Molecular Mechanism on the Block of Neurotransmitter Release with Inositol Polyphosphates

我们以前证明，突触结合蛋白的C2B结构域是一个肌醇多磷酸（IP）的结合位点，并在鱿鱼巨突触的突触传递完全阻断注射到突触前末梢的IP。这些结果表明，突触结合蛋白是一种直接参与神经递质释放的功能分子，是IP阻断突触传递的靶分子。1995年，为了验证这一想法，我们研究了C2B结构域的特异性抗体（该抗体抑制IP4与C2B结构域的结合）是否能够使用鱿鱼巨突触释放IP对神经传递的阻断。结果表明，IP和抗体共同注射可完全解除突触传递阻滞。此外，注射抗体后高频率刺激，突触反应迅速降低，突触囊泡明显减少。这些结果有力地表明，synaptotagmin是一个目标分子在阻断神经传递的IP，并参与这两种现象的胞吐和胞吞。在1996年，我们进行了类似的研究，在细胞水平上使用原代培养的哺乳动物细胞，大鼠上级颈神经节神经元（电生理方法）和牛肾上腺嗜铬细胞（儿茶酚胺释放）。结果表明，IPs的阻断效应和与C2B抗体共注射后的阻断释放现象相同。这些结果强烈表明，IP是普遍存在的调制器的突触结合蛋白的物种。

We previously demonstrated that the C2B domain of synaptotagmin is an inositol polyphosphates (IPs) binding site, and synaptic transmission in squid giant synapse is completely blocked by injection of IPs into the presynaptic terminal. From these results, we suggested that synaptotagmin is a functional molecule directly involved in the neurotransmitter release, and is a target molecule of IPs in the block of synaptic transmission. In 1995, in order to verify this idea, we investigated whether specific antibody to the C2B domain (this antibody inhibits binding of IP4 to the C2B domain) is able to release the block of neurotransmission by IPs using squid giant synapse. The result revealed that the block of synaptic transmission is completely released with coinjection of IPs and the antibody. Moreover, with high frequent stimulation after injection of the antibody, synaptic response rapidly reduced, and drastical reduction of synaptic vesicle was observed. These results strongly suggest that synaptotagmin is a target molecule in the block of neurotransmission with IPs, and is involved in both phenomenons of exocytosis and endocytosis. In 1996, we performed a similar study in cell level using the primary culture from mammalian cells, rat superior cervical ganglion neurons (electrophysiological approarch) and bovine adrenal chromaffin cells (catecholamine release). The results demonstrated that the same phenomenons were observed on the blocking effect by IPs and release of block by coinjection with the C2B antibody. These results strongly suggest that IPs are ubiquitous modulator of synaptotagmin over the species.