Madeleine Lemeignan

Evaluation of 4-aminopyridine and 3,4-diaminopyridine penetrability into cerebrospinal fluid in anesthetized rats

4-aminopyridine (4-AP) and 3,4-diaminopyridine (3,4-DAP) when injected intracisternally to anesthetized rats induced qualitatively similar central nervous system stimulant and convulsant effects at equimolar concentrations. Overall penetrability into cerebrospinal fluid of 4-AP is significantly higher than that of 3,4-DAP after single i.v. administration as evaluated by a high-performance liquid chromatographic determination. The present results can account for the lower central nervous system toxicity of 3,4-DAP when compared to 4-AP previously described after systemic administration.

Analysis of the action of 4-aminopyridine during repetitive stimulation at the neuromuscular junction

4-Aminopyridine (4-AP) increased the quantal content (m) of end-plate potentials (e.p.p.s) evoked by continual stimulation (0.2--25 Hz) in frog end-plates depressed by Mg2+. The increase in m was due to an increase in the binomial parameter n. This was interpreted to mean that 4-AP increased the number of activated release sites. In junctions blocked by d-tubocurarine, 4-AP first increased and then decreased the amplitude of e.p.p.s. elicited during a train of stimuli of increasing frequency, indicating that 4-AP increased transmitter release more than mobilization.

Inhibitory effect of kanamycin on evoked transmitter release. Reversal by 3,4-diaminopyridine.

The effect of kanamycin (Kn) on evoked transmitter release was examined in frog end-plates in vitro. By a presynaptic action, Kn (0.02 to 1 mM) significantly reduced the amount of acetylcholine liberated by nerve stimulation. In addition to its presynaptic effects, Kn (0.96 mM) decreased the size of the miniature end-plate potentials possibly by acting at the postsynaptic level. 3,4-Diaminopyridine (4.5 microM) reversed the presynaptic effects of Kn but did not modify its postsynaptic action.

Presynaptic effects of bekanamycin at the frog neuromuscular junction. Reversibility by calcium and aminopyridines

The effects of bekanamycin (5-320 microM) on evoked and spontaneous transmitter release were examined in frog neuromuscular junctions in vitro by using conventional electrophysiological techniques. Bekanamycin in a concentration-dependent fashion reduced reversibly the quantal content of the end-plate potentials while it had no observable effect on the configuration of the extracellularly recorded presynaptic action potential. The reduction in evoked transmitter release produced by bekanamycin could be antagonized either by increasing the external calcium concentration or by drugs like the aminopyridines which are to greatly enhance transmitter release from motor nerve terminals. Regarding the possible mode of action of bekanamycin on transmitter release our results are consistent with the hypothesis that there is competition between calcium and bekanamycin in the excitation-secretion coupling process and that the site of competition has a higher affinity for bekanamycin than for calcium. Bekanamycin also reduced in a dose-dependent manner the acceleration of miniature end-plate potentials induced by increasing the extracellular K+ concentration while it had no effect on spontaneous miniature end-plate potential frequency in resting junctions. Besides its inhibitory pre-junctional effects bekanamycin in concentrations higher than 40 microM decreased dose-dependently the amplitude of miniature end-plate potentials. However, presynaptic effect predominated and seemed to be quantitatively more important for the neuromuscular block than was a possible postsynaptic action of the antibiotic. It is concluded that bekanamycin exerts potent inhibitory effects on transmitter release probably by interfering with the influx of calcium that occurs during depolarization of motor nerve terminals.

Enhancement by Anemonia sulcata toxin II of spontaneous quantal transmitter release from mammalian motor nerve terminals

The action of Anemonia sulcata toxin II (ATX-II) on spontaneous quantal transmitter release from motor nerve terminals was investigated by recording miniature end-plate potentials (MEPPs) from isolated mouse phrenic nerve--hemidiaphragm nerve--muscle preparations. ATX-II (3.2 microM) when applied for 3-40 min to junctions bathed in a normal ionic medium enhanced about one hundred fold the rate of spontaneous MEPPs. Concomitantly, ATX-II depolarized the muscle fiber. The effect of the toxin on MEPP frequency was markedly reduced when junctions were exposed to Na-deficient solutions or pre-treated with dantrolene sodium (10 microM). ATX-II (0.24-3.2 microM) increased MEPP rate in junctions exposed to a Ca-free medium containing 2 mM EGTA and 2 mM Mg2+ in a dose- and time-dependent manner. Tetrodotoxin (0.2-1 microM) prevented the effects of ATX-II on MEPP frequency and on the resting membrane potential of muscle fibers. Tetrodotoxin also antagonized the acceleration of MEPP induced by ATX-II. The experimental findings suggest that ATX-II acts to increase quantal transmitter output from motor nerve terminals by enhancing Na+ influx through tetrodotoxin-sensitive presynaptic channels, since ATX-II action does not appear to depend upon entry of Ca2+ from the extracellular medium. It is likely that ATX-II, by increasing intraterminal Na+ concentration, may trigger calcium release from internal stores.

Effects of toxin II from the sea anemone Anemonia sulcata on contractile and electrical responses of frog skeletal muscle fibres

The effects of Anemonia sulcata toxin II (ATX-II) were studied on mechanical and electrical activities of frog muscle fibres isolated from semitendinosus or tibialis anterior muscles of Rana temporaria (2.8-7.7 degrees C). In concentrations ranging between 7.7 and 100 microM, ATX-II greatly potentiated the isometric twitch of single muscle fibres in a time-dependent manner. Increase in twitch amplitude by ATX-II was associated with an increase in time to peak tension and time from peak tension to half relaxation. ATX-II caused no change in maximum force production during fused tetanus, but the tension was maintained for several seconds after the cessation of stimulation. Such long tetanic contractions were also obtained in low-Na Ringer solution, but their duration was somewhat shorter. No specific action of ATX-II was detected on relaxation kinetics during a tetanus. The twitch potentiating effect of ATX-II was markedly increased by 3,4-diaminopyridine. Action potentials recorded from single muscle fibres in the presence of ATX-II showed a delayed onset of repolarization with a reduced rate of fall. In addition, ATX-II caused repetitive spontaneous firing of action potentials after the cessation of tetanic stimulation. ATX-II (3.3 - 10 microM) also increased action potential duration by reducing the rate of repolarization in surface fibres of formamide-treated sartorius or cutaneous pectoris muscles (20 degrees C) stimulated indirectly or directly. The potentiation of twitch force and the prolongation of contractions caused by tetanic stimulation can be attributed to the membrane action of ATX-II, which leads to prolongation of action potentials, to repetitive muscle firing and to the appearance of plateau potentials.

Facilitatory effects of 4-aminopyridine on strontium-mediated evoked and delayed transmitter release from motor nerve terminals.

The effect of 4-aminopyridine (4-AP) on Sr-mediated evoked and delayed transmitter release at the frog neuromuscular junction was examined using conventional electrophysiological techniques. 4-AP (5-50 microM) increased transmitter release evoked in response to conducted nerve impulses or to electrotonic depolarization of tetrodotoxin (TTX)-treated motor nerve terminals. Spontaneous quantal transmitter release was not affected by the drug as judged by the lack of effect on miniature end-plate potentials (MEPPs) frequency. However, 4-AP (10-20 microM) markedly enhanced the frequency of MEPPs appearing during conducted low rate repetitive nerve impulses or single electrotonic depolarization of TTX-blocked nerve terminals. The results suggest that 4-AP increases the influx of Sr2+ into nerve terminals in a way that modifies its sequestration and removal by the subcellular elements of the terminal.

Effects of 4-Hydroxypyridine on Transmitter Release at the Neuromuscular Junction

Publisher Summary This chapter discusses the results to analyze the effects of 4-hydroxypyridine (4-HP) on transmitter release at the neuromuscular junction. The results obtained in this study show that 4-HP (1–10 mM) greatly increased stimulus evoked transmitter release in junctions in which the normal release of transmitter was reduced by increasing extracellular Mg ++ and reducing Ca ++ . Similarly 4-HP dose dependently was able to restore from complete blockade neuromuscular transmission in curarized preparations when stimulated at low frequency. However, during titanic stimulation, only the first response in the train was facilitated, while the successive responses were depressed in comparison to the controls. 4-HP had no effect on the time course of the synaptic potentials and there was no evidence that the drug in the concentrations used had any postsynaptic effect. Focal extracellular recordings revealed that 4-HP increased the latency of the evoked responses and that this effect was mainly because of a prolongation of the synaptic delay. Spontaneous quantal release was unaffected by 4-HP in resting junctions or in junctions depolarized by increasing the external K + concentration.

4-Aminopyridine as Antagonist of the Cardiovascular and Neuromuscular Depressant Effects of Kanamycin in Rats

Publisher Summary 4-AP has been shown to antagonize the neuromuscular blockade produced by some aminoglycoside antibiotics in vivo and in vitro. This chapter discusses a study to analyze 4-aminopyridine (4-AP) as antagonist of the cardiovascular and neuromuscular depressant. This study was undertaken to study if 4-AP could antagonize both the cardiovascular and neuromuscular depressant effects of kanamycin (KM) in anesthetized and conscious rats. For this purpose, in anesthetized rats, directly and indirectly elicited maximal twitch tensions of the tibialis anterior muscle in situ were recorded together with the mean arterial blood pressure (AP) and heart rate (HR). Kanamycin infusion induced progressively a decrease in twitch tension and in HR and AP. 4-AP raised significantly the KM doses inducing cardiovascular or neuromuscular depressions. KM infusion to chronically catheterized conscious rats induced a spontaneous head drop that was taken as a sign of the neuromuscular block and a decrease in HR and AP. 4-AP also antagonized these effects of KM but was less active than in anesthetized rats.