A novel omega-conopeptide for the presynaptic localization of calcium channels at the mammalian neuromuscular junction.
A novel omega-conopeptide for the presynaptic localization of calcium channels at the mammalian neuromuscular junction.
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一种新型 omega-conopeptide，用于哺乳动物神经肌肉接头处钙通道的突触前定位。
DOI：
10.1007/bf01370157
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Ko,CP
中科院分区：
文献类型：
--
作者：
Sugiura,Y;Woppmann,A;Miljanich,GP;Ko,CP
电压敏感性Ca 2+通道对于化学突触处的递质释放是必不可少的。为了证明电压敏感性Ca ~（2+）通道与递质释放位点的关系，用α-银环蛇毒素和一种新型电压敏感性Ca ~（2+）通道探针SNX-260（ω-芋螺肽MVIIC的合成类似物）双标记小鼠神经肌肉接头。与ω-芋螺肽MVIIC类似，生物素化的SNX-260阻断小鼠神经肌肉接头处的神经刺激的递质释放。生物素化的SNX-260标记的神经末梢比乙酰胆碱受体簇更细，并由乙酰胆碱受体簇勾勒出轮廓。通过与未结合的SNX-260预孵育抑制SNX-260标记。神经肌肉接头的侧视图表明，SNX-260标记是在突触侧面对乙酰胆碱受体，而不是在非突触侧的神经末梢。这种突触前结合通过在胶原酶处理后神经切断或分离后失神经肌肉中缺乏SNX-260标记来证实。共聚焦显微镜显示SNX-260标记的斑点可能与活性区相关。SNX-260的标记模式不受未结合SNX-111（ω-芋螺肽MVIIA）（一种N型电压敏感性Ca 2+通道阻断剂）预孵育的影响。这些结果表明，SNX-260是一种新的探针定位非N型电压敏感性钙通道，这些电压敏感性钙通道定位在附近的递质释放位点在哺乳动物运动神经终末膜。这些结果与非N型、可能是P/Q型电压敏感性Ca ~（2+）通道介导哺乳动物神经肌肉接头处诱发递质释放的建议一致。
Voltage-sensitive Ca2+channels are essential to transmitter release at the chemical synapse. To demonstrate the localization of voltage-sensitive Ca2+channels in relation to the site of transmitter release, mouse neuromuscular junctions were double-labelled with α-bungarotoxin and a novel voltage-sensitive Ca2+channel probe, SNX-260, a synthetic analog of ω-conopeptide MVIIC. Similar to ω-conopeptide MVIIC, biotinylated SNX-260 blocked nerve-stimulated transmitter release at the mouse neuromuscular junction. Fluorescently-tagged biotinylated SNX-260 labelled the nerve terminal which appeared thinner than and was outlined by acetylcholine receptor clusters as seen inen faceview. This SNX-260 labelling was inhibited by preincubation with unconjugated SNX-260. Side-views of the neuromuscular junction indicated that the SNX-260 labelling was on the synaptic side facing the acetylcholine receptor rather than on the nonsynaptic side of the nerve terminal. This presynaptic binding was confirmed by the absence of SNX-260 labelling in denervated muscles following a nerve cut or disjunction after collagenase treatment. Confocal microscopy revealed spots of SNX-260 labelling that may correlate with active zones. The SNX-260 labelling pattern was not affected by preincubation with unconjugated SNX-111 (ω-conopeptide MVIIA), an N-type voltage-sensitive Ca2+channel blocker. These findings suggest that SNX-260 is a novel probe for localizing non-N type voltage-sensitive Ca2+channels and that these voltage-sensitive Ca2+channels are localized near the transmitter release sites at the mammalian motor nerve terminal membrane. The results are consistent with the suggestion that non-N, probably P/Q type voltage-sensitive Ca2+channels mediate evoked transmitter release at the mammalian neuromuscular junction.