Botulinum toxin type A: kinetics of calcium dependent paralysis of the neuromuscular junction and antagonism by drugs and animal toxins

The effect of botulinum Toxin (BoTx), which blocks the mechanism of release of acetylcholine at neuromuscular junctions and induces paralysis of muscles stimulated by nerves, is known to be Ca 2+-dependent. Amplitude of muscular contractions evoked by nerve impulse was studied in BoTx poisoned preparations. The present report notes that an increase in Ca 2+ concentration in vitro delays paralysis of muscular contractions of the frog evoked by nerve impulse. The restorative effect of different drugs on this paralysis has been tested: 4-aminopyridine, ATXII (toxin isolated and purified from the sea anemone Anemonia sulcata tentacles) and a crude venom isolated from the scorpion Androctonus australis antagonize the BoTx induced paralysis at physiological concentrations of Ca 2+ (Ca 2+ 0 = 2 mM), whereas the restorative effect observed with tetra-ethylammonium or guanidine occurs at higher concentrations of Ca 2+ (Ca 2+ 0 = 4 mM), as in mammals. ATXII restores in vivo the activity of a BoTx paralysed muscle of guinea pig and this effect is more efficient if the interval between the injection of BoTx and ATXII is shortened. These results on the frog and guinea pig are in agreement with those obtained on other biological preparations by several investigators. Moreover it is suggested that the antagonism of BoTx induced paralysis is a consequence of the increase in Ca 2+ at the nerve ending. The efficiency of 4-aminopyridine and animal toxins is explained by an action on the nerve ending, by increasing Ca 2+ from an interval compartment of the cell, whereas antagonism produced by guanidine and tetraethylammonium involves uptake of Ca 2+ from the external medium. The bathing medium must be at a higher concentration of Ca 2+ than usual. This explains the differences in antagonism obtained by these drugs and toxins in vitro and in vivo.