Gallamine triethiodide selectively blocks voltage-gated potassium channels in Ranvier nodes.

The effects of gallamine on ionic currents in single intact Ranvier nodes of the toad Xenopus were investigated. The following fully reversible effects were observed: 1. With a test concentration of 1 mmol/l the current-voltage relation of steady-state potassium currents, IK ss exhibited a complete block of IK ss up to about V = 110 mV; with stronger depolarisations the block was incomplete. The peak sodium currents, in contrast, were not affected. 2. At the same test concentration the potassium permeability constant PK was reduced by 92% from its normal value, while the sodium permeability constant PNa decreased by only 8%. 3. Concentration-response relations of the block of PK yielded an apparent dissociation constant of 30 micromol/l and a steepness parameter of unity. Patch-clamp experiments on cloned Kv1.1, Kv1.2, Kv1.3 and Kv3.1 channels yielded apparent dissociation constants of 86, 19, >>100 and 121 micromol/l, respectively. Our findings show that gallamine is particularly well suited for separating potassium and sodium currents in axonal current ensembles. They also strongly suggest that potassium currents in Ranvier nodes of Xenopus are mainly carried by an ensemble of Kv1.1 and 1.2 channels.

Effects of silperisone on the excitation process in Ranvier nodes.

The effect of silperisone on single intact Ranvier nodes of the toad Xenopus was investigated by adding it to the bathing medium. At 100 micromol/l the following fully reversible effects were observed: 1. The spike amplitude decreased in a frequency-dependent manner. 2. Both the sodium activation and the inactivation curves as well as the potential dependence of taum were slightly shifted in the negative direction, while tauh did not change. 3. The sodium permeability constant PNa decreased by 50%. 4. The potassium currents acquired a phasic time course previously described for certain psoralens. They reached a relative maximum and then approached a lower steady state value, kappa(infinity) with a time constant of about 5 ms. Concentration-related responses of PNa, PK and of k(infinity), yielded: 5. The apparent dissociation constant of block of PNa was 110 micromol/l. 6. PK proved not to be changed by silperisone in the concentration range tested, while the variable kappa(infinity) yielded a relation similar to that of PNa except that the apparent dissociation constant was 24 micromol/l. The phasic course of the potassium currents in the presence of silperisone may be due to an open channel blockade. In view of the similarities between the actions of silperisone and 5-methoxypsoralen, it is entirely conceivable that silperisone has potential for an antispastic drug, e.g., in demyelinating diseases like multiple sclerosis.

Tolperisone--a novel modulator of ionic currents in myelinated axons.

The actions of tolperisone on single intact Ranvier nodes of the toad Xenopus were investigated by means of the Hodgkin-Huxley formalism. Adding tolperisone to the bathing medium (100 micromol/l) caused the following fully reversible effects: 1. The sodium permeability P'Na was decreased by about 50% in a nearly potential-independent manner while the so-called sodium inactivation curve was shifted in the negative direction by about 3 mV. 2. The remaining parameters of the sodium system, i.e. m, taum and tauh, did not change. 3. The potassium permeability P'K decreased at strong depolarizing potentials (V > 60 mV); hence the permeability constant P(K) decreased by about 8%. However, weak depolarizations (V < 60 mV) caused P'K to increase by about 7%. 4. The potassium activation curve was shifted in the positive direction by about 9 mV and the exponent of n, b, was reduced from about 3.5 to about 1.5. Concentration-response relations for reduction of the sodium permeability constant PNa and of the potassium permeability constant P(K) yielded apparent dissociation constants of about 0.06 mmol/l and 0.32 mmol/l, respectively. The increase of P'K at V = 40 mV, however, was largely concentration-independent. Our findings show that, in contrast to the prevailing view, tolperisone cannot be said to have a so-called lidocaine-like activity, because its effect on potassium permeability in the threshold region is fundamentally different from that of other known local anaesthetics. We infer that this effect, in combination with the decrease in sodium permeability, is responsible for the tendency of tolperisone to reduce excitability and hence for the antispastic action of tolperisone documented by clinical observations.

Effects of three alkoxypsoralens on voltage gated ion channels in Ranvier nodes.

The effects of the phototoxic K+- channel blockers 8-methoxypsoralen (8-MOP) and 5-methoxypsoralen (5-MOP) on Ranvier nodes were compared to those of 5,8-diethoxypsoralen (5,8-EOP) by means of the Hodgkin-Huxley formalism. When these test substances were added individually to the bathing solution (8-MOP: 100 micromol/l; 5-MOP: 50 micromol/l; 5,8-EOP: 10 micromol/l) the following completely reversible effects were observed: 1. 8-MOP, caused a nearly potential-independent decrease of the sodium permeability, P'Na, by ca. 17%. 5-MOP and 5,8-EOP merely decreased the maximal value of P'Na, by ca. 12 and 8% respectively, whereas with weak depolarisations P'Na was unchanged. 2. In the tested potential range the potassium permeability, P'K, was caused to decrease by ca. 9% by 8-MOP, ca. 21% by 5-MOP and ca. 19% by 5,8-EOP. 3. The potassium currents acquired a phasic time course previously described for 8-MOP and 5-MOP. They reached a relative maximum and approached a lower steady-state value, kinfinity, with a time constant tauk at V = 120 mV of about 16 ms (8-MOP), 20 ms (5-MOP) and 94 ms (5,8-EOP). To obtain dose-response relations the drug-induced effects on peak P'K and on the steady state value, kinfinity, were measured. The corresponding apparent dissociation constants (in micromol/l) were 66.6 and 80.1 (for 8-MOP), 87.6 and 25.8 (for 5-MOP), and 13.5 and 6.5 (for 5,8-EOP). In view of the similarity of the actions of 5-MOP and 5,8-EOP as well as the fact that 5,8-EOP is not phototoxic, in future 5,8-EOP may well prove to be a particularly suitable K+-channel blocker for the symptomatic therapy of multiple sclerosis and other demyelinating diseases.

Reliability of ionic current measurements in Ranvier nodes.

A mathematical formalism was developed comprising four performance parameters to analyse electronic feedback systems for voltage clamp measurements in myelinated nerve fibres. The effects of the so-called Schmidt-Lanterman incisures and of the nodal series resistance were of particular interest here. Starting from passive standard data from myelinated nerve fibres, we calculated the frequency dependence of the above mentioned parameters for the measuring systems according to Nonner, to Dodge and Frankenhaeuser, and to Bohuslavizki and coworkers. It turned out that the latter system is clearly preferable to the two remaining systems for ionic current measurements in Ranvier nodes.

State-of-the-art potential clamp device for myelinated nerve fibres using a new versatile input probe.

A potential clamp device for myelinated axons is presented which, for the first time, systematically optimizes all the methodological parameters that limit the reliability of ionic current measurements. A crucial step toward this end consisted in the development of a new input probe with broad-band, extremely low-capacitance characteristics. In combination with a simple-to-use compensation criterion for optimum positive feedback, based on plausible assumptions, and with additional technological improvements, it enables the measurement of ionic currents with a reliability never previously achieved. The consequences with respect to evaluation of several earlier findings are discussed.

Mode of action of psoralens, benzofurans, acridinons, and coumarins on the ionic currents in intact myelinated nerve fibres and its significance in demyelinating diseases.

The actions of psoralens, benzofurans, acridinons and coumarins on the ionic currents in intact myelinated nerve fibres were investigated. All 6 substances blocked the potassium currents in a time-dependent manner, producing so-called K+ transients. Only 5-methoxypsoralen is a largely selective blocker of predominantly the axolemmal potassium channels, which is the characteristic required by our previously proposed working hypothesis for the mechanism of potassium-channel blockers in demyelinating diseases, in particular multiple sclerosis. If the observed K+ transients were to arise by blocking of the potassium channels of the Schwann cell, that is, by the periaxonal accumulation of K+ and a resulting collapse of the electromotive driving force for potassium-ions, according to a modified version of our previous hypothesis the other substances tested could also have a beneficial effect on the impaired impulse conduction in demyelinated axons. In this case a large number of new potential drugs would be available for the symptomatic therapy of MS.

Effects of some potassium channel blockers on the ionic currents in myelinated nerve.

The effects of some potassium channel blockers on the ionic currents and on the so-called K(+)-depolarization in intact myelinated nerve fibres were studied. 4-AP, and in particular, Flaxedil, proved to be selective K(+)-current blockers. However, TEA, a crown ether (DCH18C6), a longchained triethylammonium compound (C10-TriEA), capsaicin, and the extract from the medicinal herb Ruta graveolens proved not to be selective K(+)-current blockers; they all block Na(+)-currents as well, although to a lesser extent. The sodium inactivation curve did not change under TEA and Flaxedil but was shifted on the potential axis in negative direction by DCH18C6, 4-AP, capsaicin and the Ruta extract whereas C10-TriEA caused a shift of both sodium inactivation and activation parameters in positive direction. Regarding to the kinetics of the persisting K(+)-current fraction, two different kinds of blockade were found: 1. Unchanged K(+)-kinetic which is typical for the effects of TEA, 4-AP, Flaxedil, and C10-TriEA. 2. Clearly changed K(+)-kinetic, characterized by K(+)-transients; which is typical for the effects of capsaicin and in particular, for those of DCH18C6 and of the Ruta extract. The possibly different modes of action of both groups of blockers are discussed in terms of current models for the action of potassium channel blockers.

High standard one-loop potential clamp device for Ranvier nodes.

Starting from the observation that using a conventional potential clamp device for membrane current measurements in Ranvier nodes neither the kinetics of sodium currents nor the constant field concept agree satisfactorily with the Hodgkin-Huxley-Frankenhaeuser (HHF)-formalism, an extendend measuring system has been developed. The extensions introduced base largely on physical implications of myelinated nerve fibres which give rise 1. to systematic distortions of any current records at the high frequency end and 2. to current proportional deviations of the membrane potential from desired potential values. In addition, we provided to meet any unwanted current load during membrane current measurements and to push the time resolution of the measuring system to the highest possible value. After having tested thoroughly the new circuitry by appropriate physical methods, from sodium current measurements the following conclusions were drawn: 1. Occasional deviations of sodium current kinetics near the sodium equilibrium potential from the predictions of the HHF-formalism are measurement errors. 2. The constant field formalism holds for sodium currents in the potential range of biological relevance only. 3. Instantaneous sodium current measurements, however, are of unsatisfactory significance because for this kind of experiments the time resolution of the measuring system used might be still too low.

Inconsistent sodium current records derived on Ranvier nodes with a commercially available potential clamp device according to Nonner.

We tried to reproduce some basic implications of the Hodgkin-Huxley-Frankenhaeuser formalism by measuring sodium currents in single myelinated nerve fibres with a commercially available version of the potential clamp device according to Nonner. The following contradictory observations were made: 1. The potential dependence of the time to peak sodium currents showed a discontinuity around the sodium equilibrium potential. 2. Defining the sodium permeability PNa by the constant field equation and fitting the peak PNa-voltage relation by a sigmoid function we obtained unbelievable high values of PNa at rest. 3. Testing PNa as calculated by the constant field equation by so-called "sodium tail current" experiments we obtained instantaneous changes of PNa. Summing up, neither the kinetics of sodium currents nor the constant field concept as tested with the equipment used seem to agree satisfactorily with the standard data of sodium currents in Ranvier nodes.

[The effect of aminobenzoic acid derivatives on the ion current of myelinated neurons. 2. Inactivation of sodium permeability]. / Wirkung von Aminobenzoesäure-Derivaten auf die Ionenströme myelinisierter Nervenfasern. 2. Mitteilung: Inaktivierung der Natrium-Permeabilität.

Effects of Aminobenzoic Acid Derivatives on the Ionic Currents in Myelinated Nerve/2nd Communication: Sodium inactivation. Derivatives of aminobenzoic acid similar to metacaine were modified and their effects on the sodium inactivation and on the sodium permeability constant, PNa, of single potential clamped myelinated nerve fibres were tested. Comparing the observed shift of the inactivation curves and the reduction of PNa with the octanol/water partition coefficients of the drugs tested, a minimum lipid solubility seems to be necessary for the substances being efficacious. It is assumed that the uncharged aminobenzoic acids reduce PNa by disturbing the lipid matrix in an unspecific manner. The observed shift of the inactivation curves is explained in terms of the fixed charges at the inside of the membrane being moved apart by the uncharged compounds thus lowering the effective membrane potential. Moreover, no indications for a receptor-mediated mechanism were found.