Selective modification of sodium channel gating in lobster axons by 2, 4, 6-trinitrophenol: Evidence for two inactivation mechanisms

Trinitrophernol (TNP) selectively alters the sodium conductance system of lobster giant axons as measured in current clamp and voltage clamp experiments using the double sucrose gap technique. TNP has no measurable effect on potassium currents but reversibly prolongs the time-course of sodium currents during maintained depolarizations over the full voltage range of observable currents. Action potential durations are increased also. Tm of the Hodgkin-Huxley model is not markedly altered during activation of the sodium conductance but is prolonged during removal of activation by repolarization, as observed in sodium tail experiments. The sodium inactivation versus voltage curve is shifted in the hyperpolarizing direction as is the inactivation time constant curve, measured with conditioning voltage steps. This shift speeds the kinetics of inactivation over part of the same voltage range in which sodium currents are prolonged, a contradiction incompatible with the Hodgkin-Huxley model. These results are interpreted as support for a hypothesis of two inactivation processes, one proceeding directly from the resting state and the other coupled to the active state of sodium conductance.     

Gating of cardiac Na+ channels in excised membrane patches after modification by alpha-chymotrypsin.

Single cardiac Na+ channels were investigated after intracellular proteolysis to remove the fast inactivation process in an attempt to elucidate the mechanisms of channel gating and the role of slow inactivation. Na+ channels were studied in inside-out patches excised from guinea-pig ventricular myocytes both before and after very brief exposure (2-4 min) to the endopeptidase, alpha-chymotrypsin. Enzyme exposure times were chosen to maximize removal of fast inactivation and to minimize potential nonspecific damage to the channel. After proteolysis, the single channel current-voltage relationship was approximately linear with a slope conductance of 18 +/- 2.5 pS. Na+ channel reversal potentials measured before and after proteolysis by alpha-chymotrypsin were not changed. The unitary current amplitude was not altered after channel modification suggesting little or no effect on channel conductance. Channel open times were increased after removal of fast inactivation and were voltage-dependent, ranging between 0.7 (-70 mV) and 3.2 (-10 mV) ms. Open times increased with membrane potential reaching a maximum at -10 mV; at more positive membrane potentials, open times decreased again. Fast inactivation appeared to be completely removed by alpha-chymotrypsin and slow inactivation became more apparent suggesting that fast and slow inactivation normally compete, and that fast inactivation dominates in unmodified channels. This finding is not consistent with a slow inactivated state that can only be entered through the fast inactivated state, since removal of fast inactivation does not eliminate slow inactivation. The data indicate that cardiac Na+ channels can enter the slow inactivated state by a pathway that bypasses the fast inactivated state and that the likelihood of entering the slow inactivated state increases after removal of fast inactivation.     

Effects of physostigmine on the voltage dependent ionic conductances of skeletal muscle fibres

The voltage dependent ionic conductances were studied by analysing the phase plane trajectories of action potentials evoked by electrical stimulation of the sartorius muscles of the frog (Rana esculenta). The delayed outward potassium current was measured also under voltage clamp conditions on muscle fibres of either the frog (Rana esculenta) or Xenopus laevis. On analysing the effect of physostigmine decreasing the peak amplitude, the rate of both the rising and falling phases of the action potentials, it was revealed that the alkaloid at a concentration of 1 mmol/l reduced significantly both the delayed potassium conductance and the outward ionic current values during the action potentials. The inhibition of sodium conductance and inward ionic current was less expressed. The maximum value of delayed potassium conductance measured under voltage clamp conditions was decreased by 1 mmol/l physostigmine. The time constant determined from the development of delayed potassium conductance was increased at a given membrane potential. The voltage vs. n relationship describing the membrane potential dependence of the delayed rectifier was not influenced by physostigmine. It has been concluded that physostigmine changes the time course of the action potentials by decreasing the value of both voltage dependent ionic conductances and by slowing down their kinetics. It is discussed that results obtained from the phase plane analysis of complex pharmacological effects can only be accepted with some restrictions.     

Ionic mechanism of action of a spin-labeled local anesthetic on squid axon membranes.

The ionic mechanism of action of a spin-labeled local anesthetic (SLA), 2-[N-methyl-N-(2,2,6,6-tetramethylpiperidonooxyl)]-ethyl 4-ethoxylbenzoate, was studied by means of voltage clamp technique with squid giant axons in comparison with the parent compound without spin label moiety, 2-(N,N-dimethyl)ethyl 4-ethoxylbenzoate (GS-01). Like other local anesthetics, they suppressed both sodium and potassium conductance increases. However, three remarkable differences have been noted between SLA and GS-01: (1) SLA is more effective than GS-01 in suppressing the sodium and potassium conductance increases; (2) SLA induces a potassium inactivation, whereas GS-01 is lacking this ability; (3) SLA has no effect on the time to peak sodium current, whereas GS-01 prolongs it. GS-01 resembles procaine with respect to (2) and (3) above. SLA will become a useful probe for the study of the molecular mechanism of local anesthetic aciton and of ionic channel function.     

Modelling action potentials and membrane currents of mammalian skeletal muscle fibres in coherence with potassium concentration changes in the T-tubular system

During prolonged activity the action potentials of skeletal muscle fibres change their shape. A model study was made as to whether potassium accumulation and removal in the tubular space is important with respect to those variations. Classical Hodgkin-Huxley type sodium and (potassium) delayed rectifier currents were used to determine the sarcolemmal and tubular action potentials. The resting membrane potential was described with a chloride conductance, a potassium conductance (inward rather than outward rectifier) and a sodium conductance (minor influence) in both sarcolemmal and tubular membranes. The two potassium conductances, the Na-K pump and the potassium diffusion between tubular compartments and to the external medium contributed to the settlement of the potassium concentration in the tubular space. This space was divided into 20 coupled concentric compartments. In the longitudinal direction the fibre was a cable series of 56 short segments. All the results are concerned with one of the middle segments. During action potentials, potassium accumulates in the tubular space by outward current through both the delayed and inward rectifier potassium conductances. In between the action potentials the potassium concentration decreases in all compartments owing to potassium removal processes. In the outer tubular compartment the diffusion-driven potassium export to the bathing solution is the main process. In the inner tubular compartment, potassium removal is mainly effected by re-uptake into the sarcoplasm by means of the inward rectifier and the Na-K pump. This inward transport of potassium strongly reduces the positive shift of the tubular resting membrane potential and the consequent decrease of the action potential amplitude caused by inactivation of the sodium channels. Therefore, both potassium removal processes maintain excitability of the tubular membrane in the centre of the fibre, promote excitation-contraction coupling and contribute to the prevention of fatigue. Received: 5 May 1998 / Revised version: 27 October 1998 / Accepted: 19 January 1999     

Slow Changes of Potassium Permeability in the Squid Giant Axon

A slow potassium inactivation i.e. decrease of conductance when the inside of the membrane is made more positive with respect to the outside, has been observed for the squid axon. The conductance-potential curve is sigmoid shaped, and the ratio between maximum and minimum potassium conductance is at least 3. The time constant for the change of potassium conductance with potential is independent of the concentration of potassium in the external solution, but dependent upon potential and temperature. At 9 degrees C and at the normal sea water resting potential, the time constant is 11 sec. For lower temperature or more depolarizing potentials, the time constant is greater. The inactivation can be described by modifying the Hodgkin-Huxley equation for potassium current, using one additional parameter. The modified equation is similar in form to the Hodgkin-Huxley equation for sodium current, suggesting that the mechanism for the passive transport of potassium through the axon membrane is similar to that for sodium.     

Slow sodium inactivation in nerve after exposure to sulhydryl blocking reagents

Exposure to N-ethylmaleimide (NEM), a reagent that binds covalently to protein sulfhydryl groups, results in a specific reduction in sodium conductance in crayfish axons. Resting potential, the delayed rise in potassium conductance, and the selectivity of the sodium channel are unaffected. Sodium currents are only slightly increased by hyperpolarizing prepulses of up to 50 ms duration, but can be restored to about 70% of their value before treatment if this duration is increased to 300-800 ms. The time to peak sodium current and the time constant of decay of sodium tail currents are unaffected by NEM, suggesting that the sodium activation system remains unaltered. Kinetic studies suggest that NEM reacts with a "slow" sodium inactivation system that is present in normal axons and that may be seen after depolarization produced by lowered the holding potential or increasing the external potassium concentration. NEM also perturbs the fast h inactivation system, and in a potential-dependent manner. At small depolarizations tauh is decreased, while at strong depolarizations it is increased over control values. Experiments with structural analogs of NEM suggest that sulfhydryl block is involved, but do not rule out an action similar to that of local anesthetics, p- Chloromercuriphenylsulfonic acid (PCMBS), another reagent with high specificity for SH groups, also blocks sodium currents, but restoration with prolonged hyperpolarizations is not possible.     

Simulation of the electrical activity of an excitable membrane with an electronic analog (author's transl)

1. The sodium and potassium conductances of the HODGKIN-HUXLEY model are simulated by a field effect transistor with a series resistor. This arrangement leads to a simple analog model of the excitable membrane (fig. 1 and 2). 2. Normally, the model is silent (fig. 3), but it becomes automatic (fig. 4) when the decay time (de-activation) of the potassium conductance is at least twice the recovery from inactivation time of the sodium conductance (taud greater than 2 tauri). 3. The effects of changes in sodium (fig. 5 and 6) and potassium (fig. 7, 8 and 9) concentration gradients upon the membrane potential and the ionic currents are easily studied when the model is silent or automatic. 4. When automatic, an increase in the potassium concentration gradient induces a lengthening of the period and ultimately, when the gradient is very high, spontaneous activity is blocked (fig. 9). On the other hand, increases of sodium gradient over 30% of normal value do not modify the period (fig 6). 5. The potassium concentration gradient modifies the excitability solely through membrane polarization (fig. 8), while sodium concentration has no effect on it (fig. 5). 6. Results with the model strengthen the hypothesis that tetraethylammonium (TEA) acts on both the maximum potassium conductance (gK) and the mechanism of sodium conductance inactivation (Tauh) to lengthen the action potential as observed on the Ranvier node (fig. 10). Effects of TEA on potassium conductance activation are also discussed. 7. Because of its simplicity and accuracy, this model lends itself easily to many other simulations.     

Gating of Na channels. Inactivation modifiers discriminate among models

Macroscopic Na currents were recorded from N18 neuroblastoma cells by the whole-cell voltage-clamp technique. Inactivation of the Na currents was removed by intracellular application of proteolytic enzymes, trypsin, alpha-chymotrypsin, papain, or ficin, or bath application of N-bromoacetamide. Unlike what has been reported in squid giant axons and frog skeletal muscle fibers, these treatments often increased Na currents at all test pulse potentials. In addition, removal of inactivation gating shifted the midpoint of the peak Na conductance-voltage curve in the negative direction by 26 mV on average and greatly prolonged the rising phase of Na currents for small depolarizations. Polypeptide toxins from Leiurus quinquestriatus scorpion and Goniopora coral, which slow inactivation in adult nerve and muscle cells, also increase the peak Na conductance and shift the peak conductance curve in the negative direction by 7-10 mV in neuroblastoma cells. Control experiments argue against ascribing the shifts to series resistance artifacts or to spontaneous changes of the voltage dependence of Na channel kinetics. The negative shift of the peak conductance curve, the increase of peak Na currents, and the prolongation of the rise at small depolarization after removal of inactivation are consistent with gating kinetic models for neuroblastoma cell Na channels, where inactivation follows nearly irreversible activation with a relatively high, voltage-independent rate constant and Na channels open only once in a depolarization. As the same kind of experiment does not give apparent shifting of activation and prolongation of the rising phase of Na currents in adult axon and muscle membranes, the Na channels of these other membranes probably open more than once in a depolarization.     

Voltage gated ionic channels in rat cultured astrocytes, reactive astrocytes and an astrocyte-oligodendrocyte progenitor cell

Astrocytes (both type 1 and type 2), cultured from the central nervous system of newborn or 7 day old rats show voltage gated sodium and potassium channels that are activated when the membrane is depolarized to greater than -40 mV. The sodium channels in these cells have an h-infinity curve similar to that of nodal membranes but the activation (peak current-voltage) curves are shifted along the voltage axis by about +30 mV. These sodium currents are blocked only by high concentrations of tetrodotoxin. The voltage activated potassium currents in both types of astrocyte show at least two components; an inactivating component that is suppressed at holding potentials of greater than -40 mV and a persistent, non-inactivating current. Several types of single channel currents were observed in outside-out membrane patches from type 2 astrocytes. One type of potassium channel showed inactivation on depolarization and may contribute to the whole-cell inactivating current. In contrast, oligodendrocytes showed no obvious voltage gated membrane channels. The properties of the type 2 astrocyte-oligodendrocyte progenitor cell were investigated in two ways: 1) by examination of cells just beginning to differentiate along the "electrically silent" oligodendrocyte pathway or 2) by recording from progenitor cells cultured for 24 hours in the presence of cycloheximide to block the appearance of new membrane channels. In both cases, voltage gated inward (sodium) and outward (potassium) currents were noted. The outward current response showed both an inactivating and a non-inactivating component. Similar voltage activated inward and outward membrane currents were noted in reactive astrocytes freshly isolated (3-6 hours) from lesioned areas of adult rat brains.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of N-alcohols on potassium conductance in squid giant axons

The effect of bath application of several short chain N-alcohols on voltage-dependent potassium conductance has been studied in intact giant axons of Loligo forbesi under voltage-clamp conditions. All tested alcohols (methanol, ethanol, propanol, butanol, heptanol and octanol) were found to depress potassium conductance only at concentrations much larger than those necessary to reduce sodium conductance. The efficacy of the different molecules was correlated with the carbon-chain length. In all cases the effects were found to be at least partly reversible. Low concentrations of propanol (100 mM) or heptanol (1 mM) were found to increase potassium conductance whereas higher concentrations had the usual depressing effect. The two alcohols were found to induce a slow inactivation of the potassium conductance. A detailed analysis of the time course of the turning-on of the potassium current for various pulse potentials in the presence of TTX revealed that, for membrane potential values more positive than -20 mV, the time constant of activation was reduced in the presence of propanol or heptanol. The delay which separates the change in potential and the turning-on of the potassium current, which was systematically analysed for different pulse and prepulse potential values, was increased by the two alcohols, the curve relating this delay to prepulse potential being shifted towards larger (positive) delays. This high degree of complexity in the effects on potassium conductance suggests that the alcohol molecules modify several more or less independent mechanisms associated with the turning-on of the potassium current.     

Effect of calcium ions on outward currents in the somatic membrane of mollusk giant neurons

Potassium currents through the somatic membrane of giant neurons ofHelix pomatia in normal (10 mM Ca) Ringer's solution and low-calcium (1 mM Ca) solution were studied by the voltage clamp method. With a decrease in the Ca concentration to 1 mM peak potassium conductance versus membrane, potential curves and inactivation curves were shifted along the voltage axis in the negative direction by about 10 mV. Inactivation of the delayed potassium current was slowed in low Ca solution. The effect of a decrease in external calcium concentration on volt-ampere and inactivation characteristics increased with a rise in external pH. These effects of a low Ca concentration on potassium mechanisms of the giant neuron somatic membrane can be attributed to changes in the negative surface potential in the region of the potassium channels.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Institute of Biology, Hungarian Academy of Sciences, Tihany. Translated from Neirofiziologiya, Vol. 8, No. 4, pp. 400–409, July–August, 1976.     

Opioids activate both an inward rectifier and a novel voltage-gated potassium conductance in the hippocampal formation

Opioid receptors were found to activate two different types of membrane potassium conductance in acutely dissociated neurons from the CA1/subiculum regions of the adult rat hippocampal formation. Opioid-responsive neurons were distinguished based on their morphology and electrophysiological responses. In one population of neurons having a multipolar, nonpyramidal cell shape, mu-selective opioid agonists increased an inward rectifying potassium current. Opioid activation of the inward rectifying conductance resulted in small outward potassium currents at resting membrane potentials and increased inward currents at hyperpolarized potentials. In a second population of nonpyramidal neurons, mu opioid agonists increased a novel voltage-gated potassium current. This current was blocked by internal CsCl2, unaffected by external BaCl2 or CdCl2, irreversibly activated by intracellular GTP-gamma-S, and inactivated by sustained depolarization. In contrast to the inward rectifying conductance, the voltage-gated conductance was not activated at resting membrane potentials or hyperpolarized potentials. The opioid-activated, voltage-gated conductance represents a new class of G protein-regulated potassium current in the brain.     

Effect of diethylamine analog of ethmozine on parameters of sodium current in isolated rat cardiomyocytes

Voltage clamp experiments were made on ezymically isolated and internally perfused rat cardiac cells. The effect of a diethylamine analog of ethmozine (DAAE) on sodium current (INa) was tested when the drug was applied inside or outside the cell. It was found that the effect of DAAE (8 X 10(-6) g/ml) on INa was asymmetrical: after DAAE addition outside the cell, the amplitude of INa was effectively suppressed. Thus, 5 minutes after DAAE action the maximal value of INa in a voltage-current relationship was 20% of the control value without significant changes in the kinetics of INa. When the DAAE was added inside the cell preferentially, the inactivation time constant was increased without significant changes in the amplitude of the maximal INa. The same results were obtained with pronase (1 mg/ml) added inside the cell. It was supposed that as compared to ethmozine, the DAAE possesses a supplementary binding site on the cardiac cell membrane possibly linked to the structures responsible for inactivation processes.     

A study of the crustacean axon repetitive response. 3. A comparison of the effect of veratrine sulfate solution and potassium-rich solutions

The crustacean single nerve fiber gives rise to trains of impulses during a prolonged depolarizing stimulus. It is well known that the alkaloid veratrine itself causes a prolonged depolarization; and consequently it was of interest to investigate the effect of this chemically produced depolarization on repetitive firing in the single axon and compare it with the effect of depolarization by an applied stimulating current or by a potassium-rich solution. It was found that veratrine depolarization, though similar in some respects to a potassium-rich depolarization of depolarizing current effect, was in many respects quite different. (1) At low veratrine concentration, less than 1 Mg%, the negative after potential following a spike action potential was prolonged and augmented. At higher concentrations or after a long period of time, veratrine caused a prolonged steady state depolarization of the membrane, the “veratrine response”. The prolonged plateau depolarization response could be elicited with or without an action potential spike by a short or long duration stimulating pulse, but only if the veratrine depolarization was prevented or offset by an applied conditioning hyperpolarizing inward current. (2) The “veratrine response” resembled the potassium-rich solution response in the plateau-like contour of the depolarization and the very low membrane resistance during this plateau phase. Like the potassium response, it was possible to obtain a typical hyperpolarizing response with an inwardly directed current pulse if applied during the plateau phase. During the negative after potential augmented with veratrine, however, this hyperpolarizing response was not observed. (3) In contrast to the potassium response, however, the “veratrine response” is intimately associated with the sodium concentration in the external medium. The depolarization in millivolts is linearly related to the log of the concentration of external sodium. Moreover, during veratrine action there is a continuous and progressive inactivation of the sodium mechanism which ultimately terminates repetitive firing and abolishes the spike action potential. Then even with conditioning hyperpolarization only the slow response may be elicited in veratrine, occasionally with a spike superimposed if sodium is present, but without repetitive firing. (4) It is concluded that veratrine action is the result of a chemical or metabolic reaction by the alkaloid in the membrane. It is suggested that veratrine may inhibit the sodium extrusion mechanism, or may itself compete for sites in the membrane with calcium and/or sodium. This explains the inhibiting effect of high calcium, the abolition of the “veratrine response” with low temperature and high calcium combined and the progressive inactivation of the sodium system.     

Inactivation of a volume-sensitive basolateral potassium conductance in turtle colon: effect of metabolic inhibitors.

Previous work has shown that the basolateral membrane of turtle colon epithelium contains a quinidine-sensitive potassium conductance which can be activated by osmotic cell swelling. In this work and in the present study, potassium flow across the basolateral membrane was measured as a short-circuit current across intact pieces of epithelial tissue in which amphotericin B was used to permeabilize the apical membrane. Quinidine-sensitive currents were generated when the mucosal bath contained chloride, a permeant anion. Replacement of chloride by sulfate or addition of sucrose to the bathing solutions abolished 75-90% of the current and caused the quinidine-inhibitable fraction of the current to go from over 90% to around 6%--suggesting that decreases in cell volume had brought about inactivation of the quinidine-sensitive conductance. When metabolic inhibitors were present, inactivation of the conductance by these maneuvers was prevented. Activation of the conductance by replacement of mucosal SO4 by Cl, however, was not affected.     

A voltage-dependent persistent sodium current in mammalian hippocampal neurons

Currents generated by depolarizing voltage pulses were recorded in neurons from the pyramidal cell layer of the CA1 region of rat or guinea pig hippocampus with single electrode voltage-clamp or tight-seal whole-cell voltage-clamp techniques. In neurons in situ in slices, and in dissociated neurons, subtraction of currents generated by identical depolarizing voltage pulses before and after exposure to tetrodotoxin revealed a small, persistent current after the transient current. These currents could also be recorded directly in dissociated neurons in which other ionic currents were effectively suppressed. It was concluded that the persistent current was carried by sodium ions because it was blocked by TTX, decreased in amplitude when extracellular sodium concentration was reduced, and was not blocked by cadmium. The amplitude of the persistent sodium current varied with clamp potential, being detectable at potentials as negative as -70 mV and reaching a maximum at approximately -40 mV. The maximum amplitude at -40 mV in 21 cells in slices was -0.34 +/- 0.05 nA (mean +/- 1 SEM) and -0.21 +/- 0.05 nA in 10 dissociated neurons. Persistent sodium conductance increased sigmoidally with a potential between -70 and -30 mV and could be fitted with the Boltzmann equation, g = gmax/(1 + exp[(V' - V)/k)]). The average gmax was 7.8 +/- 1.1 nS in the 21 neurons in slices and 4.4 +/- 1.6 nS in the 10 dissociated cells that had lost their processes indicating that the channels responsible are probably most densely aggregated on or close to the soma. The half-maximum conductance occurred close to -50 mV, both in neurons in slices and in dissociated neurons, and the slope factor (k) was 5-9 mV. The persistent sodium current was much more resistant to inactivation by depolarization than the transient current and could be recorded at greater than 50% of its normal amplitude when the transient current was completely inactivated. Because the persistent sodium current activates at potentials close to the resting membrane potential and is very resistant to inactivation, it probably plays an important role in the repetitive firing of action potentials caused by prolonged depolarizations such as those that occur during barrages of synaptic inputs into these cells.     

Photodynamic Alteration of Sodium Currents in Lobster Axons

Photodynamic alteration of lobster giant axons drastically changed the magnitude and kinetics of sodium currents seen under voltage clamp using the sucrose gap technique. Illumination of axons following treatment with acridine orange or eosin Y decreased the maximum sodium conductance to a zero asymptote as an exponential function of illumination time. Normal sodium inactivation was slowed, with τ_h more than doubled depending on experimental conditions. A second slower inactivation rate developed occasionally. τ_h was altered little, if at all. Sodium current "tails" were not prolonged. At maximum light intensity and with eosin Y as sensitizer leakage current increased after 4–10 sec in light. These changes were irreversible. Decreases in maximum sodium conductance correlated highly with increases in time to peak sodium current. The magnitude of change varied linearly with light intensity. The action spectra for eosin Y and acridine orange peaked near 545 and 505 nm, respectively. The magnitude of change varied with preillumination dye exposure time in a quasi-exponential approach to a maximum effect. Sodium dithionite protected the axon from photodynamic change.     

Multiple mechanisms of spike-frequency adaptation in motoneurones.

Spike-frequency adaptation is the continuous decline in discharge rate in response to a constant stimulus. We have described three distinct phases of adaptation in rat hypoglossal motoneurones: initial, early and late. The initial phase of adaptation is over in one or two intervals, and is primarily due to summation of the calcium-activated potassium conductance underlying the medium duration afterhyperpolarization (mAHP). The biophysical mechanisms underlying the later phases of adaptation are not well understood. Two of the previously-proposed mechanisms for adaptation are an increase in outward current flowing through calcium-activated potassium channels and increasing outward current produced by the electrogenic sodium-potassium pump. We found that neither of these mechanisms are necessary for the expression of the early and late phases of adaptation. The magnitude of the initial phase of adaptation was reduced when the calcium in the external solution was replaced with manganese, but the magnitudes of the early and late phases were consistently increased under these conditions. Partial blockade of the sodium-potassium pump with ouabain had no significant effect on any of the three phases of adaptation. Our current working hypothesis is that the magnitude of late adaptation depends upon the interplay between slow inactivation of sodium currents, that tends to decrease discharge rate, and the slow activation or facilitation of a calcium current that tends to increase discharge rate. Adaptation is often associated with a progressive decrease in the peak amplitude and rate of rise of action potentials, and a computer model that incorporated slow inactivation of sodium channels reproduced this phenomenon. However, the time course of adaptation does not always parallel changes in spike shape, indicating that the progressive activation of another inward current might oppose the decline in frequency caused by slow sodium inactivation.     

Effect of toxin 1 from Androctonus australis Hector on sodium currents in giant axons of Loligo forbesi

The effects of external application of micromolar concentrations of toxin 1 of the scorpion, Androctonus australis Hector, on the sodium conductance of squid giant axons have been studied quantitatively using the voltage clamp technique. Toxin concentrations which induce long plateau action potentials under current clamp conditions were found to simultaneously decrease the peak conductance and increase the delayed sodium conductance. Return to holding potential level after step depolarizations was accompanied by large exponential tails of current. The toxin-induced maintained sodium conductance increased with membrane depolarization independently of the peak conductance. Depolarizing conditioning prepulses to - 30 mV were found to almost totally inactivate the peak sodium current but to leave the delayed conductance unaffected. This property was taken as an indication that the total current is made of the added contributions of two distinct populations on sodium channels : fast activating and inactivating channels and slow activating channels. These two channel populations were separated from each other and analysed. It was found that the fast channels were almost identical to normal channels whereas the slow channels had a much slower (nearly exponential) kinetics and activated for more positive values of membrane potential. These observations strongly support the second hypothesis of Gillespie and Meves (1980) that the peak conductance and maintained conductance reflect the existence of two separate populations of channels. They further indicate that slow channels probably originate from the modification by the toxin of normal voltage-sensitive channels.