Local Anesthetics Affect Gramicidin A Channels via Membrane Electrostatic Potentials

The effects of local anesthetics (LAs), including aminoamides and aminoesters, on the characteristics of single gramicidin A (GA) channels in 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) bilayers were studied. Aminoamides, namely lidocaine (LDC), prilocaine (PLC), mepivacaine (MPV), and bupivacaine (BPV), reduced the conductance of GA channels. Aminoesters influenced the current fluctuations induced by GA differently; procaine (PC) did not affect the fluctuations, whereas tetracaine (TTC) distinctly reduced the conductance of single GA channels. Using electrophysiological technique, we estimated the changes in the membrane boundary potential at the adsorption of LAs; LDC, PLC, MPV, BPV, and TTC substantially increased, while PC did not affect it. To elucidate which component of the membrane boundary potential, the surface or dipole potential, is responsible for the observed effects of LAs, we employed a fluorescence assay. We found that TTC led to a significant increase in the membrane dipole potential, whereas the adsorption of LDC, PLC, MPV, BPV, and PC did not produce any changes in the membrane dipole potential. We concluded that aminoamides affected the surface potential of lipid bilayers. Together, these data suggest that the effects of LAs on the conductance of single GA channels are caused by their influence on membrane electrostatic potentials; the regulation of GA pores by aminoamides is associated with the surface potential of membranes, whereas TTC modulation of channel properties is predominantly due to changes in dipole potential of lipid bilayers. These data might provide some significant implications for voltage-gated ion channels of cell membranes.     

Cell Membrane Potentials of Higher Plants: Changes Induced by Wounding

Cell membrane potentials of cultured soybean cells, freshlycut soybean cotyledons, and freshly cut Jerusalem artichoketubers ranged from –20 to –45 mV. Readings fromthe cultured soybean cells remained within this range over aperiod of several days after the cells were subcultured. However,the membrane potentials of the soybean cotyledonary chlorenchymaand of the Jerusalem artichoke storage parenchyma exhibiteda striking hyperpolarization over a period of hours after thesetissues were sliced. The soybean cotyledons exhibited a morerapid hypopolarization than the artichoke tubers, developinga maximum average potential of –80 mV within 3 h afterwounding. These readings were obtained with the cells bathedin a complete culture medium known to permit sustained cellproliferation from soybean cotyledons. However, the wound-inducedhyperpolarization was not dependent upon any particular componentof the medium, including auxin or cytokinin. The hyperpolarizationwas observed even when cotyledon slices were incubated in isosmoticKCl.     

Membrane Potentials in the Xylem in Roots of Intact Plants

The membrane potential differences (PDs) of root cells of intact,illuminated Trifolium repens L. and Lolium perenne L. have beenmeasured. In T. repens the PDs were the same for all cell typesexcept for the xylem vessels, which were more positive, andfor some cells immediately adjacent to the xylem vessels whichwere 10 mV more negative. The mean PD for all cells was emdash164.6 ± 0.6 mV and the mean for cells adjacent to thexylem vessels with elevated PDs was 178.4 ± 2.4 mV. Whenthe electrode tip was in a xylem vessel a low but stable PD(mean = emdash 89.9 mV) was recorded. The results for L. perennewere similar except that there were no cells with elevated PDsadjacent to the xylem vessels. An inhibitor of ion transport from the root to the shoot, p-fluorophenylalanine(p-FPA), caused a depolarization of 10 mV in the cell PDs butin the xylem vessels the depolarization was 50 mV. The possibility that the elevated PDs of cells adjacent to thexylem vessels are related to the transport of ions into thevessels is discussed.     

The Effect of Electrotonus on the Olfactory Epithelium

The effect of electrotonus on the slow potential of the olfactory epithelium of the frog was studied. The "on"-slow potential induced by a general odor like amyl acetate increased its magnitude in accordance with increase of anodal current, while it decreased its magnitude with increase of cathodal current. Similar relations were also found in the case of the vapors of organic solvents like ethyl ether of low concentrations. Conversely, the on-slow potential induced by the vapors of organic solvents of high concentration decreased its magnitude in accordance with the increase of anodal current, while it increased its magnitude with the increase of cathodal current. The "off"-slow potential induced by the vapors of organic solvents of high concentration showed a potential change under the action of electrotonic currents which is similar to the change of the on-slow potential induced by general odors. It was concluded that there are two receptive processes in the olfactory cell. One is an ordinary excitatory process which produces an electronegative slow potential in response to general odors. The other is a process of a different kind which is activated only by the vapor of an organic solvent of high concentration and which shows an entirely opposite reaction from that generally found in excitable tissues when an electrotonic current is applied.     

Passive Electrical Properties of Microorganisms: II. Resistance of the Bacterial Membrane

Studies of the effective, homogeneous, dielectric constants of bacteria are used to show that the resistances of their cytoplasmic membranes are too great to explain the low-frequency conductivities which have been observed for these organisms. This reaffirms the conclusion that at low frequencies the conductivities of bacteria reflect properties of their cell walls. In the organisms studied, the conductivities of the cell wall region are as great as the conductivities of the cytoplasm. This is true even though the ion concentration in the environment is much less than that in the cells. The mobile ions of the wall are presumed to be counterions for fixed charges in this region.     

Membrane Potentials in Excitable Cells of Aldrovanda vesiculosa Trap-Lobes

The resting membrane potential in excitable cells of Aldrovandatrap-lobes is composed of diffusion and electrogenic potentials.The diffusion potential, about –100 mV in artificial pondwater, was determined from the external K⁺ and Na⁺ concentrations.The permeability ratio, P_Na/P_K of the membrane was estimatedto be about 0.3. The electrogenic potential hyperpolarized themembrane to about –140 mV. The peak value of the actionpotential increased by +26 mV with a tenfold increase in theexternal Ca²⁺ concentration. The action potential was blockedby an application of the Ca²⁺ chelater or the Ca channel blocker,LaCl₃. Cells showed additional Ca²⁺ influx (7.8 pmole/cm² impulse)during membrane excitation. These facts suggest that the transientincrease in Ca²⁺ influx causes the action potential presentin cells of Aldrovanda trap-lobes. ¹ Present address: Jerry Lewis Neuromuscular Research Center,School of Medicine, University of California Los Angeles, LosAngeles, CA90024, U.S.A. ² Present address: Biological Laboratory, Kyoritsu Women's University,Hachioji 193, Japan. (Received September 21, 1983; Accepted September 7, 1984)     

Membrane Permeability: Generalization of the Reflection Coefficient Method of Describing Volume and Solute Flows

The reflection coefficient method for describing volume and solute fluxes through membranes is generalized to take into account the nonideality of the solutions bathing the membrane and/or multicomponent systems. The reflection coefficient of the impermeable species in these systems is less than unity by a coefficient γ. The reflection coefficient obtained solely from the volume flow equation, σ^v, will always be less than the reflection coefficient obtained from the solute flow equation, σ^8v. These two coefficients are related by σ^8v = σ^v + γ.     

Influence of Phenolic Acids on Ion Uptake: IV. Depolarization of Membrane Potentials

The membrane potentials of aged, excised barley (Hordeum vulgare L.) root cells were rapidly depolarized by the addition of salicylic acid (o-hydroxybenzoic acid) to the buffered medium bathing root segments. Initial values for membrane potentials were restored very slowly (within 100 minutes) by replacing the phenolic solution by phenolic-free buffer. Several other naturally occurring benzoic and cinnamic acids depolarized cell membrane potentials. The cinnamic acids consistently caused a greater depolarization than the correspondingly substituted benzoic acids. A strong positive correlation was found between the depolarization values (ΔE) for the benzoic acids and their lipid solubilities. This study supports the hypothesis that the inhibition of ion uptake brought about by naturally occurring phenolic acids is caused by a generalized increase in membrane permeability to inorganic ions.     

A Generalization of the GMANOVA-Model

It is shown that in some experimental designs the MANOVA- and the GMANOVA-model are too restrictive either to yield all hypothesis tests of interest or to reflect all known features of the design. An extension of these models is derived by relating the response vectors with the unknown model parameters by linear equations which may be completely different for each of the p components of the response vector and for each of the n independent vectors. For situations, in which a Wishart-distributed estimator for the underlying common covariance matrix is attainable, a test for any s-dimensional linear hypothesis on the model parameters is derived.     

Membrane Potentials at Zero Current: The Significance of a Constant Ionic Permeability Ratio

The possibility has been examined that the Goldman-Hodgkin-Katz equation for V(0), the total membrane potential at zero current, can be derived with constant permeability ratios from a thermodynamic treatment. The flux equations have been integrated under zero current conditions subject only to the restriction that the total membrane potential should be independent of internal concentration profiles, which is the requirement for the premeability ratios to be phenomenological constants, independent of solution conditions. No assumptions have been made concerning the electric potential profile. It was found that a constant permeability ratio can only be characteristic of systems satisfying certain relationships between ionic conductances and chemical potentials. From these relationships it was possible to define the permeability ratio in terms of the thermodynamic properties of the membrane quite generally and to identify the permeability ratio as the product of mobility ratio and ratio of partition coefficients. Moreover, the ionic conductance ratio at any point in the membrane has been shown to be expressable explicitly in terms of the permeability ratio and the activities of an external solution which would be in equilibrium with the point under consideration. Lastly, a number of conclusions have been reached regarding the range of applicability of the Goldman-Hodgkin-Katz equation with constant permeability ratios.     

Membrane Potentials of the Lobster Giant Axon Obtained by Use of the Sucrose-Gap Technique

A method similar to the sucrose-gap technique introduced be Stäpfli is described for measuring membrane potential and current in singly lobster giant axons (diameter about 100 micra). The isotonic sucrose solution used to perfuse the gaps raises the external leakage resistance so that the recorded potential is only about 5 per cent less than the actual membrane potential. However, the resting potential of an axon in the sucrose-gap arrangement is increased 20 to 60 mv over that recorded by a conventional micropipette electrode when the entire axon is bathed in sea water. A complete explanation for this effect has not been discovered. The relation between resting potential and external potassium and sodium ion concentrations shows that potassium carries most of the current in a depolarized axon in the sucrose-gap arrangement, but that near the resting potential other ions make significant contributions. Lowering the external chloride concentration decreases the resting potential. Varying the concentration of the sucrose solution has little effect. A study of the impedance changes associated with the action potential shows that the membrane resistance decreases to a minimum at the peak of the spike and returns to near its initial value before repolarization is complete (a normal lobster giant axon action potential does not have an undershoot). Action potentials recorded simultaneously by the sucrose-gap technique and by micropipette electrodes are practically superposable.     

Electrophysiological Techniques and the Magnitudes of the Membrane Potentials and Resistances of Nitella translucens

The membrane resistance of internodal cells of Nitella translucensincreased by 50 per cent during the first 5 h after insertionof two microelectrodes into the vacuole even when precautionswere taken to eliminate external disturbances. The insertionof a third microelectrode into the cytoplasm did not affectthe resistance. In artificial pond water the final value forthe plasmalemma resistance was 112 k cm² and that for the tonoplastwas 6 k cm². The increase in the membrane potential after thefirst hour was less than 10 per cent. A recent suggestion that accurate measurements of the plasmalemmaresistance can be made with a microelectrode outside the plasmalemma,but in close contact with it, is criticized. Tests were made of the claim that leakage of current at thepoint where microelectrodes enter the cytoplasm gives rise toa local increase in current density at the tonoplast and henceleads to an overestimate of the tonoplast resistance. Valuesfor the tonoplast resistance obtained when the cytoplasmic microelectrodewas inserted through the plasmalemma were similar to those observedwhen it was pushed across the cell and inserted through thetonoplast at a point remote from the postulated current leakage.Furthermore, the tonoplast resistance stayed remarkably constantwhen the plasmalemma resistance varied in a way which wouldcause different proportions of the applied current to pass throughthe leak resistance and produce variations in the apparent tonoplastresistance. It is concluded that published values of the tonoplastresistance are not grossly inaccurate.     

Studies on Mechano-Perception in Characeae: Decrease in Electrical Membrane Resistance in Receptor Potentials

A polyacrylate chamber with three pools was developed to measurethe changes in electrical membrane resistance during generationof receptor potentials upon mechanical stimulation in Charainternodal cells. The membrane resistance decreased at the peakof the receptor potentials. The present results support thehypothesis that the receptor potential is generated by activationof ion channel(s). This is the first success in demonstratinga change in the membrane resistance in receptor potential inducedmechanically in plants. (Received June 17, 1997; Accepted September 12, 1997)     

Membrane Potential Dependent Duration of Action Potentials in Cultured Rat Hippocampal Neurons

(1) Fluctuations of the membrane potential states are essential for the brain functions from the response of individual neurons to the cognitive function of the brain. It has been reported in slice preparations that the action potential duration is dependent on the membrane potential states. (2) In order to examine whether dependence of action potential duration on the membrane potential could happen in isolated individual neurons that have no network connections, we studied the membrane potential dependence of the action potential duration by artificially setting the membrane potentials to different states in individual cultured rat hippocampal neurons using patch-clamp technique. (3) We showed that the action potential of individual neurons generated from depolarized membrane potentials had broader durations than those generated from hyperpolarized membrane potentials. (4) Furthermore, the membrane potential dependence of the action potential duration was significantly reduced in the presence of voltage-gated K⁺ channel blockers, TEA, and 4-AP, suggesting involvement of both delayed rectifier I _K and transient I _A current in the membrane potential dependence of the action potential duration. (5) These results indicated that the dependence of action potential duration on the membrane potential states could be an intrinsic property of individual neurons. Bo Gong and Mingna Liu contributed equally to this work.     

Interactions of Calcium with Sodium and Potassium in Membrane Potentials of the Lobster Giant Axon

Experiments were performed on the lobster giant axon to determine the relation between intracellular spike amplitude and external calcium ion concentration. Action potential decline in low external calcium is greatly accelerated by simultaneous removal of external sodium ion. Correlation of the time course of spike decline in low calcium-low sodium solution with the time courses of spike decline in low calcium alone and in low sodium alone indicates that the effect of simultaneous removal of both ions is significantly greater than the sum of the individual effects. For a given time of treatment, spike amplitude was a function of external calcium concentration. While spike height is proportional to the log of the external calcium concentration over the range 2.5 to 50 millimolar, the proportionality constant is dependent upon the sodium concentration. Under the conditions of low external sodium (50 per cent reduction) the slope of the linear relationship between the spike height and the log of the external calcium concentration is about 5 times greater than in normal external sodium. Decreasing external calcium concentration and simultaneously increasing external potassium concentration produce a greater spike reduction than the arithmetic sum of spike reductions in low calcium alone and in high potassium alone. It is suggested that calcium interacts strongly with sodium and potassium in the spike-generating mechanism. A theoretical basis for these results is discussed.     

Effect of Root Position on Measured Root Position on Measured Root Membrane Potentials

Root membrane potentials were measured by interposing a plantroot between two KCI solutions of different concentrations.The potentials measured across the two calomel electrodes werefound to depend on the position of the root. The potentialswere found to be lower when the root tip rather than the rootbase was in contact with the more concentrated solution. Thisindicates that the two parts of the root do not have the samein transport properties. Using an approximate theoretical treatmentthe observed potentials could be accounted for.