Hua S Y, Raciborska D A, Trimble W S, Charlton M P
Physiology Department, University of Toronto, Toronto, Ontario, M5S 1A8 Canada.
J Neurophysiol. 1998 Dec;80(6):3233-46. doi: 10.1152/jn.1998.80.6.3233.
Different VAMP/synaptobrevin complexes for spontaneous and evoked transmitter release at the crayfish neuromuscular junction. J. Neurophysiol. 80: 3233-3246, 1998. Although vesicle-associated membrane protein (VAMP/synaptobrevin) is essential for evoked neurotransmitter release, its role in spontaneous transmitter release remains uncertain. For instance, many studies show that tetanus toxin (TeNT), which cleaves VAMP, blocks evoked transmitter release but leaves some spontaneous transmitter release. We used recombinant tetanus and botulinum neurotoxin catalytic light chains (TeNT-LC, BoNT/B-LC, and BoNT/D-LC) to examine the role of VAMP in spontaneous transmitter release at neuromuscular junctions (nmj) of crayfish. Injection of TeNT-LC into presynaptic axons removed most of the VAMP immunoreactivity and blocked evoked transmitter release without affecting nerve action potentials or Ca2+ influx. The frequency of spontaneous transmitter release was little affected by the TeNT-LC when the evoked transmitter release had been blocked by >95%. The spontaneous transmitter release left after TeNT-LC treatment was insensitive to increases in intracellular Ca2+. BoNT/B-LC, which cleaves VAMP at the same site as TeNT-LC but uses a different binding site, also blocked evoked release but had minimal effect on spontaneous release. However, BoNT/D-LC, which cleaves VAMP at a different site from the other two toxins but binds to the same position on VAMP as TeNT, blocked both evoked and spontaneous transmitter release at similar rates. The data indicate that different VAMP complexes are employed for evoked and spontaneous transmitter release; the VAMP used in spontaneous release is not readily cleaved by TeNT or BoNT/B. Because the exocytosis that occurs after the action of TeNT cannot be increased by increased intracellular Ca2+, the final steps in neurotransmitter release are Ca2+ independent.
小龙虾神经肌肉接头处用于自发和诱发递质释放的不同VAMP/突触小泡蛋白复合物。《神经生理学杂志》80: 3233 - 3246, 1998年。尽管囊泡相关膜蛋白（VAMP/突触小泡蛋白）对于诱发神经递质释放至关重要，但其在自发递质释放中的作用仍不确定。例如，许多研究表明，切割VAMP的破伤风毒素（TeNT）会阻断诱发递质释放，但仍会保留一些自发递质释放。我们使用重组破伤风和肉毒杆菌神经毒素催化轻链（TeNT-LC、BoNT/B-LC和BoNT/D-LC）来研究VAMP在小龙虾神经肌肉接头（nmj）自发递质释放中的作用。将TeNT-LC注入突触前轴突可去除大部分VAMP免疫反应性，并阻断诱发递质释放，而不影响神经动作电位或Ca2+内流。当诱发递质释放被阻断>95%时，TeNT-LC对自发递质释放频率的影响很小。TeNT-LC处理后剩余的自发递质释放对细胞内Ca2+的增加不敏感。BoNT/B-LC与TeNT-LC在相同位点切割VAMP，但使用不同的结合位点，也能阻断诱发释放，但对自发释放的影响最小。然而，BoNT/D-LC在与其他两种毒素不同的位点切割VAMP，但与TeNT在VAMP上的结合位置相同，以相似的速率阻断诱发和自发递质释放。数据表明，诱发和自发递质释放使用不同的VAMP复合物；自发释放中使用的VAMP不易被TeNT或BoNT/B切割。由于TeNT作用后发生的胞吐作用不会因细胞内Ca2+增加而增强，神经递质释放的最终步骤是Ca2+非依赖性的。
