Tetrodotoxin binding to normal depolarized frog muscle and the conductance of a single sodium channel.

Almers, Wolfhard; Levinson, S. Rock

doi:10.1113/jphysiol.1975.sp010943

Cited by 91 publications

(52 citation statements)

References 44 publications

(78 reference statements)

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“…nM-1) that was much smaller than in earlier experiments with [3H]TTX, allowing the saturable component to be readily identified in intact muscle. The value of M, which is about ten times the previously reported estimate from experiments with [3H]TTX, is similar in size to that of frog muscle (Almers & Levinson, 1975;Jaimovich, Venosa, Shrager & Horowicz, 1976;Ritchie et al 1976).…”

Section: Psupporting

confidence: 72%

Proceedings of the Physiological Society, 2‐3 July 1976, Cambridge Meeting: Communications

1976

The Journal of Physiology

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Section: Psupporting

confidence: 72%

Proceedings of the Physiological Society, 2‐3 July 1976, Cambridge Meeting: Communications

1976

The Journal of Physiology

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“…fibres is 535 + 43/11m2 (Hansen Bay & Strichartz, 1980). If, as in frog nerve, only half the sodium channels are open at the time of the peak current (Sigworth, 1978) (Almers & Levinson, 1975), and nerve (Conti, Hille, Neumcke, Nonner and Stampfli, 1976) which range from 1 to 8 p-mho.…”

Section: Discussionmentioning

confidence: 99%

Voltage‐clamp experiments in normal and denervated mammalian skeletal muscle fibres.

Pappone¹

1980

The Journal of Physiology

278

203

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SUMMARY1. The vaseline-gap voltage-clamp method has been applied to the study of ionic currents in cut pieces from innervated and 5-7 day denervated rat skeletal muscle fibres.2. Kinetic analysis of sodium currents in innervated rat muscle showed them to be similar to those in frog muscle, except that rat sodium channels are activated at slightly more negative potentials. Peak sodium conductances of 40-50 m-mho/cm2 were measured, corresponding to values of GNa of 100-120 m-mho/cm2.3. The permeability sequence of the sodium channel to several organic and inorganic cations is Li+ > Na+ > hydroxylammonium > hydrazinium > guanidiniumã mmonium > K+. TMA+ and Ca2+ were not measurably permeant.4. Denervation appears to shift activation and inactivation parameters of sodium currents by 10 mV to more negative potentials, but does not appreciably affect the maximum peak sodium conductance or the time constants for activation and inactivation.5. Dose-response curves for block by tetrodotoxin in innervated fibres are fitted well by assuming binding of toxin to a single population of channels with a dissociation constant of about 5 nm. In denervated fibres there appears in addition a second population of channels with a dissociation constant in the micromolar range. These relatively toxin-insensitive channels respond less rapidly to potential changes, and can contribute up to 25-30 0% of the total sodium conductance.6. The potassium currents of innervated rat muscle were similar to those of frog muscle in their voltage dependence of activation.7. The time constant for inactivation of the sodium current, Th, at -13 mV showed a temperature dependence measured between 10 and 20 'C equivalent to an average Q10 of 2*3. The Q1o for the time constant of activation of the potassium current, Tn, averaged 2-5 between -40 mV and + 40 mV, measured over the same temperature range.

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“…The number of Na channels in the electroplaque of the electric eel has been estimated in terms of the number of the binding sites for tetrodotoxin which was measured by a sensitive bio-assay Almers & Levinson, 1975) and by tritiated toxin binding (Reed & Raftery, 1976;Agnew et al 1978). Reported values range from 15 to 148 p-mole/g tissue.…”

Section: Discussionmentioning

confidence: 99%

“…However, the difficulty was largely overcome in the present experiments by making use of iodinated toxin with a high specific radioactivity (Catterall, 1977;Ray, Morrow & Catterall, 1978), and of electroplaque from the electric eel Electrophorus electricus as a source of membrane preparation. This material is shown to be enriched with Na channels on the basis of electrophysiological and pharmacological studies (35-91 mA/cm2 of peak Na current density; Nakamura, Nakajima & Grunfest, 1965 and 15-148 pmole/g tissue of the tetrodotoxin binding sites; Almers & Levinson, 1975;Reed & Raftery, 1976;Agnew, Levinson, Brabson & Raftery, 1978).…”

mentioning

confidence: 99%

Binding of scorpion toxin to sodium channels in vitro and its modification by β‐bungarotoxin

Okamoto

1980

The Journal of Physiology

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SUMMARY1. Binding of a purified scorpion toxin to membrane fragments isolated from electroplaque of an electric eel Electrophorus electricus was studied using a radioiodinated toxin.2. A scorpion toxin was purified from the venom of Leiurus quinquestriatus and iodinated with 125I in a lactoperoxidase-catalysed reaction. Monoiodinated toxin, isolated by an ion exchange chromatography, retarded the inactivation kinetics of Na current to a similar extent as the native toxin, indicating that radioiodination did not appreciably affect physiological and binding properties of the native toxin.3. Analyses of binding properties by Scatchard plots showed the presence of two classes of binding sites (with low and high affinities) in the membrane preparation from eel electroplaque; similar preparation from an electric skate, of which the electroplaque is known to be devoid of Na channels, possessed only the low affinity sites.4. The number of high affinity sites in the eel preparation was 41-8 + 10-5 p-mole/g tissue; the value was within the range reported for tetrodotoxin binding to similar preparations (15-148 p-mole/g tissue).5. A variety of cations (Na+, Mn2+ and La3+) inhibited the high affinity scorpion toxin binding, as indicated for the toxin binding to Na channels by a previous electrophysiological study. KD value in the presence of 120 mM-Na+ (approx. 8 nM) agreed reasonably with that (approx. 10 nM) reported for the scorpion toxin binding to excitable neuroblastoma cells or synaptic nerve ending particles under conditions where membrane potential was depolarized by the addition of 135 mM-KCl.6. Pretreatment of the eel membrane preparation with fl-bungarotoxin (7-44 ng/ ml.) in the presence of Ca ions (10-200 /M) resulted in a substantial loss of high affinity binding of scorpion toxin. When phospholipase A2 activity of the fl-toxin was inactivated by a chemical modification with p-bromophenacyl bromide, the inhibitory action of the ,-bungarotoxin was abolished.7. It is concluded that a high affinity binding of scorpion toxin to the eel electroplaque membrane fragments represents the binding to Na channels in vitro, and that phospholipase A2 activity of f8-bungarotoxin interferes with the binding of scorpion toxin to Na channels.

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Tetrodotoxin binding to normal depolarized frog muscle and the conductance of a single sodium channel.

Cited by 91 publications

References 44 publications

Proceedings of the Physiological Society, 2‐3 July 1976, Cambridge Meeting: Communications

Proceedings of the Physiological Society, 2‐3 July 1976, Cambridge Meeting: Communications

Voltage‐clamp experiments in normal and denervated mammalian skeletal muscle fibres.

Binding of scorpion toxin to sodium channels in vitro and its modification by β‐bungarotoxin

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