Inhibition of neuromuscular transmission in isolated mouse phrenic nerve-diaphragm by the enterotoxin of Clostridium perfringens type A.

The enterotoxin of Clostridium perfringens type A, a channel forming protein toxin, inhibited neuromuscular transmission under conditions of low calcium. Twitch tension of isolated phrenic nerve-diaphragm preparations elicited by electrical stimulations to the phrenic nerve was recorded isometrically, and the preparations were exposed to the purified enterotoxin. In Krebs solution containing 0.5 mM calcium, the enterotoxin (20 micrograms/ml) reduced within 10 min the amplitude of the twitch tension to 34 +/- 7% (mean +/- S.D., n = 11) of that recorded before the treatment. The effects of the enterotoxin on the twitch tension were irreversible and proceeded independently of stimulation. The reduction of the twitch tension by the enterotoxin was apparent in Krebs solution containing less than 0.6 mM calcium and the degree of reduction was inversely related to the concentration of calcium. The reduction of the twitch tension by the enterotoxin was also dependent on temperature and concentration of the toxin. At temperatures below 20 degrees C, no obvious reduction of twitch tension was observed with 20 micrograms/ml of the enterotoxin. Enterotoxin at a concentration of 0.4 micrograms/ml caused 16 +/- 2% (mean +/- S.D., n = 4) reduction of twitch tension, and the degree of the reduction in twitch tension increased with toxin concentration, reaching a plateau of 65 +/- 4% (mean +/- S.D., n = 7) at 6.5 micrograms/ml of the enterotoxin. The effects of the enterotoxin were antagonized by 2 microM physostigmine. Unlike curare, pretreatment of the preparation with enterotoxin did not antagonize the neuromuscular block by decamethonium. Neither the tension of muscular twitch elicited by direct electrical stimulation to the muscle nor the resting membrane potentials of muscle fibers recorded intracellularly were affected by the enterotoxin. The enterotoxin (2.2 micrograms/ml) reduced the frequency, but not mean amplitude or amplitude distribution, of miniature end-plate potentials, from 0.91 +/- 0.07/sec to 0.72 +/- 0.07 (mean +/- S.E., n = 5). The results suggest that the enterotoxin will provide a novel tool for the studies on the mechanism of the neuromuscular transmission because of the unique characteristics of the inhibition and of the known mechanism of its action on the cell membrane.