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D A Haydon

Publications and source records attributed to D A Haydon.

At least 19 recordsLinked to original sources
PubMed1991

The effects of short-chain phospholipids on the acetylcholine-activated ion channel.

The effects of a homologous series of short-chain phospholipids, the phosphatidylcholines from dihexanoylglycerophosphocholine (Hxo2GroPCho) to didecanoylglycerophosphocholine, on the nicotinic acetylcholine-activated ion channel in cultured rat muscle cells were investigated. Standard patch-clamp techniques were used to measure single-channel currents in excised patches. All phospholipids investigated markedly reduced the frequency of channel opening in a concentration-dependent manner. Other parameters, such as the mean open time, the duration and frequency of brief closures within an opening, and channel amplitude, were not significantly affected. This effect was independent of the side of the membrane to which the phospholipid was added. Dose/response curves were obtained for Hxo2-, diheptanoyl(Hpo2)- and dinonanoyl(Nno2)GroPCho. The concentration leading to 50% reduction in channel activity decreased upon ascending the homologous series from 16.69 microM Hxo2GroPCho to 4.52 microM and 0.043 microM for Hpo2- and Nno2GroPCho, respectively. The more hydrophobic the molecule the more effective it was, and hence the higher its affinity to the binding site. Calculation of the standard free-energy change of adsorption into the site led to a value of -3.1 kJ/mol, which indicates a very hydrophobic binding site. In conclusion, the phospholipids interact in a non-specific way with the lipid membrane thereby disturbing proper channel function.

Acetylcholine↗
PubMed1991

Effects of anesthetics and convulsants on the resting potassium conductance in squid nerve.

The effects of some fluorinated anesthetics and convulsants on the ionic conductances of the axon membrane of the squid Loligo forbesi are described. The substances studied were the inhalation anesthetics enflurane (CF2HOCF2CClFH) and isoflurane (CF2HOCClHCF3) and the convulsants flurothyl (CF3CH2OCH2CF3) and trichlorofluormethane (CCl3F). At low concentrations (0.4-0.8 mM), none of these substances significantly affected the voltage-dependent Na and K channels of the nerve. However, at these concentrations each substance produced a depolarization of the resting potential and reduced the potassium conductance of the resting membrane. This was associated with a tendency to cause axonal hyperexcitability. The potassium conductance of the resting membrane was separated into a component arising from residual open Hodgkin-Huxley delayed rectifier K channels and another voltage-independent component (g'k). The former component was insensitive to the test substances at 0.4-0.8 mM, while g'k was inhibited by 40-80%. It is suggested that the convulsant activity of certain small fluorinated molecules, and the proconvulsant actions of certain clinical anesthetics at low concentrations, may be related to the inhibition of a resting, voltage-independent potassium conductance system in the nerve membrane.

Action Potentials↗
PubMed1991

Actions of n-alcohols on nicotinic acetylcholine receptor ion channels in cultured rat muscle cells.

The effects of the n-alcohols from pentanol to dodecanol on nAChR channel function were resolved at the single channel level. ACh-activated channel activity was recorded from isolated membrane patches using the patch clamp method. The intermediate-chain alcohols (C5-C8) had two main effects: (1) They caused channel openings to be interrupted by brief shut or blocked periods, the duration of which was dependent on chain length of the alcohol but independent of concentration. (2) They caused a reduction in the duration of bursts of openings. The long-chain alcohols (C9-C11) produced only the second effect, and there was a decline in activity beyond undecanol. Results were consistent with a mechanism of channel blockade and were analyzed in terms of an open channel block model with a long-lived closed-blocked state beyond the blocked state. Affinity for the binding site increased with chain length up to octanol. The standard free energy per methylene group for adsorption to the site was calculated to be -3.3 kJ/mol, indicating the very hydrophobic nature of the site.

Alcohols↗
PubMed1991

Actions of n-alcohols on nicotinic acetylcholine receptor channels in cultured rat myotubes.

1. The actions of the n-alcohols from pentanol to dodecanol on nicotinic acetylcholine receptor (nAChR) channels were investigated by recording single ACh-activated channel activity from inside-out membrane patches isolated from cultured rat myotubes. Alcohols were applied to the cytoplasmic side of the membrane; aqueous concentrations ranged from 11.7 mM-pentanol to 0.02 mM-dodecanol. 2. The intermediate-chain alcohols (pentanol to octanol) caused channel currents to fluctuate between the fully open and closed state level so that openings occurred in bursts interrupted by brief gaps. Closed time distributions were fitted well with two exponential components, the fast component representing the closures within a burst. The number of gaps within a burst was dependent on alcohol concentration whereas gap duration was independent of concentration but increased with increasing chain length of the alcohol up to octanol. 3. Nonanol and decanol reduced the mean duration of bursts of openings but did not cause an increase in the number of short closed intervals within a burst. Beyond decanol there was a decline in the ability of the n-alcohols to affect channel function. A saturated solution of undecanol (0.07 mM) reduced the mean open time by 33 +/- 17%, whereas a saturated solution of dodecanol had no significant effect. 4. The current integral per burst was reduced by all the n-alcohols between pentanol and undecanol. The IC50S were as follows: hexanol, 0.53 +/- 0.14 mM; heptanol, 0.097 +/- 0.02 mM; octanol, 0.04 mM and nonanol, 0.16 +/- 0.035 mM. 5. The results were analysed in terms of an open channel block model with a long-lived closed-blocked state beyond the blocked state. Over the range of concentrations tested this describes the effects of all the n-alcohols (C5 to C12) on channel gating reasonably well. 6. Blocking rate constants (k+B) for pentanol through to nonanol were calculated to be between 2.8 and 5.7 X 10(6) M-1 S-1. These values are based on the assumption that the concentration of the alcohols at their site(s) of action was equal to the aqueous concentration applied to the membrane. 7. Equilibrium dissociation constants (KD), calculated from the blocking and unblocking rate constants (KD = k-B/k+B), decreased with increasing chain length from 8 mM for pentanol to 0.15 mM for octanol. The standard free energy per methylene group for adsorption to the site of action was calculated to be about -3.3 kJ mol-1.(ABSTRACT TRUNCATED AT 400 WORDS)

1-Octanol↗
PubMed1989

The effects of homologous series of anaesthetics on a resting potassium conductance of the squid giant axon.

The effects of n-alkanes (n-pentane to n-octane), n-alkanols (n-pentanol to n-undecanol) and two carboxylic esters (methyl pentanoate and methyl octanoate) on the conductance of squid giant axons in a high potassium, zero sodium bathing solution have been examined. Sodium and delayed rectifier potassium channels were as far as possible pharmacologically blocked. A substantial fraction of the measured conductance is attributed to a recently-described, voltage-independent, potassium channel. Anaesthetics block this channel but its sensitivity is markedly different from those of other squid axon ion channels.

Anesthetics↗
PubMed1988

The potassium conductance of the resting squid axon and its blockage by clinical concentrations of general anaesthetics.

1. The effects of some neutral clinical and experimental general anaesthetics on the resting potential of normal squid axons and squid axons exposed to tetrodotoxin and 3,4-diaminopyridine have been studied. 2. Depolarizations of 1-4 mV were produced by all the anaesthetics at 'clinical' concentrations in the normal axon. Larger depolarizations (5-11 mV) were produced by the same anaesthetic concentrations in axons exposed to tetrodotoxin and 3,4-diaminopyridine. 3. The conductance of axons exposed to tetrodotoxin and either tetraethyl-ammonium or 3,4-diaminopyridine in zero Na+, 430 mM-K+ artificial sea water was examined by voltage clamp and AC bridge techniques. 4. The evidence that this conductance is due predominantly to K+ is discussed. 5. Pre-pulse protocols under voltage clamp have been used to show that part of this conductance arises from the incompletely blocked delayed rectifier. 6. Substantial reductions in this conductance are produced by anaesthetics at 'clinical' concentrations. 7. It is concluded that there is a component of the K+ conductance of the resting squid axon other than the Hodgkin-Huxley delayed rectifier which is extremely sensitive to anaesthetics and which to an appreciable extent determines the resting potential.

4-Aminopyridine↗
PubMed1988

Excitation of the squid giant axon by general anaesthetics.

1. The effects of 'clinical' concentrations of some general anaesthetics on the minimum stimulus required to produce an action potential in the squid giant axon have been examined as a function of time from exposure to the anaesthetic. The resting potential in these experiments was also monitored. 2. The minimum stimulus varied with time in different ways for different anaesthetics. For chloroform, diethyl ether, n-pentanol, halothane and cyclopropane the stimulus initially declined, reached a minimum after about 3 min and then recovered to near-normal values at 10-15 min. For n-pentane, cyclopentane and, to a lesser extent methoxyflurane, the stimulus often declined to such low values that the axon exhibited spontaneous action potentials which persisted until the anaesthetic was removed. For one substance, the experimental local anaesthetic diheptanoyl phosphatidylcholine, the stimulus increased considerably over the 10-15 min required to reach the steady state. In all instances the axons reverted to normal behaviour after removal of the anaesthetic although the time course by which they did so was more variable than for the initial exposure. 3. For all anaesthetics the resting potential changed in the positive direction monotonically by ca. 1-5 mV and reached a steady state in approximately 3 min. On removal of the anaesthetic the resting potential returned to normal, also monotonically. 4. The voltage-gated Na+ and K+ currents were significantly affected even at the low anaesthetic concentrations used. Estimates of the changes in the Hodgkin-Huxley parameters were obtained partly by direct experiment and partly from results previously obtained for higher anaesthetic concentrations. 5. The time dependencies of the minimum stimuli have been accounted for semi-quantitatively in terms of the resting potential changes and the voltage shifts in the Na+ current steady-state activation, and the time dependencies respectively of these two parameters. 6. Quantitative calculations of the resting potential changes for comparison with experiment have been made based on the changes in K+ conductance determined in the preceding paper (Haydon, Requena & Simon, 1988) and changes in the Hodgkin-Huxley parameters of the Na+ and delayed-rectifier K+ currents. 7. Calculations of the minimum stimulus in the steady state have been made from the experimental resting potential changes and from the anaesthetic-affected Hodgkin-Huxley parameters. Good agreement with the experimental stimuli was found, especially in the prediction of high and low values.

Action Potentials↗
PubMed1987

The actions of halogenated ethers on the ionic currents of the squid giant axon.

The effects of fourteen halogenated ethers on the sodium and potassium currents of voltage-clamped squid giant axons have been examined. Effects under open-circuit were also studied. In voltage-clamped axons, the ethers tended to reduce potassium currents at least as much, if not more, than sodium currents. This finding distinguishes the halogenated ethers from many other general anaesthetics. Certain, but not all, halogenated ethers induced a pronounced maximum in potassium current traces as a function of time. This property can be formally described if an inactivation term is added to the Hodgkin-Huxley equation for potassium currents. Large shifts in the sodium-current inactivation parameter h infinity were produced in some instances. Two fully halogenated methyl ethyl ethers, known to produce convulsions in mice, depressed both sodium and potassium currents, but with a very slow time course of action. The electrophysiological effects of the halogenated ethers investigated appear to depend on the position and number of hydrogen bonds that can be formed.

Animals↗
PubMed1987

Local anaesthetic effects of benzene and structurally related molecules, including benzocaine, on the squid giant axon.

(1). The effects of benzene and several of its derivatives on sodium currents in the voltage-clamped squid giant axon have been studied. Substances tested were benzene, aniline, benzyl alcohol, propiophenone, 4-amino-propiophenone, methyl benzoate, ethyl benzoate, and 4-amino ethyl benzoate (benzocaine). (2.) All substances tested reduced the sodium current in both intact axons and axons internally perfused with CsF. (3.) There were four major actions of benzene on the sodium current: (a) an increase in the resting level of inactivation, (b) an increase in the depolarization required to produce the maximum current, (c) a decrease in the maximum sodium conductance, and (d) an increase in the rate of inactivation. (4.) 4-amino ethyl benzoate (benzocaine) had actions on the sodium current which were very similar to those of benzene with the exception that the rate of inactivation was scarcely affected and, at comparable shifts, the slope of the steady state inactivation curve was slightly smaller. (5.) The results obtained with the substances structurally intermediate between benzene and 4-amino ethyl benzoate allow some conclusions to be drawn as to the role of each functional group.

Anesthetics, Local↗
PubMed1987

The mechanisms of sodium current inhibition by benzocaine in the squid giant axon.

(1) The effects of benzocaine on the ionic currents in the voltage-clamped squid giant axon have been examined under various conditions; intact axons, axons internally perfused with CsF and axons dialysed with tetraethylammonium ions were used. (2) Both the steady state outward (potassium) current and the early transient (sodium) current were reduced by ca. 50% by benzocaine (1 mM). (3) Plots of the changes produced by benzocaine (1 mM) in the Hodgkin-Huxley parameters for the steady state activation (m infinity), the steady state inactivation (h infinity) and the time constants (tau m and tau h) for activation and inactivation of the sodium current are shown. The m infinity and h infinity curves are shifted in positive and negative directions respectively on the voltage axis. The time constants are not greatly affected. (4) In axons in which the sodium current inactivation had been largely removed by treatment with chloramine T, the sodium current was still reduced by ca. 50% by 1 mM benzocaine and the positive shift in activation remained unchanged. (5) The dependence on benzocaine concentration (for less than or equal to 2 mM) of the peak sodium current reduction and the shift in steady state inactivation have been determined. (6) It is concluded that in the squid axon the effects on inactivation are not the main reason for the reduction of the sodium current by benzocaine and that, in common with many other neutral anaesthetics, there are at least two sites at which benzocaine acts.

Animals↗
PubMed1987

Local anesthetic action of carboxylic esters: evidence for the significance of molecular volume and for the number of sites involved.

The effects of the homologous series of carboxylic esters, methyl propionate to methyl decanoate, on the steady-state inactivation of the sodium current in squid axons have been studied. The esters moved the relationship between the inactivation parameter, h infinity, and the membrane potential in the hyperpolarizing direction, thus reducing the number of sodium channels available at the resting potential. The concentration dependence of the shift at the mid-point of the curve of h infinity against potential has been measured for all esters except decanoate, which was almost inactive. Two aspects of these concentration dependences suggest that molecular volume is an important determinant of the effectiveness of each ester. Firstly, there is a sharp decline in activity above methyl hexanoate. This cut-off in activity resembles that for hydrocarbons where it has been suggested [e.g., Haydon, D.A., Urban, B.W. 1983, J. Physiol. (London) 341:411-427] to a result from a decrease in uptake with increasing molecular volume. (Further data for the hydrocarbons n-butane to n-heptane are reported here.) Secondly, the smallest compounds, methyl propionate and methyl butyrate, are less effective than would be predicted if equal membrane concentrations of each ester produced the same shift. The aqueous concentration dependences for these esters indicate that below methyl hexanoate, as the series is descended, progressively higher membrane concentrations are required to produce a given shift. This would be expected if the volume of ester in the membrane, rather than the number of molecules, is important.(ABSTRACT TRUNCATED AT 250 WORDS)

Anesthetics, Local↗
PubMed1986

Surface potential changes in lipid monolayers and the 'cut-off' in anaesthetic effects of N-alkanols.

The effects of 0.09 saturated solutions of the n-alkanols n-hexanol to n-tridecanol on the surface (compensation) potential of lipid monolayers have been examined. Actions on monolayers spread from pure egg phosphatidylcholine have been compared with effects on a system containing 2:1 mole ratio of egg phosphatidylcholine and cholesterol. The mean compensation potential for the pure phospholipid system was 475 +/- 9 mV; addition of cholesterol increased the potential to 503 +/- 10 mV. All n-alkanols tested reduced the surface potential in both systems. The reduction was larger in the pure phospholipid system but the difference in effect between lipid systems declined as the n-alkanol chainlength increased, becoming negligible by n-tridecanol. These results are considered in relation to the 'cut-off' in biological activity of n-alkanols around n-tridecanol.

Anesthetics↗
PubMed1986

The actions of some general anaesthetics on the potassium current of the squid giant axon.

A number of small organic molecules with general anaesthetic action have been examined for their effects on the voltage-dependent potassium current of the squid giant axon. They include representatives of the three classes of anaesthetics examined in previous studies on the sodium current (Haydon & Urban, 1983a, b, c), i.e. the non-polar molecules n-pentane, cyclopentane and CCl4, several n-alkanols and the inhalation anaesthetics chloroform, halothane, diethyl ether and methoxyflurane. Potassium currents under voltage clamp were recorded in intact and in intracellularly perfused axons before, during and after exposure to the test substances, and the records were fitted with equations similar to those proposed by Hodgkin & Huxley (1952). Shifts in the curves of the steady-state activation against membrane potential and reductions in the potassium conductance at 60 or 70 mV membrane potential have been tabulated. On the same intact axons, all the anaesthetics with the exception of methoxyflurane reduced potassium currents less than sodium currents by about a factor of two or more. For the n-alkanols, butanol to decanol, the concentrations required to reduce the potassium current at 60 mV membrane potential by 50% were determined. For n-butanol to n-heptanol, the standard free energy per CH2 for adsorption to the site of action was estimated to be -2.91 kJ mol-1 as compared with -3.04 kJ mol-1 for reduction of the sodium current. The magnitude of the free energy decreased for alkanols with longer chain lengths. At anaesthetic concentrations that reduce the sodium current by 50%, the hydrophobic substances n-pentane and cyclopentane reduced the maximal sodium conductance, gNa, and the potassium conductance at 70 mV, gK70, equally by about a third, while the n-alkanols reduced both parameters by less than 10%. By contrast, diethyl ether and methoxyflurane were more effective in reducing the maximal potassium conductance. All of the test substances examined, except n-pentane and n-hexane, shifted the voltage dependence of the potassium steady-state activation in the depolarizing direction. A broad qualitative correlation was found between the shifts in the activation curves for sodium and potassium currents but, quantitatively, the agreement between the two shifts was poor. In n-decanol and methoxyflurane solutions, the voltage-clamped potassium currents exhibited pronounced inactivation-like behaviour. These currents can be fitted by the Hodgkin-Huxley formalism if an inactivation term analogous to the sodium current inactivation is added.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1985

A quantitative explanation of the effects of some alcohols on gramicidin single-channel lifetime.

The effects of n-decanol, n-hexadecanol, n-octyl(oxyethylene)3 alcohol and cholesterol on gramicidin single-channel lifetime in planar lipid bilayers have been determined. The bilayers used were formed from a solution of monoolein in squalene. Measurements have also been made of the above compounds' effects on membrane thickness (as measured by electrical capacity and optical reflectance technique) and surface tension (as derived from bulk interfacial tension and bilayer-lens contact angle measurements). The reduction in single-channel lifetime caused by the n-alkanols may be accounted for quantitatively in terms of the effects of these compounds on bilayer thickness and surface tension. The n-octyl(oxyethylene)3 alcohol caused an increase in single-channel lifetime which is also consistent with the thickness/tension theory. The reduction in channel lifetime caused by cholesterol, however, was much larger than would be predicted from its effects on bilayer thickness and surface tension.

Alcohols↗
PubMed1985

Inactivation of the sodium current in squid giant axons by hydrocarbons.

The voltage dependence of the steady state inactivation parameter (h infinity) of the sodium current in the squid giant axon is known to be shifted in the hyperpolarizing direction by hydrocarbons and it has been suggested that the shifts arise from thickness changes in the axon membrane, analogous to those produced in lipid bilayers (Haydon, D. A., and J. E. Kimura, 1981, J. Physiol. [Lond.], 312:57-70; Haydon, D. A., and B. W. Urban, 1983, J. Physiol. [Lond.], 338:435-450; Haydon, D. A., J. R. Elliott, and B. M. Hendry, 1984, Curr. Top. Membr. Transp., 22:445-482). This hypothesis has been tested systematically by examining the effects of a range of concentrations of cyclopentane on the high-frequency capacitance per unit area both of the axonal membrane and of lipid bilayers formed from monoolein plus squalene. A similar comparison has been made for cyclopropane and n-butane, both at a pressure of 1 atm. The results are consistent with the notion that thickness increases in the axolemma produce the shifts in h infinity. Except at very high concentrations, however, the thickness changes in the lipid bilayer were too small to account for the h infinity shifts. A possible explanation of this finding is discussed.

Animals↗
PubMed1985

Further evidence that membrane thickness influences voltage-gated sodium channels.

The short-chain phospholipid, diheptanoyl phosphatidylcholine, at 520 microM, reduced the maximum inward sodium current in voltage-clamped squid giant axons by greater than 50%. Analysis of these currents by means of the Hodgkin-Huxley equations showed this reduction to be mainly the result of a large depolarizing shift in the voltage dependence of the steady state activation parameter, m infinity. The voltage dependence of the steady state inactivation parameter, h infinity, was also moved in the depolarizing direction and the axonal membrane capacitance per unit area measured at 100 kHz was increased. A longer chain length derivative, didecanoyl phosphatidylcholine, had no significant effect on the axonal sodium current at concentrations of 3.7 and 18.5 microM. Dioctanoyl phosphatidylcholine was intermediate in its effects, 200 microM producing approximately the same current suppression as 520 microM diheptanoyl phosphatidylcholine, together with depolarizing shifts in m infinity and h infinity. These effects may be contrasted with those of the normal and cyclic alkanes (1-3), which tend to move both m infinity and h infinity in the hyperpolarizing direction and to reduce the capacitance per unit area at 100 kHz. The above results are all consistent with the hypothesis that small hydrocarbons thicken, while short-chain phospholipids thin, the axonal membrane. Thus membrane thickness changes may be of considerable importance in determining the behavior of the voltage-gated sodium channel.

Animals↗
PubMed1985

The admittance of the squid giant axon at radio frequencies and its relation to membrane structure.

The admittance of the squid giant axon membrane has been measured, using an intracellular electrode, at frequencies up to 40 MHz. The existence of a radio frequency dispersion, previously detected with extracellular electrodes (Cole, 1976) and attributed to the Schwann cell layer, has been confirmed and followed to higher frequencies. For a comparable method of analysis, membrane parameters similar to those given by Cole (1976) have been calculated. The radio frequency dispersion has a centre frequency at approximately 1.8 MHz, and the properties of a parallel combination of a 28 nF cm-2 capacity and a 3.3 omega cm2 resistance. When the axon membrane capacity is calculated, taking into account the radio frequency dispersion, as described above, the capacity remains frequency dependent throughout the range studied. If it is assumed that at high frequencies the axolemma capacity becomes constant at approximately the value for a lipid bilayer, a radio frequency dispersion is found which cannot be accounted for in terms of a simple equivalent circuit with two passive components, but appears to arise from a network with a distribution of relaxation times. This result could be consistent with the morphology of the Schwann cell layer. The radio frequency dispersion referred to in (4) can be described reasonably well by a circuit with two dispersions having centre frequencies of 250 kHz and 3.2 MHz respectively. The corresponding axolemma capacity (100-500 kHz) would be approximately 0.6 microF cm-2. It is argued that between 50 and 100 kHz the geometrical capacity arising from the non-polar regions of the membrane is a major contributor to the axon membrane capacity, and that capacity variations arising from compositional changes in the lipid bilayer are best monitored in this frequency range.

Action Potentials↗