Synaptosomes isolated from the rat brain corpus striatum and locust head and thoracic ganglia were loaded with radioactive neurotransmitter ([3H]dopamine and [3H]acetylcholine, respectively) and then treated with α-latrotoxin and other fractions (fractions C, D and E of Frontali et al.8) obtained by Sephadex G200 column chromatography from black widow spider venom gland homogenates. As shown by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, α-latrotoxin is a high Mr protein, whereas fractions C-E are mixtures of several proteins, that include small amounts of contaminating α-latrotoxin (especially in fraction C). In rat synaptosomes α-latrotoxin induced massive neurotransmitter release, and some release was induced also by high concentrations of fractions C and D. These responses were blocked almost completely by a monospecific anti-α-latrotoxin serum, indicating that they were all due to α-latrotoxin. Release of [3H]acetylcholine from locust synaptosomes was induced by the various preparations investigated. α-Latrotoxin was about 10-fold less potent in locust than in rat synaptosomes. The effects of fractions C-E tended to disappear with storage. The most active batches of fractions C and E were even more potent than α-latrotoxin, while the D fraction was approximately 5-fold less potent. The anti-α-latrotoxin antiserum inhibited part of the responses elicited by fractions C and E, but left fraction D almost unaffected. Release by D and E fractions was maintained even when Ca2+ was removed from the incubation medium. It is concluded that locust nerve endings are sensitive not only to α-latrotoxin, but also to other venom components: a high Mr toxin (fractions C, D), for which the name of β-latrotoxin is proposed; and, possibly, a third toxin, recovered in the E fraction, that might be immunologically related to α-latrotoxin.
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