Abstract
Leptinotarsin, a toxin found in the hemolymph of the beetle Leptinotarsa haldemani, can stimulate release of acetylcholine from synaptic termini. Leptinotarsin causes an increase in the frequency of miniature end plate potentials (mepps) of the rat phrenic nerve-diaphragm preparation. The increase in the frequency of mepps induced by leptinotarsin is biphasic: about 10% of the total mepps are released in an initial burst that lasts about 90 sec, after which the remaining mepps are released over a period of 10-20 min. Tetrodotoxin has no effect upon the release induced by leptinotarsin, but low-Ca(2+) conditions abolish the first phase. The two phases of release may represent two presynaptic pools of acetylcholine, both of which can be released in quantized form. In a second study, rat brain synaptosomes were incubated with [(3)H]choline and were immobilized on Millipore filters. Leptinotarsin induced release of [(3)H]acetylcholine from this preparation, confirming the release seen by using neurophysiological methods. The ability of leptinotarsin to induce release from either intact nerve terminals or synaptosomes was abolished when the toxin was heated. The releasing activity of leptinotarsin from synaptosomes was also partially dependent upon the presence of Ca(2+) in the perfusing solution. Release from synaptosomes followed first-order kinetics, and was not inhibited by commercial antibodies to black widow spider antigens. The data suggest that leptinotarsin acts as a presynaptic neurotoxin and may be of value as a mechanistic probe in understanding the storage and release of neurotransmitters.