Inhibition of neurotransmitter release in the lamprey reticulospinal synapse by antibody-mediated disruption of SNAP-25 function.
Inhibition of neurotransmitter release in the lamprey reticulospinal synapse by antibody-mediated disruption of SNAP-25 function.
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DOI：
10.1016/s0171-9335(99)80029-6
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发表时间：
1999-11
影响因子：
6.6
通讯作者：
P. Löw;T. Norlin;C. Risinger;D. Larhammar;V. Pieribone;O. Shupliakov;L. Brodin
中科院分区：
文献类型：
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作者：
P. Löw;T. Norlin;C. Risinger;D. Larhammar;V. Pieribone;O. Shupliakov;L. Brodin
七鳃鳗巨网状脊髓突触可通过突触前微量注射来操纵突触小泡吐出的分子机制。在这里，我们测试了扰乱SNARE蛋白SNAP-25功能的效果。体外实验表明，SNAP-25多克隆抗体可抑制Synaxin与SNAP-25的结合。当在突触前显微注射时，这些抗体对突触反应产生了强有力的抑制。突触前轴突中记录的Ba2+棘波没有改变，表明这种影响不是由于突触前钙进入减少所致。电子显微镜分析显示，抗体注射后的轴突突触内的突触囊泡结构与对照轴突相似，与突触前质膜明显接触的突触囊泡数量也相似。在注射SNAP-25抗体后，通常出现在受刺激突触周围质膜上的笼状蛋白包裹的凹坑没有被检测到，这与囊泡循环被阻断一致。因此，SNAP-25抗体破坏了与突触素的相互作用，抑制了神经递质的释放，而不影响质膜上突触小泡的数量。这些结果进一步支持了SNARE复合体的形成是膜融合的关键，而不是突触小泡靶向突触前膜的观点。
The lamprey giant reticulospinal synapse can be used to manipulate the molecular machinery of synaptic vesicle exocytosis by presynaptic microinjection. Here we test the effect of disrupting the function of the SNARE protein SNAP-25. Polyclonal SNAP-25 antibodies were shown in an in vitro assay to inhibit the binding between syntaxin and SNAP-25. When microinjected presynaptically, these antibodies produced a potent inhibition of the synaptic response. Ba2+spikes recorded in the presynaptic axon were not altered, indicating that the effect was not due to a reduced presynaptic Ca2+entry. Electron microscopic analysis showed that synaptic vesicle clusters had a similar organization in synapses of antibody-injected axons as in control axons, and the number of synaptic vesicles in apparent contact with the presynaptic plasma membrane was also similar. Clathrin-coated pits, which normally occur at the plasma membrane around stimulated synapses, were not detected after injection of SNAP-25 antibodies, consistent with a blockade of vesicle cycling. Thus, SNAP-25 antibodies, which disrupt the interaction with syntaxin, inhibit neurotransmitter release without affecting the number of synaptic vesicles at the plasma membrane. These results provide further support to the view that the formation of SNARE complexes is critical for membrane fusion, but not for the targeting of synaptic vesicles to the presynaptic membrane.