The mechanism of beta-bungarotoxin on spontaneous transmitter release at developing neuromuscular synapse.

• Main Authors: Kai-Hsiang Kang, 康凱翔
• Other Authors: Jau-Cheng Liou
• Format: Others
• Language: zh-TW
• Published: 2003
• Online Access: http://ndltd.ncl.edu.tw/handle/33787669643968085726

碩士 === 國立中山大學 === 生物科學系研究所 === 91 === beta-Bungarotoxin (beta-BuTx), the presynaptic neurotoxin purified from the venom of Bungarus multicinctus, consists of two dissimilar polypeptide subunits. A phospholipase A2 subunit named A chain, and a non-phospholipase A2 subunits named B chain. The A chain and B chain are covalently linked by one disulfide bridge. Although it has been widely accepted that the toxic effect of beta-BuTx is attributed to the disturbance of presynaptic transmitter release, however the inhibition of transmitter release by beta-BuTx is still obscure. Here we investigate the mechanism that mediates facilitation of transmitter release at the neuromuscular junction induced by beta-BuTx, using Xenopus nerve-muscle coculture. Application of beta-BuTx and isotoxins BM12, BM13 led to a marked increase in the frequency of spontaneous synaptic currents (SSCs) after a short period (12~18 min) of latency. The synaptic potentiation induced by these toxins was abolished when Ca2+ in the medium is substituted by Ba2+ (a potent phospholipase A2 inhibitor). Application of PLP-BM12 and PLP-BM13, which have been chemical-modification to lose their PLA2 activity from BM12 and BM13, failed to potentiate the transmitter release. ��-BuTx-induced synaptic potentiation was not affect in the present of cycloxygenase inhibitor indomethacin and lipoxygenase inhibitor NDGA, indicating that COX and LPO metabolite was not involved in the potentiation. Bath application of XeC, a membrane-permeable inhibitor of inositol 1,4,5-trisphosphate (IP3) receptor, effectively occluded the increase of SSC frequency elicited by beta-BuTx. Treating cells with phospholipase C inhibitor U73122 or phosphatidylinositol 3-kinase inhibitors wortmannin and LY294002 abolishes beta-BuTx-induced synaptic potentiation. Furthermore, the potentiation effect of beta-BuTx is abolished while presynaptic loading with Non-hydrolyzable GDP analog GDP-beta-s. Taken together, these results suggest that beta-BuTx potentiates neurotransmitter secretion by stimulating Ca2+ release from IP3-sensitive receptor via G protein coupled, PI3 kinase and/or PLC signaling cascade.