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R D Keynes

Publications and source records attributed to R D Keynes.

At least 55 records · Page 3Linked to original sources
PubMed1975

On the relation between displacement currents and activation of the sodium conductance in the squid giant axon.

The early time course of the current passing across the membrane in squid giant axons in which the ionic currents have been blocked reveals substantial asymmetries during and after the application of hyperpolarizing and depolarizing voltage-clamp pulses of identical size. Since the integral of the 'on' and 'off' current transients is zero, these currents must result from charge movements confined to the membrane and, therefore, they are nonlinear displacement currents. The steady state rearrangement of the charges as a consequence of sudden displacements of the membrane potential is consistent with a Boltzmann distribution of charges between two states characterized by different energy levels. Following changes in membrane potential the charges undergo a first order transition between these states. The relaxation time constant for the transition at a given temperature is a function of membrane potential. We propose that these displacement currents arise from a redistribution of the charges involved in the sodium gating system.

Action Potentials↗
PubMed1975

The ionic channels in excitable membranes.

The ionic channels in excitable membranes are of two classes: those that open and close when the membrane potential alters and those that respond to the release of an appropriate chemical transmitter. The former are responsible for the conduction of impulses in nerve and muscle fibres and the latter for synaptic transmission. It is now clear that the sodium and potassium channels in electrically excitable membranes are functionally distinct, since each can be blocked without affecting the behaviour of the other. It has recently proved possible to study, in the voltage-clamped squid giant axon, the movements of the mobile charges or dipoles that form the voltage-sensitive portion of the sodium channels, which give rise to the so-called 'gating' current. Detailed comparisons can now be made between the kinetics of the ionic conductances as described by Hodgkin & Huxley, and the steady-state distribution and kinetics of the charged controlling particles, which should lead to useful conclusions about the intramolecular organization of the sodium channels and the conformational changes that take place under the influence of the electric field. There is as yet little information about the chemical nature of the electrically excitable channels, but significant progress has been made towards the isolation and characterization of the acetylcholine receptors in muscle and electric organ.

Animals↗
PubMed1975

The heat production associated with the passage of a single impulse in pike olfactory nerve fibres.

1. A study has been made of the temperature changes associated with the passage of a single impulse in the non-myelinated fibres of the pike olfactory nerve. 2. The initial heat occurs in two phases: a burst of positive heat, followed by an evolution of negative heat. The positive and negative heats, and the net initial heat, are temperature-dependent. 3. At 0 degrees C the measured positive initial heat is 44.2 mucal/g.impulse; and the corresponding negative initial heat is 48.9 mucal/g.impulse. There is thus a net initial heat that is negative, of about 4.7 mucal/g.impulse. 4. The positive heat has a positive temperature coefficient, being increased by a factor of 1.86 when the temperature is rasied from 0 degrees C to 10 degrees C. 5. The negative initial heat also increases when the temperature is raised, but less than the positive initial heat. As a result, the net initial heat tends to become positive at higher temperatures. 6. Because of temporal dispersion of the action potential over the face of the thermopile, the observed temperature changes are smaller than those that occur at a single point in the nerve close to the stimulating cathode. The value of the positive heat at 0 degrees C corrected for temporal dispersion is estimated to be about 62 mucal/g.impulse: the corresponding value for the negative heat is about 67 mucal/g.impulse. 7. All records were analysed in terms of only two phases of initial heat (one positive, one negative). No analysis required four phases; but it is unclear whether this finding reflects a true absence of four phases, or merely the inability of the recording equipment to resolve them. 8. The positive heat seems to be derived from two sources. First, there is a dissipation of the free energy stored in the membrane capacity. Secondly, there is an evolution of heat corresponding with a decrease in entropy of the membrane dielectric with depolarization.

Action Potentials↗
PubMed1975

The effect of ouabain on ion transport across isolated sheep rumen epithelium.

1. The net flux of Na from the lumen to the blood side of isolated sheep rumen epithelium was reduced to zero in the third 30 min period after treatment with ouabain. 2. The net flux of K from blood to lumen side of the epithelium was reduced from 0-34 to 0-13 mumole/cm2. hr in the third 30 min period after ouabain treatment. 3. The net flux of Cl from lumen to blood side of the epithelium was reduced from 1-4 to 0-72 mumole/cm2. hr in the third 30 min period after ouabain treatment. 4. The results support the existence of a Cl pump independent of Na transport and also a K pump in sheep rumen epithelium.

Animals↗
PubMed1974

Kinetics and steady-state properties of the charged system controlling sodium conductance in the squid giant axon.

1. Asymmetries in the early time course of the displacement current passing across the membrane after application of equal voltage-clamp pulses in the two directions have been investigated in the squid giant axon. Before making the measurements, Na current was blocked by removal of external Na and treatment with tetrodotoxin. Potassium current was usually blocked by perfusion with CsF, but some experiments were done with intact axons. A signal averaging technique was used to eliminate the symmetrical components of the membrane current.2. The asymmetrical current had a contribution of appreciable size attributed to the movement of mobile charges or dipoles in the membrane. This was manifested as an outward current rising rapidly to a peak on depolarization of the membrane and then declining exponentially to zero, followed at the end of the pulse by an inward surge of current with a similar time course. There was also a sustained flow of current outwards during the pulse, arising from ionic leakage with a rectifying characteristic.3. The identification of the exponentially changing current component with the displacement of charged particles forming an integral part of the membrane was supported by the demonstration that the total transfer of charge was equal and opposite at the beginning and end of the pulse, that it reached saturation when the internal potential was taken to a sufficient positive value, and that its size was unaffected by temperature, although its time constant had a large temperature coefficient.4. The disposition of the mobile charges in the steady state was shown to obey a Boltzmann distribution. At the midpoint of the distribution curve, the proportion of the charge displaced underwent an e-fold change for a 19 mV change in potential. The effective valency of the particles, that is their actual charge multiplied by the fraction of the electric field acting on them, was therefore 1.3.5. The total quantity of mobile charge was estimated as about 1500 x 10(-12) C for 0.05 cm(2) of membrane, corresponding to some 1900 charges/mum(2).6. The identification of these mobile charges with the gating particles responsible for controlling Na conductance was supported by the findings that (a) their time constants were the same as those of Hodgkin & Huxley's ;m' system, both in absolute magnitude and in their dependence on potential and temperature, (b) the transition potential at which the charges were evenly distributed on the two sides of the membrane also agreed with that for the ;m' system in intact axons, and its value was similarly shifted in a positive direction by a reduction in internal ionic strength or by raising the external Ca concentration, (c) comparison of the steepness of the curves governing on the one hand the steady-state distribution of the mobile charges and on the other the Na conductance, suggested that an effective cooperation of the charges in groups of three was involved, again in excellent agreement with the ;m' system.7. Displacement of the mobile charges was unaffected by external pH over the range 5-8, but preliminary observations showed that 1% procaine reduced the total charge transfer to somewhat less than 40% of the initial value, and roughly halved the time constant.

Animals↗
PubMed1973

Calcium and potassium systems of a giant barnacle muscle fibre under membrane potential control.

1. Single barnacle muscle fibres from Megabalanus psittacus (Darwin) were internally perfused and the effects of various internal and external solutions on voltage clamp currents were examined.2. The usual internal solution was 180 mM-K(+) aspartate (osmotic pressure adjusted to 1000 m-osmole by adding sucrose). Fibres perfused with this solution gave an average resting potential of -55 +/- 5 mV (all potentials are referred to the external solutions as ground). Further increase in internal K concentration depolarized the fibres.3. With membrane current control the total capacitance, referred to apparent membrane surface area, was 21.2 +/- 2.4 muF/cm(2).4. Under normal conditions, with demonstrably good longitudinal space clamp control, voltage clamp currents associated with certain depolarizing pulses showed oscillations. These oscillations were reduced in frequency and magnitude by lowering the temperature from 20 to 10 degrees C, by eliminating the inward currents with external Ca-free saline or by reducing the outward currents with internal tetraethylammonium (TEA) or replacement of internal K by Cs.5. With a Na- and Ca-free, 60 mM-MgCl(2) solution outside depolarizing voltage clamp pulses produced only outward currents. On repolarization the current tail reversed direction at about -70 mV for pulses of less than 50 msec duration. For longer pulses this reversal potential was less negative, suggesting an accumulation of external, or depletion of internal K.6. Both the size of the outward currents and the rate at which they reached their maximum value increased with temperature. The activation energy for the rate constant was about 63 kJ/mole.7. Fibres bathed in Na- and Mg-free, 60 mM-CaCl(2) saline were excitable. After replacement of the internal K(+) with Cs(+) or adding 60 mM-TEA to the internal solution only sustained inward currents were recorded with depolarization.8. Sustained inward currents could be reduced by external application of 5 mM-LaCl(3). Tetrodotoxin was not effective even at a concentration of 1000 nM.9. The rate at which these inward currents reached a maximum value increased with increase in temperature of the bathing solution with an activation energy of the order of 42 kJ/mole.10. The reversal potential of the inward currents changed with the level of internal Ca ions. For a fibre perfused without ethyleneglycol-bis (beta-aminoethyl ether) N,N'-tetraacetic acid (EGTA) this reversal potential was 175 mV (internal free Ca 5 x 10(-7)M), and was 196 mV for a fibre perfused with 20 mM-Tris EGTA (internal free Ca 0.26 x 10(-8)M).11. We propose an electrical equivalent circuit to account for most of the observed electrical properties of barnacle muscle fibres. In this model the Ca and the K system are located at different anatomical places and they interact through a series resistance.

Animals↗
PubMed1972

Ion transport across an isolated preparation of sheep rumen epithelium.

1. The fluxes of isotopically labelled sodium, potassium and chloride passing in each direction across isolated sheets of rumen epithelium from the sheep have been measured under short-circuit conditions.2. With both sides of the epithelium bathed in chloride Ringer the mean sodium fluxes were 2.85 mumole/cm(2).hr from rumen to blood and 1.28 mumole/cm(2).hr in the reverse direction. In sulphate Ringer the sodium fluxes were 1.64 mumole/cm(2).hr from rumen to blood and 0.54 mumole/cm(2).hr from blood to rumen.3. In chloride Ringer the mean potassium fluxes were 0.18 mumole/cm(2).hr from rumen to blood and 0.54 mumole/cm(2).hr from blood to rumen. In sulphate Ringer the potassium fluxes were 0.07 mumole/cm(2).hr from rumen to blood and 0.35 mumole/cm(2).hr from blood to rumen.4. In chloride Ringer the mean chloride fluxes were 4.89 mumole/cm(2).hr from rumen to blood and 3.78 mumole/cm(2).hr from blood to rumen.5. In chloride Ringer the mean value of the short-circuit current was 13 muA/cm(2), corresponding to a flux of 0.49 muequiv/cm(2).hr. When sulphate was substituted for chloride, the short-circuit current was increased by about 40%, and the net flux of sodium from rumen to blood fell by 30%.6. Neither the sodium nor the chloride fluxes changed significantly when the epithelium was temporarily open-circuited.

Animals↗
PubMed1972

Changes in light scattering that accompany the action potential in squid giant axons: potential-dependent components.

1. To obtain information about structural events that occur in axons, changes in light scattering from squid giant axons were measured during action potentials and voltage-clamp steps.2. The scattering changes were measured at several scattering angles. Because the changes in scattering divided by the resting scattering were between 10(-6) and 10(-5), signal-averaging techniques were used to increase the signal-to-noise ratio.3. The scattering changes during the action potential were different at different angles. Two types were found, one at 10-30 degrees (forward angles) and the other at 60-120 degrees (right angles).4. At forward angles, there was a transient scattering decrease during the action potential. The time course of the change was similar to that of the action potential; this change was thought to be potential-dependent.5. At right angles, there was a transient scattering increase during the action potential followed later by a second, longer-lasting increase. Indirect evidence indicated that neither component could be totally potential-dependent.6. To further analyse these effects, scattering was measured during voltage-clamp steps. The changes seen during hyperpolarizing steps were presumed to be potential-dependent; again two different changes were found, one at forward angles and one at right angles.7. The potential-dependent change at right angles occurred with a time course that could be approximated by a single exponential with a time constant tau = 24 musec. The change at forward angles required two exponentials, tau(1) = 23 musec, tau(2) = 900 musec, to represent its time course.8. The size of both potential-dependent changes was proportional to the square of potential. The change at right angles, but not that at forward angles, was increased in size by the addition of butanol or octanol to the bathing solution.

Action Potentials↗
PubMed1972

Changes in axon light scattering that accompany the action potential: current-dependent components.

1. When light scattering was measured during hyperpolarizing and depolarizing voltage-clamp steps, relatively large scattering changes were found during the depolarizing steps. These large changes were found to depend on the time integral of the ionic current and not on the changes in conductance or potential.2. The current-dependent changes were examined at several scattering angles, and three distinct time courses were found. At 30-120 degrees , the main change occurred after the current when steps of 2-5 msec duration were used. This change was called I-90 degrees . At 15-30 degrees , the change occurred with the same time course as the time integral of the current. This change was called I-25 degrees . At 5-15 degrees the scattering change occurred with a time course intermediate between that of I-90 degrees and I-25 degrees . This change was called I-10 degrees .3. In all experiments, outward potassium and outward sodium currents led to similar light scattering changes indicating that specific effects of the cation carrying the current across the membrane were not involved.4. The size of I-90 degrees was reduced by 29% when an isethionate artificial sea water was substituted for the normal chloride artificial sea water. This reduction equalled the reduction predicted for a transport number effect at the membrane-solution interface. The time course of I-90 degrees was similar to the predicted time course for a volume change in the periaxonal space, and such volume changes were tentatively identified as the origin of I-90 degrees .5. Because of difficulties in measuring the time course of I-25 degrees , it was not possible to distinguish between a water of hydration effect and a transport number effect as the cause of this change. Similarly, the origins of I-10 degrees were not identified. Only I-10 degrees was altered in size and time course when the external refractive index was increased with bovine albumin.6. When the scattering changes during the action potential were examined in light of the voltage-clamp experiments, we concluded that the forward-angle change was potential-dependent and that the long-lasting change at right angles probably represented a swelling of the periaxonal space resulting from the fact that chloride carried a significant fraction of the outward current during the action potential.

Action Potentials↗
PubMed1971

Changes in light scattering associated with the action potential in crab nerves.

1. Changes in light scattering from stimulated crab leg nerves were measured in an effort to study changes in structure that occur during the action potential.2. The peak changes in scattering after individual stimuli were 10(-4)-10(-5) of the resting scattering; signal averaging techniques were used to increase the signal-to-noise ratio.3. During the compound action potential there was a transient increase in the 90 degrees scattering which reached a peak about 20 msec after the stimulus. This increase was probably not due to a change in axon volume. Its time course was somewhat slower than that of birefringence changes measured at the same time.4. In normal sea water the transient increase was followed by a longer lasting (persistent) decrease in scattering. This persistent decrease could be converted to an increase by reducing the tonicity of the sea water or by raising the external refractive index. This scattering change may represent a swelling of the axon resulting from the ionic exchanges that occur during the action potential.

Action Potentials↗
PubMed1971

The binding of tetrodotoxin to nerve membranes.

1. The% reduction in size of the externally recorded action potential produced by concentrations of tetrodotoxin (TTX) in the range 6-300 nM was determined for the small non-myelinated fibres of the rabbit cervical vagus nerve and of the walking leg nerves of crab and lobster. The concentration of TTX for 50% reduction was around 80 nM for rabbit vagus and 14 nM for crab nerve.2. Bio-assay procedures were devised to measure the amount of TTX taken up by a nerve when it was exposed to a very small volume of a solution whose TTX content was just great enough to produce 100% block of conduction. The extracellular space of each nerve was determined with [(14)C]sugar so that an allowance could be made for extracellular dilution.3. The TTX binding by rabbit, crab and lobster nerve was respectively 0.064, 0.053 and 0.036 p-mole/mg wet weight of nerve.4. The binding of saxitoxin was measured in rabbit vagus nerve, and found to be much the same as that of TTX.5. Control experiments on rabbit sciatic nerve, where the area of excitable membrane was much smaller, showed that there was relatively little unspecific binding of TTX.6. In view of the evidence presented here and elsewhere that the blocking of sodium conductance by TTX involves the attachment of only one TTX molecule at each sodium site, and that unspecific binding of TTX does not cause serious errors, these results suggest that in 1 mum(2) of nerve membrane the number of sodium sites is 75 for rabbit, 49 for crab, and 36 for lobster nerve.

Action Potentials↗
PubMed1971

Analysis of the potential-dependent changes in optical retardation in the squid giant axon.

1. An analysis has been made of the change in optical retradation of the membrane elicited by the application of voltage-clamp pulses in squid giant axons.2. The retardation response consists of three separate voltage-dependent components. For freshly mounted axons, defined as being in state 1, hyperpolarizing pulses give a rapid increase in the light intensity measured with crossed polarizers which has been termed the fast phase. This is followed by a rather slow return towards the base line termed the rebound. On treatment of the axon with certain agents that include tetrodotoxin, high calcium and terbium, the rebound disappears and the fast phase slows down, increases in size, and has a new slow component added to it. This transition from state 1 to a second state, 2, appears to be irreversible.3. In state 1, the time constant of the fast phase is 20-40 musec at 13 degrees C; it has a very large negative temperature coefficient (Q(10) = Ca.(1/8)). The size of the retardation change is independent of temperature and varies as the square of the applied voltage, but the voltage-retardation curve is symmetrical about a point well beyond zero membrane potential, at an internal potential of around + 70 mV. In state 2, the time constant is about five times larger, and varies much less markedly with temperature; the apex of the voltage-retardation curve is shifted to + 200 mV.4. The rebound has a time constant of the order of 20 msec at 13 degrees C. A 10 degrees rise in temperature more than halves the time constant and roughly doubles the amplitude of the rebound. The voltage dependence of the rebound differed from that of the fast phase.5. The slow component of state 2 has a time constant of about 2 msec which does not change noticeably between 10 and 25 degrees C. The size of this component seems to be linearly dependent on the applied voltage, rather than obeying a square law.6. A tenfold increase in external calcium concentration had no discernible effect on the fast and slow phases, but reversibly reduced the amplitude of the rebound nearly to half.7. In experiments on perfused axons, the retardation response was not measurably altered by any of the modifications made to the composition of the perfusing fluid.8. There was some indication of the possible existence of a small current- or conductance-dependent component of the retardation response.9. These phenomena seem likely to originate either from molecular relaxation processes analogous with the Kerr effect, or from changes in membrane thickness under the influence of the pressure exerted by the electric field. However, the specific molecules involved in the retardation response cannot yet be identified.

Animals↗
PubMed1970

Changes in axon birefringence during the action potential.

1. Observations have been made on the changes in optical retardation accompanying the passage of impulses along crab leg nerves and squid giant axons.2. The nerves were mounted on the stage of a polarizing microscope, at 45 degrees to the planes of polarization and analysis, brightly illuminated with white light. During the nerve impulse the intensity of the light passing the analyser decreased temporarily by 1 part in 10(3)-10(6). Signal-averaging techniques were used to obtain an acceptable ratio of signal to noise.3. The changes in light intensity recorded under these conditions were shown to arise almost entirely from alterations in retardation, with little or no interference from scattering, absorption, linear dichroism or optical rotation effects; the occurrence of stimulus and coupling artifacts was also ruled out.4. In the squid giant axon, the retardation change was shown to be located in a thin cylinder immediately surrounding the axoplasm, and to have a radially oriented optic axis.5. The time course of the decrease in optical retardation was very similar to that of the action potential recorded with an intracellular electrode, suggesting that the retardation closely followed the electrical potential across the membrane.

Action Potentials↗