-K+ voltage-current curve changes from superlinear to sublinear. These results imply that, above 10(-5) M phloretin, the nonactin-5+ transport across the membrane becomes interfacially limited.     

Theoretical investigation of the behavior of titratable groups in proteins.

This paper presents a theoretical analysis of the titration behavior of strongly interacting titratable residues in proteins. Strongly interacting titratable residues exist in many proteins such as for instance bacteriorhodopsin, cytochrome c oxidase, cytochrome bc(1), or the photosynthetic reaction center. Strong interaction between titratable groups can lead to irregular titration behavior. We analyze under which circumstances titration curves can become irregular. We demonstrate that conformational flexibility alone can not lead to irregular titration behavior. Strong interaction between titratable groups is a necessary, but not sufficient condition for irregular titration curves. In addition, the two interacting groups also need to titrate in the same pH-range. These two conditions together lead to irregular titration curves. The mutation of a single residue within a cluster of interacting titratable residues can influence the titration behavior of the other titratable residues in the cluster. We demonstrate this effect on a cluster of four interacting residues. This example underlines that mutational studies directed at identifying the role of a certain titratable residue in a cluster of interacting residues should always be accompanied by an analysis of the effect of the mutation on the titration behavior of the other residues.     

A theory of ion transport across cell surfaces by a process analogous to electron transport across liquid-solid interfaces

A kinetic theory of ion transport across cell surfaces has been developed in a form analogous to the kinetic theory of electron transport across solid-liquid interfaces of biological particles. The ionic theory is based on the observation that, at least in one instance, the voltage-current behavior for ion conduction across a cell surface is describable by the Tafel equation, in analogy to the conduction of electrons across solid-liquid interfaces. The theory predicts that the kinetics of ion transport across cell surfaces should conform to the Elovich rate equation, which is shown to be true for various experimental data. The opinions and conclusions contained in this report are those of the author. They are not to be construed as necessarily reflecting the views or the endorsement of the Navy Department.     

Bile salt alteration of ion transport across jejunal mucosa.

The effect of conjugated dihydroxy and trihydroxy bile salts on electrolyte transport across isolated rabbit jejunal mucosa was studied. Both taurochenodeoxycholic acid and taurocholic acid increased the short-circuit current (Isc) in bicarbonate-Ringer solution but not in a bicarbonate-free, chloride-free solution. Taurochenodeoxycholic acid was significantly more effective than taurocholic acid in increasing Isc. The presence of theophylline prevented the taurochenodeoxycholic acid- and taurocholic acid-induced increase in Isc. Transmural ion fluxes across jejunal mucosa demonstrated that 2 mM taurochenodeoxycholic acid decreased net Na+ absorption, increased net Cl- secretion and increased the residual flux (which probably represents HCO3- secretion). These studies support the hypothesis that cyclic AMP may be a mediator of intestinal electrolyte secretion.     

Regulation of ion transport across the pollen tube plasmalemma by hydrogen peroxide

The polar growth of the pollen tube is a key stage in the life cycle of seed plants, which is critical for successful sexual reproduction. One of the most important components of this process is ion transport across the cell membrane coordinated in time and space. Different classes of signal molecules, including reactive oxygen species, as has been found recently, participate in regulation of ion transmembrane transport. In this study, based on the model system of subprotoplasts isolated from pollen tubes, we showed that hydrogen peroxide can regulate two targets located on the plasma membrane: nifedipine-sensitive Ca²⁺ channels and ion transport, both of which control the membrane potential. The interaction of hydrogen peroxide with these targets resulted in an increase in an intracellular Ca²⁺ concentration and hyperpolarization of the plasma membrane. Faster regeneration of the cell wall was a consequence of elevation of the Ca²⁺ intracellular concentration.     

Millimeter microwave effect on ion transport across lipid bilayer membranes

The effects of millimeter microwaves in the frequency range of 54–76 GHz on capacitance and conductance of lipid bilayer membranes (BLM) were studied. Some of the membranes were modified by gramicidin A and amphotericin B or by tetraphenylboron anions (TPhB_?). The millimeter microwaves were pulse-modulated (PW) at repetition rates ranging from 1 to 100 pps, PW at 1000 pps, or unmodulated continuous waves (CW). The maximum output power at the waveguide outlet was 20 mW. It was found that CW irradiation decreased the unmodified BLM capacitance by 1.2% ± 0.5%. At the same time, membrane current induced by TPhB transport increased by 5% ± 1%. The changes in conductance of ionic channels formed by gramicidin A and amphotericin B were small (0.6% ± 0.4%). No “resonance-like” effects of mm-wave irradiation on membrane capacitance, ionic channel currents, or TPhB transport were detected. All changes in membrane capacitance and currents were independent of the modulation employed and were equivalent to heating by approximately 1.1 °C. © 1995 Wiley-Liss, Inc.