Ion Selectivity Sequence Research Articles

A Comprehensive Study on the Ion-Selective Behavior of MnOx for Electrochemical Deionization

Manganese oxide is an effective active material in several electrochemical systems including batteries, supercapacitors, and electrochemical deionization (ECDI). This work conducts a comprehensive study on the ion-selective behavior of MnOx to fulfill the emptiness in the energy and environmental science field. Furthermore, it broadens the promising application of MnOx in the ion-selective ECDI system. With the support of a microfluidic cell and the electrochemical quartz crystal microbalance (EQCM) analysis, we proposed a time-dependent multi-mechanism ion-selective behavior with the following guidelines. (1) Hydrated radius is the most critical factor for ions with the same valence, and MnOx tends to capture cations with a small hydrated radius. (2) The importance of charge density rises when comparing cations with different valences, and MnOx prefers to capture divalent cations with a large hydrated radius but a strong electrostatic attraction. In addition, ion-swapping may occur where divalent cations replace monovalent cations in this circumstance. (3) NH4 + triggers MnOx dissolution, leading to performance and stability decay. The clear EQCM evidence has directly verified the proposed mechanisms. Moreover, the EQCM data provide a novel but simple method to judge ion selectivity preference. The overall ion selectivity sequence is Ca2+ > Mg2+ > K+ > NH4 + > Na+ > Li+ with the highest βCa//Li and βCa//Na values around 3 at the deionization time = 10 min.

Comprehensive Study on the Ion-Selective Behavior of MnOx for Electrochemical Deionization.

Manganese oxide is an effective active material in several electrochemical systems, including batteries, supercapacitors, and electrochemical deionization (ECDI). This work conducts a comprehensive study on the ion-selective behavior of MnOx to fulfill the emptiness in the energy and environmental science field. Furthermore, it broadens the promising application of MnOx in the ion-selective ECDI system. We propose a time-dependent multimechanism ion-selective behavior with the following guidelines by utilizing a microfluidic cell and the electrochemical quartz crystal microbalance (EQCM) analysis. (1) Hydrated radius is the most critical factor for ions with the same valence, and MnOx tends to capture cations with a small hydrated radius. (2) The importance of charge density rises when comparing cations with different valences, and MnOx prefers to capture divalent cations with a strong electrostatic attraction at prolonged times. Under this circumstance, ion swapping may occur where divalent cations replace monovalent cations. (3) NH4+ triggers MnOx dissolution, leading to performance and stability decay. The EQCM evidence has directly verified the proposed mechanisms, and these data provide a novel but simple method to judge ion selectivity preference. The overall ion selectivity sequence is Ca2+ > Mg2+ > K+ > NH4+> Na+ > Li+ with the highest selectivity values of βCa//Li and βCa//Na around 3 at the deionization time = 10 min.

Osmotically Activated Anion Current of Phycomyces Blakesleeanus-Filamentous Fungi Counterpart to Vertebrate Volume Regulated Anion Current.

Studies of ion currents in filamentous fungi are a prerequisite for forming a complete understanding of their physiology. Cytoplasmic droplets (CDs), obtained from sporangiophores of Phycomyces blakesleeanus, are a model system that enables the characterization of ion currents in the native membrane, including the currents mediated by the channels not yet molecularly identified. Osmotically activated anionic current with outward rectification (ORIC) is a dominant current in the membrane of cytoplasmic droplets under the conditions of hypoosmotic stimulation. We have previously reported remarkable functional similarities of ORIC with the vertebrate volume regulated anionic current (VRAC), such as dose-dependent activation by osmotic difference, ion selectivity sequence, and time and voltage dependent profile of the current. Using the patch clamp method on the CD membrane, we further resolve VRAC-like ORIC characteristics in this paper. We examine the inhibition by extracellular ATP and carbenoxolone, the permeation of glutamate in presence of chloride, selectivity for nitrates, and activation by GTP, and we show its single channel behavior in excised membrane. We propose that ORIC is a functional counterpart of vertebrate VRAC in filamentous fungi, possibly with a similar essential role in anion efflux during cell volume regulation.

D433 Does Not Determine Ion Selectivity in ASIC1

ASICs (Acid Sensing Ion Channels) are ion channels activated by external protons. They belong to the family of ion channels known as ENaC/Degenerin, which has a distinctive high selectivity for Na+ ions. The atomic structure of chicken ASIC1 solved by Gonzales (Gonzales et al., Nature 460: 599604, 2009) shows the transmembrane segments, TM1 and TM2, of three subunits around the pore. The TM2s are tilted ∼50 degrees to the normal of the membrane producing a constriction that closes the lumen of the pore near the external side of the ion pathway. The side chain of D433 in TM2 defines the narrowest segment of the pore in the desensitized state and the carboxyl group coordinates a Cs+ ion, suggesting that D433 forms both the desensitization gate and the selectivity filter. We show here that most substitutions of residue D433 reduce or abolish ASIC1 currents and change the kinetics of desensitization. The polar residues Q and N decrease the mean duration of open events to a few milliseconds but maintain the ion selectivity sequence: Na+>Li+>K+>>>Cs+, and the value of the unitary conductance: 17 pS (120 mM symmetric Na+, V range −80 to −20 mV) indicating that D433 participates in shutting the pore but not in setting ion selectivity. Hence, the narrowest segment of the ion pathway is not the same in the open and desensitized states. Together, the results predict a significant change in the architecture of the transmembrane segments when the channel adopts the open conformation.

Ca2+-Activated Cl−Current in Retinal Arteriolar Smooth Muscle

To characterize the biophysical, pharmacologic, and functional properties of the Ca(2+)-activated Cl(-) current in retinal arteriolar myocytes. Whole-cell perforated patch-clamp recordings were made from myocytes within intact isolated arteriolar segments. Arteriolar tone was assessed using pressure myography. Depolarizing of voltage steps to -40 mV and greater activated an L-type Ca(2+) current (I(Ca(L))) that was followed by a sustained current. Large tail currents (I(tail)) were observed on stepping back to -80 mV. The sustained current and I(tail) reversed close to 0 mV in symmetrical Cl(-) concentrations. The ion selectivity sequence for I(tail) was I(-)> Cl(-)> glucuronate. Outward I(tail) was sensitive to the Cl(-) channel blockers 9-anthracene-carboxylic acid (9-AC; 1 mM), 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS; 1 mM), and disodium 4,4'-diisothiocyanatostilbene-2,2'-disulfonate (DIDS; 1 mM), but only DIDS produced a substantial (78%) block of inward tail currents at -100 mV. I(tail) was decreased in magnitude when the normal bathing medium was substituted with Ca(2+)-free solution or if I(Ca(L)) was inhibited by 1 microM nimodipine. Caffeine (10 mM) produced large transient currents that reversed close to the Cl(-) equilibrium potential and were blocked by 1 mM DIDS or 100 microM tetracaine. DIDS had no effect on basal vascular tone in pressurized arterioles but dramatically reduced the level of vasoconstriction observed in the presence of 10 nM endothelin-1. Retinal arteriolar myocytes have I(Cl(Ca)), which may be activated by Ca(2+) entry through L-type Ca(2+) channels or Ca(2+) release from intracellular stores. This current appears to contribute to agonist-induced retinal vasoconstriction.

Metal Ion Selectivity of Bifunctional Cation Exchange Resins Containing Phosphonate and Sulfonate Derived from Crosslinked Chloromethylstyrene-styrene Copolymers

Three phosphonate/sulfonate bifunctional resins having different molar ratios of both functional groups were prepared from three kinds of chloromethylstyrene-styrene-divinylbenzene copolymers which were prepared by suspension polymerization of three different molar mixtures of chloromethylstyrene and styrene in the presence of 10 mol % divinylbenzene as crosslinker. Selectivity sequences of these resins toward Ba (II), Cu (II) and Pb (II) were evaluated by measuring their distribution ratios as a function of equilibrium pH of aqueous phases and columnar competitive adsorption of these metal ions as well. It was revealed that the metal ion selectivity sequence of these bifunctional resins was governed by both functional groups and their molar ratios strongly affect the metal ion selectivity sequence; the selectivity sequence of the bifunctional resins approaches that of the monofunctional sulfonate resin with an increase in molar ratio of sulfonate/phosphonate and does that of the monofunctional phosphonate resin with a decrease in molar ratio of sulfonate/phosphonate. These results indicate that the concept of bifunctionality is useful for design of a resin with a novel metal ion selectivity sequence.

Regulation of Kir channels by intracellular pH and extracellular K(+): mechanisms of coupling.

ROMK channels are regulated by internal pH (pHi) and extracellular K+ (K+ o). The mechanisms underlying this regulation were studied in these channels after expression in Xenopus oocytes. Replacement of the COOH-terminal portion of ROMK2 (Kir1.1b) with the corresponding region of the pH-insensitive channel IRK1 (Kir 2.1) produced a chimeric channel (termed C13) with enhanced sensitivity to inhibition by intracellular H+, increasing the apparent pKa for inhibition by ∼0.9 pH units. Three amino acid substitutions at the COOH-terminal end of the second transmembrane helix (I159V, L160M, and I163M) accounted for these effects. These substitutions also made the channels more sensitive to reduction in K+ o, consistent with coupling between the responses to pHi and K+ o. The ion selectivity sequence of the activation of the channel by cations was K+ ≅ Rb+ > NH4 + >> Na+, similar to that for ion permeability, suggesting an interaction with the selectivity filter. We tested a model of coupling in which a pH-sensitive gate can close the pore from the inside, preventing access of K+ from the cytoplasm and increasing sensitivity of the selectivity filter to removal of K+ o. We mimicked closure of this gate using positive membrane potentials to elicit block by intracellular cations. With K+ o between 10 and 110 mM, this resulted in a slow, reversible decrease in conductance. However, additional channel constructs, in which inward rectification was maintained but the pH sensor was abolished, failed to respond to voltage under the same conditions. This indicates that blocking access of intracellular K+ to the selectivity filter cannot account for coupling. The C13 chimera was 10 times more sensitive to extracellular Ba2+ block than was ROMK2, indicating that changes in the COOH terminus affect ion binding to the outer part of the pore. This effect correlated with the sensitivity to inactivation by H+. We conclude that decreasing pHI increases the sensitivity of ROMK2 channels to K+ o by altering the properties of the selectivity filter.

A Highly Selective Salicylate Electrode Based on Schiff Base Complexes of Cobalt(III)

The present article describes a new solvent polymeric membrane anion selective electrode based on Schiff base complexes of Co(III) as neutral carriers, which demonstrates excellent selectivity toward the salicylate ion (Sal−) and anti-Hofmeister selectivity sequence: salicylate>thiocyanate>perchlorate>benzoate>iodide>acetate> nitrite>nitrate>bromide>chloride>citrate>sulfate. The electrode exhibits a near-Nernstian potentiometric response in the range 1.0 × 10−1 ∼1.6 × 10−6 mol/L Sal− with a detection limit of 8.0 × 10−7 mol/L and a slope of −57.2 mV/pSal− in pH 5.5 of phosphate buffer solution at 20°. The response mechanism of the electrode was also studied with the a.c. impedance and spectroscopic techniques. The electrode can be applied to the determination of salicylate in human urine and pharmaceutical preparations with satisfactory results.

METAL ION SELECTIVITY OF MACRORETICULAR CHELATING CATION EXCHANGE RESINS WITH PHOSPHONIC ACID GROUPS ATTACHED TO PHENYL GROUPS OF A STYRENE-DIVINYLBENZENE COPOLYMER MATRIX

To clarify the metal ion selectivity sequence of the phosphonic acid resin named RSPO, in which the functional groups were introduced into phenyl groups of a macroreticular poly(styrene-co-divinylbenzene) matrix, the distribution of 20 kinds of metal ions from 0.0001 mol/L solution of each metal ion onto RSPO was studied in nitric, hydrochloric, sulfuric, and phosphoric acid media. Distribution ratios of divalent metal ions Ba(II), Ca(II), Cd(II), Co(II), Cu(II), Mg(II), Mn(II), Ni(II), Pb(II), Sr(II), and Zn(II) normally decreased with solutions of increased acidity and were not significantly affected by acid species. The clear effect of acid species was observed in the distribution of Fe(III), Mo(VI), and U(VI); although their distributions from hydrochloric and sulfuric acid media normally decreased with increasing acidity of solutions, the minimum was observed in their distribution from nitric acid media, indicating that they were distributed onto the resin from approximately 6 mol/L nitric acid through the mechanism similar to solvation extraction of their nitrate complexes with organophosphorous compounds. Phosphoric acid markedly depressed the distribution of Fe(III), Mo(VI), and U(VI) but not the distribution of Al(III), Bi(III), Cr(III), Gd(III), La(III), and Lu(III). The overall metal ion selectivity sequence of RSPO is the order of Mo(VI) ≅ Fe(III) ≅ U(VI) > Bi(III) ≅ Lu(III) > Al(III) ≅ Gd(III) > La(III) > Cr(III) > Pb(II) > Mn(II) ≅ Cd(II) ≅ Cu(II) > Ca(II) ≅ Co(II) ≅ Zn(II) ≅ Ba(II) ≅ Sr(II) > Ni(II) ≅ Mg(II). The metal ion selectivity sequence was also illustrated on the basis of uptake of U(VI), Pb(II), Ca(II), Ba(II), Ni(II) from 0.01 mol/L solutions of each metal ion.

MRNA encoding ‘ClC-K1, a kidney Cl −- channel’ is expressed in marginal cells of the stria vascularis of rat cochlea: its possible contribution to Cl − currents

The cochlear stria vascularis is essential for the normal function of hair cells. mRNA encoding the ClC-K1 Cl − channel, previously thought to be found only in the kidney, was detected in epithelial marginal cells of the rat stria vascularis by single cell RT-PCR. When Cl − currents were recorded from rat marginal cells by the whole-cell patch clamp method, the steady-state currents showed weak outward rectification and an ion selectivity sequence of SCN −>Br −=Cl −>F −>NO 3 −>I −>gluconate −. The Cl − currents were regulated by extracellular Ca 2+ and pH. These characteristics resemble those reported for the currents recorded from Xenopus oocytes expressing ClC-K1 Cl − channels. These data together suggest that ClC-K1 Cl − channels may contribute to the whole-cell currents of marginal cells.

Permeation Properties of Inward-Rectifier Potassium Channels and Their Molecular Determinants

The structural domains contributing to ion permeation and selectivity in K channels were examined in inward-rectifier K(+) channels ROMK2 (Kir1.1b), IRK1 (Kir2.1), and their chimeras using heterologous expression in Xenopus oocytes. Patch-clamp recordings of single channels were obtained in the cell-attached mode with different permeant cations in the pipette. For inward K(+) conduction, replacing the extracellular loop of ROMK2 with that of IRK1 increased single-channel conductance by 25 pS (from 39 to 63 pS), whereas replacing the COOH terminus of ROMK2 with that of IRK1 decreased conductance by 16 pS (from 39 to 22 pS). These effects were additive and independent of the origin of the NH(2) terminus or transmembrane domains, suggesting that the two domains form two resistors in series. The larger conductance of the extracellular loop of IRK1 was attributable to a single amino acid difference (Thr versus Val) at the 3P position, three residues in front of the GYG motif. Permeability sequences for the conducted ions were similar for the two channels: Tl(+) > K(+) > Rb(+) > NH(4)(+). The ion selectivity sequence for ROMK2 based on conductance ratios was NH(4)(+) (1.6) > K(+) (1) > Tl(+) (0.5) > Rb(+) (0.4). For IRK1, the sequence was K(+) (1) > Tl(+) (0.8) > NH(4)(+) (0.6) >> Rb(+) (0.1). The difference in the NH(4)(+)/ K(+) conductance (1.6) and permeability (0.09) ratios can be explained if NH(4)(+) binds with lower affinity than K(+) to sites within the pore. The relatively low conductances of NH(4)(+) and Rb(+) through IRK1 were again attributable to the 3P position within the P region. Site-directed mutagenesis showed that the IRK1 selectivity pattern required either Thr or Ser at this position. In contrast, the COOH-terminal domain conferred the relatively high Tl(+) conductance in IRK1. We propose that the P-region and the COOH terminus contribute independently to the conductance and selectivity properties of the pore.

Electrophysiological characterization of the rat epithelial Na+ channel (rENaC) expressed in MDCK cells. Effects of Na+ and Ca2+.

The epithelial Na+ channel (ENaC), composed of three subunits (alpha, beta, and gamma), is expressed in several epithelia and plays a critical role in salt and water balance and in the regulation of blood pressure. Little is known, however, about the electrophysiological properties of this cloned channel when expressed in epithelial cells. Using whole-cell and single channel current recording techniques, we have now characterized the rat alpha beta gamma ENaC (rENaC) stably transfected and expressed in Madin-Darby canine kidney (MDCK) cells. Under whole-cell patch-clamp configuration, the alpha beta gamma rENaC-expressing MDCK cells exhibited greater whole cell Na+ current at -143 mV (-1,466.2 +/- 297.5 pA) than did untransfected cells (-47.6 +/- 10.7 pA). This conductance was completely and reversibly inhibited by 10 microM amiloride, with a Ki of 20 nM at a membrane potential of -103 mV; the amiloride inhibition was slightly voltage dependent. Amiloride-sensitive whole-cell current of MDCK cells expressing alpha beta or alpha gamma subunits alone was -115.2 +/- 41.4 pA and -52.1 +/- 24.5 pA at -143 mV, respectively, similar to the whole-cell Na+ current of untransfected cells. Relaxation analysis of the amiloride-sensitive current after voltage steps suggested that the channels were activated by membrane hyperpolarization. Ion selectivity sequence of the Na+ conductance was Li+ > Na+ >> K+ = N-methyl-D-glucamine+ (NMDG+). Using excised outside-out patches, amiloride-sensitive single channel conductance, likely responsible for the macroscopic Na+ channel current, was found to be approximately 5 and 8 pS when Na+ and Li+ were used as a charge carrier, respectively. K+ conductance through the channel was undetectable. The channel activity, defined as a product of the number of active channel (n) and open probability (Po), was increased by membrane hyperpolarization. Both whole-cell Na+ current and conductance were saturated with increased extracellular Na+ concentrations, which likely resulted from saturation of the single channel conductance. The channel activity (nPo) was significantly decreased when cytosolic Na+ concentration was increased from 0 to 50 mM in inside-out patches. Whole-cell Na+ conductance (with Li+ as a charge carrier) was inhibited by the addition of ionomycin (microM) and Ca2+ (1 mM) to the bath. Dialysis of the cells with a pipette solution containing 1 microM Ca2+ caused a biphasic inhibition, with time constants of 1.7 +/- 0.3 min (n = 3) and 128.4 +/- 33.4 min (n = 3). An increase in cytosolic Ca2+ concentration from <1 nM to 1 microM was accompanied by a decrease in channel activity. Increasing cytosolic Ca2+ to 10 microM exhibited a pronounced inhibitory effect. Single channel conductance, however, was unchanged by increasing free Ca2+ concentrations from <1 nM to 10 microM. Collectively, these results provide the first characterization of rENaC heterologously expressed in a mammalian epithelial cell line, and provide evidence for channel regulation by cytosolic Na+ and Ca2+.

Cl- and K+ conductances activated by cell swelling in primary cultures of rabbit distal bright convoluted tubules.

Ionic currents induced by cell swelling were characterized in primary cultures of rabbit distal bright convoluted tubule (DCTb) by the whole cell patch-clamp technique. Cl- currents were produced spontaneously by whole cell recording with an isotonic pipette solution or by exposure to a hypotonic stress. Initial Cl- currents exhibited outwardly rectifying current-voltage relationship, whereas steady-state currents showed strong decay with depolarizing pulses. The ion selectivity sequence was I- = Br- > Cl- >> glutamate. Currents were inhibited by 0.1 mM 5-nitro-2-(3-phenylpropylamino) benzoic acid and 1 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid and strongly blocked by 1 mM diphenylamine-2-carboxylate. Currents were insensitive to intracellular Ca2+ but required the presence of extracellular Ca2+. They were not activated in cells pretreated with 200 nM staurosporine, 50 microM LaCl3, 10 microM nifedipine, 100 microM verapamil, 5 microM tamoxifen, and 50 microM dideoxyforskolin. Staurosporine, tamoxifen, verapamil, or the absence of external Ca2+ was without effect on the fully developed Cl- currents. Osmotic shock also activated K+ currents in Cl- free conditions. These currents were time independent, activated at depolarized potentials, and inhibited by 5 mM BaCl2. The activation of Cl- and K+ currents by an osmotic shock may be implicated in regulatory volume decrease in DCTb cells.

Calcium-activated chloride currents in primary cultures of rabbit distal convoluted tubule.

Chloride (Cl-) conductances were studied in primary cultures of rabbit distal convoluted tubule (very early distal "bright" convoluted tubule, DCTb) by the whole cell patch-clamp technique. We identified a Cl- current activated by 2 microM extracellular ionomycin. The kinetics of the macroscopic current were time dependent for depolarizing potentials with a slow developing component. The steady state current presented outward rectification, and the ion selectivity sequence was I- > Br- > > Cl > glutamate. The current was inhibited by 0.1 mM 5-nitro-2-(3-phenylpropyl-amino)benzoic acid, 1 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, and 1 mM diphenylamine-2-carboxylate. To identify the location of the Cl- conductance, 6-methoxy-N-(3-sulfopropyl)quinolinium fluorescence experiments were carried out in confluent cultures developed on collagen-coated permeable filters. Cl- removal from the apical solution induced a Cl- efflux that was stimulated by 10 microM forskolin. Forskolin had no effect on the basolateral Cl- permeability Cl- substitution in the basolateral solution induced an efflux stimulated by 2 microM ionomycin or 50 microM extracellular ATP Ionomycin had no effect on the apical Cl- fluxes. Thus cultured DCTb cells exhibit Ca(2+)-activated Cl- channels located in the basolateral membrane. This Cl- permeability was active at a resting membrane potential and could participate in the Cl- reabsorption across the DCTb in control conditions.

Pore properties of Lymnaea stagnalis neuron stretch-activated K+ channels

In many neurons, variations in membrane excitability are determined by a resting K+ conductance whose magnitude is modulated via neurotransmitters. The S-channel in Aplysia californica mechanosensory neurons is such a conductance, but it has also been shown to be a stretch-activated K+ channel. In this, it resembles stretch-activated K+ channels common to all molluscan neurons. Comparable channels are widespread, having been reported in molluscan and insect muscle and various vertebrate cells. The pore properties of the S-channel and similar stretch-activated K+ channels have received only sporadic attention. Here we examine, at the single-channel level, the permeation characteristics of a stretch-activated K+ channel from neurons of the mollusc Lymnaea stagnalis. Michaelis&shy;Menten constants (Km) for the conductance, obtained separately for inward (28 mmol l-1) and outward (91 mmol l-1) K+ currents, suggest that the channel presents to the external medium, where [K+] is lower, a higher-affinity site than it presents to the cytoplasmic medium. This may help to ensure that influx is not diffusion-limited at potentials near the resting potential, i.e. near the K+ equilibrium constant. Anomalous mole fraction behavior, observed when the ratio of permeant ion (K+ and Rb+) was varied, indicated that the stretch-activated K+ channel is a multi-ion pore. The ion selectivity sequence determined using reversal potentials under bi-ionic conditions was Cs+>K+>Rb+>NH4+>Na+>Li+, and using relative conductance in symmetrical solutions, the sequence was Tl+=K+>Rb+>NH4+>>Na+=Li+=Cs+. Extreme variations in extracellular pH from 4.7 to 11.4 had no effect on stretch-activated K+ channel conductance, whereas normal concentrations of extracellular Mg2+ reduced inward K+ current. Intracellular, but not extracellular, Ba2+ produced a slow, open channel block with an IC50 of 140&plusmn;80 &micro;mol l-1. These pore properties are compared with those of other stretch-activated K+ channels and of K+ channels in general. In spite of a greater than half order of magnitude difference in the cytoplasmic [K+] in marine (Aplysia californica) and freshwater (Lymnaea stagnalis) molluscs, the conductances of stretch-activated K+ channels from the two groups are very similar.

Functional role of a conserved aspartate in the external mouth of voltage-gated potassium channels

Mutation of the glycines in a conserved Gly-Tyr-Gly-Asp sequence in the P-region of voltage-gated K channels has identified determinants of Na/K selectivity. But the function of the negatively charged Asp is not known because mutations at this position are not tolerated, owing to the fourfold replication of mutations in a tetrameric channel. We have successfully mutated Asp378-->Thr in a tandem dimer Kv2.1 construct to yield a twofold neutralization of charge at this site. When expressed in Xenopus oocytes, the mutated channels showed markedly altered ion conduction and blockade. Potassium conduction in the inward direction was selectively reduced, so that the instantaneous current-voltage relationship obtained in isotonic KCl became strongly outwardly rectifying. The relative permeability to Na+, PNa/PK, increased from 0.02 to 0.10 without changing the ion selectivity sequence K > Rb >> Cs >> Na. The IC50 for block by external tetraethylammonium (TEA) increased more than 100-fold without affecting block by internal TEA. We conclude that Asp378 is an essential part of a potassium ion binding site associated with the Na/K selectivity filter at the external mouth of the pore.

A chloride current associated with swelling of cultured chick heart cells.

1. Cultured chick heart cells challenged by hyposmotic stress underwent regulatory volume decrease (RVD) that was attenuated by prior depletion of intracellular chloride. 2. During hyposmotic swelling, cell aggregates experienced an initial increase in spontaneous contractile activity followed by eventual quiescence. Conventional microelectrode studies revealed an underlying increase in spontaneous electrical activity, followed by a sustained depolarization beyond threshold. 3. Whole-cell patch clamp studies, with K+ currents blocked, indicated that exposure of cells to hyposmotic solution (NaCl reduction) resulted in a rapid osmotic swelling followed by a substantial increase in whole-cell conductance which persisted for the duration of hyposmotic exposure and was almost completely reversed on return to isosmotic bath solution. 4. For a variety of Cl- concentrations, the reversal potentials (Erev) of the measured swelling-activated current closely followed the calculated Cl- equilibrium potential (ECl) with a linear regression slope of 0.82. When estimated by the Nernst equation, the relationship between Erev and the [Cl-]i/[Cl-]o ratio fitted well with a slope of 51 mV per decade change in the concentration ratio, consistent with a Cl(-)-selective conductance. 5. The permeability ratios of this swelling-activated conductance to chloride, methanesulphonate (MSA) and aspartate (Asp) were calculated as PCl:PMSA:PASP = 1:0.36:0.02, with the ion selectivity sequence of Cl- > MSA- >> Asp-, which suggests the swelling-activated conductance is slightly permeable to other anions. 6. Application of a Cl- channel blocker, diphenylamine-2-carboxylate (DPC, 200 microM), substantially suppressed the swelling-activated current without shifting the Erev of this current. The effect of DPC was independent of membrane potential. 7. This evidence demonstrates that hyposmotic swelling of cultured chick heart cells activates a channel-mediated Cl- conductance which may be associated with the integrated response of volume-regulatory mechanisms.

Polyvalent cations inhibit human neutrophil chemotaxis by interfering with the polymerization of actin.

The effect of a series of di- and trivalent cations on the locomotor response of human neutrophils to the chemotactic tripeptide N-formyl-methionyl-leucyl-phenylalanine (FMLP) was investigated. Migration was assessed by the leading front method. The cations inhibited FMLP-stimulated chemotaxis in the rank order: Ni2+ approximately Co2+ greater than Sr2+ greater than Zn2+ greater than Mn2+ approximately La3+ greater than Cd2+ approximately Ba2+ much greater than Mg2+. Benzamil, which blocks Na+/Ca2+ exchange, did not alter chemotaxis by itself but prevented the suppressive effects of each of the polyvalent cations on motility. The ion selectivity sequence and the lack of activity of benzamil are strikingly different than for O(-2) generation, thereby implying different modes of action in the two functional expressions. The F-actin content of the cells was monitored by the fluorescence of rhodamine-phalloidin. Each of the cations displayed comparable efficacy in blocking the polymerization of actin in FMLP-activated cells. Likewise, benzamil exhibited a protective effect, completely overcoming the inhibitory action of the polyvalent cations. The results indicate that these foreign ions gain access to the cell interior via a benzamil-sensitive pathway, namely Na+/Ca2+ exchange. Upon entry into the cytosol, they then interfere with the formation of filaments from actin monomers. These studies help to shed light on the interaction of divalent cations with cytoskeletal and contractile elements in cell motility.

Facilitated transport of alkali metal picrates through acetylcellulose blend membranes containing synthetic macrolides

AbstractPermeation of alkali metal picrates through acetylcellulose membranes containing synthetic macrolides (4 and 5) was examined in tris‐buffer solutions (pH 7,8) to evaluate the effectiveness of the macrolides as ion carriers. The macrolides 4 and 5 are 20‐ and 25‐membered macrocyclic oligoesters, respectively, consisting of alternating tetrahydropyran ring and ester moieties, which were prepared by cationic ring‐opening polymerization of 6,8‐dioxabicyclo[3.2.1]octan‐7‐one (1). Ion permeability through the blend membranes increased with increasing macrolide content. The macrolide 5 facilitated ion transport more effectively than the macrolide 4. Dependence of ion permeability on solute concentration in a source phase was analyzed by using Lineweaver‐Burk's equation, and it was revealed that the ion transport through the blend membranes obeyed kinetics of a Michaelis‐Menten type. Ion selectivity sequence for the permeation of alkali metal picrates through the membrane containing 5 was, although not remarkable, Na+ &gt; K+ &gt; Cs+, in sharp contrast to the previously reported ion selectivity sequence for the transport mediated by the macrolides through liquid organic membranes.

Channel properties of the purified acetylcholine receptor from Torpedo californica reconstituted in planar lipid bilayer membranes

The electrophysiological properties of the cation channel of the purified nicotinic acetylcholine receptor (AChR) reconstituted in planar lipid bilayers were characterized. Single-channel currents were activated by acetylcholine, carbamylcholine and suberyldicholine. The single channel conductance (28 pS in 0.3 M NaCl) was ohmic and independent of the agonist. Single channel currents increased with Na+ concentration to a maximum conductance of 95 pS and showed a half-saturation point of 395 mM. The apparent ion selectivity sequence, derived from single-channel current recordings, is: NH+4 greater than Cs+ greater than Rb+ greater than or equal to Na+ Cl-, F-, SO2-(4). The distribution of channel open times was fit by a sum of two exponentials, reflecting the existence of at least two distinct open states. The time constants depend on the choice of agonist, being consistently longer for suberyldicholine than for carbamylcholine. Similar channel properties were recorded in bilayers formed from monolayers at the tip of patch pipets . Single-channel currents occur in paroxysms of channel activity followed by quiescent periods. This pattern is more pronounced as the agonist concentration increases, and is reflected in histograms of channel-opening frequencies. Computer simulations with a three-state model, consisting of two closed (unliganded and liganded) and one open state, do not resemble the recorded pattern of channel activity, especially at high agonist concentration. Inclusion of a desensitized liganded state reproduces the qualitative features of channel recordings. The occurrence of paroxysms of channel activity thus seems to result from the transit of AChR through its active conformation, from which it can open several times before desensitizing.