Ca^<2+> Channels at the Presynaptic Terminal; Distributions and Functions

Ca^<2> 突触前末梢通道；

• 批准号: 17300126
• 负责人: KIDOKORO Yoshiaki
• 金额: $ 7.3万
• 依托单位: Gunma University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2006
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17300126/
• 关键词: synaptic transmission; Ca^<2+>channel; Drosophila; mutants; cacophony; Presynaptic terminal; neuromuscular junction; synapse; Ca^<2+>チャネル

1.Synaptic transmission in cacopholy(cac)-null mutant embryosIt has been demonstrated that Ca^<2+> influx through Drosophila N-type Ca^<2+> channels, encoded by the cacophony (cac) gene, triggers fast synaptic transmission. A remaining question is ; Is the cac Ca^<2+> channel the sole type of Ca^<2+> channel for fast synaptic transmission? Since the cac^<null> mutation is lethal, this question can only be answered by examining synaptic transmission in cac^<null> embryos. At the cac^<null> neuromuscular junction (NMJ), no fast synchronous synaptic transmission was detected upon nerve stimulation while infrequent delayed quantal synaptic events were observed. When the wild-type cac gene was introduced in the cac^<null> background, fast synaptic transmission recovered to 50% of heterozygous control embryos (cac^<null>/+). Thus, cac Ca^<2+> channels are necessary and sufficient for fast synaptic transmission. Unexpectedly, even in cac^<null> embryos nerve stimulation induced delayed releas … More e of synaptic vesicles in the minority of cells in HL3 solution (1.5 mM Ca^<2+>) and all cells examined in 5 mM [Ca^<2+>]_e. These delayed events started to appear at around 8 ms after stimulation, quickly increased in frequency to a peak at around 13 ms and gradually decreased to the baseline after 〜80 ms. The delayed events were abolished by 10 nM PLTXII (a spider toxin analog) suggesting that voltage-gated Ca^<2+> channels, other than cac Ca^<2+> channels, are contributing to the delayed release, but were not affected by 50 μM La^<3+> that preferentially blocks one type of Ca^<2+> channel in the Drosophila presynaptic terminal. Taken together, the cac Ca^<2+> channel is the sole Ca^<2+> channel for fast synaptic transmission and another type of Ca^<2+> channel, non-cac, PLTXII-sensitive Ca^<2+> channel, is contributing to delayed transmitter release in cac^<null> embryos.2.Distribution of cac Ca^<2+> channel clusters and the number of channels in each cluster at the presynaptic terminal.Using a Drosophila transformant that expresses EGFP-tagged cac Ca^<2+> channels, we have visualized the distribution of channel clusters at the presynaptic terminal. The number of Ca^<2+> in each cluster was estimated by examining the bleaching process of fluorescence intensity. In the line scan mode of confocal laser microscopy, the fluorescence intensity of a cluster declined due to bleaching. During this process, we found that the intensity declines in steps, which indicates a contribution of each EGFP molecule. By counting the number of steps we should be able to estimate the number of Ca^<2+> channels in each cluster. It turned out that the number is relatively small, around 10, in agreement with previous results in other systems obtained using completely different methods. This information is indispensable for simulation of the process of exocytosis at each release site. Less

1. 不和谐(cac)缺失突变体胚胎中的突触传递已证明，由不和谐(cac)基因编码的Ca^<2+>通过果蝇N型Ca^<2+>通道流入，触发快速突触传递。剩下的问题是； cac Ca^<2+> 通道是用于快速突触传递的唯一 Ca^<2+> 通道类型吗？由于 cac^<null> 突变是致命的，因此这个问题只能通过检查 cac^<null> 胚胎中的突触传递来回答。在cac^<null>神经肌肉接头(NMJ)处，在神经刺激时未检测到快速同步突触传递，而观察到罕见的延迟量子突触事件。当将野生型 cac 基因引入 cac^<null> 背景时，快速突触传递恢复到杂合对照胚胎的 50% (cac^<null>/+)。因此，cac Ca ^ 2+ 通道对于快速突触传递是必要且充分的。出乎意料的是，即使在 cac^<null> 胚胎中，神经刺激也会诱导 HL3 溶液 (1.5 mM Ca^<2+>) 中少数细胞的突触小泡延迟释放，并且所有细胞均在 5 mM [Ca^<2+>]_e 中检查。这些延迟事件在刺激后约 8 ms 开始出现，频率迅速增加，在 13 ms 左右达到峰值，并在〜80 ms 后逐渐下降至基线。延迟事件被 10 nM PLTXII（一种蜘蛛毒素类似物）消除，表明除 cac Ca^<2+> 通道之外的电压门控 Ca^<2+> 通道有助于延迟释放，但不受 50 μM La^<3+> 的影响，50 μM La^<3+> 优先阻断果蝇突触前末端的一种类型的 Ca^<2+> 通道。综上所述，cac Ca^<2+> 通道是用于快速突触传递的唯一 Ca^<2+> 通道，而另一种类型的 Ca^<2+> 通道（非 cac、PLTXII 敏感的 Ca^<2+> 通道）有助于 cac^<null> 胚胎中递质的延迟释放。 2. cac Ca^<2+> 通道簇的分布以及每个簇中通道的数量 突触前末端。使用表达 EGFP 标记的 cac Ca^<2+> 通道的果蝇转化体，我们可视化了突触前末端通道簇的分布。通过检查荧光强度的漂白过程来估计每个簇中Ca ^ 2+ 的数量。在共焦激光显微镜的线扫描模式下，簇的荧光强度由于漂白而下降。在此过程中，我们发现强度逐步下降，这表明每个 EGFP 分子的贡献。通过计算步数，我们应该能够估计每个簇中 Ca^<2+> 通道的数量。事实证明，这个数字相对较小，约为 10 个，这与之前使用完全不同方法在其他系统中获得的结果一致。该信息对于模拟每个释放位点的胞吐作用过程是必不可少的。较少的

项目成果

Gsα is involved in sugar perception in Drosophila melanogaster

• DOI: 10.1523/jneurosci.0857-06.2006
• 发表时间: 2006-06-07
• 期刊: JOURNAL OF NEUROSCIENCE
• 影响因子: 5.3
• 作者: Ueno, K;Kohatsu, S;Kidokoro, Y
• 通讯作者: Kidokoro, Y

Analysis of conditional paralytic mutants in Drosophila sarco-endoplasmic reticulum calcium ATPase reveals novel mechanisms for regulating membrane excitability

• DOI: 10.1534/genetics.104.031930
• 发表时间: 2005-02-01
• 期刊: GENETICS
• 影响因子: 3.3
• 作者: Sanyal, S;Consoulas, C;Ramaswami, M
• 通讯作者: Ramaswami, M

Synaptic Vesicle trafficking and recycling at the neuromuscular Junction; Two pathways for endocytosis.

神经肌肉连接处的突触小泡运输和回收；

• 发表时间: 2006
• 期刊: Drosophila Neuromuscular Junction (Book Chapter) (ed. V. Butnik and C. Ruiz) Elsevier 75
• 作者: Steinert JR;Kuromi H;Hellwig A;Knirr M;Wyatt AW;Kidokoro Y;Schuster CM;Kidokoro. Y.
• 通讯作者: Kidokoro. Y.

Nerve-evoked synchronous release and high K^+-induced quantal events are regulated separately by synaptotagmin I at Drosophila neuromuscular junction.

神经诱发的同步释放和高 K 诱导的量子事件由果蝇神经肌肉接头处的突触结合蛋白 I 分别调节。

• 发表时间: 2007
• 期刊: Journal of Neurophysiology 97
• 作者: Tamura;T.;Hou;J.;Reist;NE;Kidokoro.Y.
• 通讯作者: Kidokoro.Y.

External Ca2+ dependency of synaptic transmission in drosophila synaptotagmin I mutants.

• DOI: 10.1152/jn.00205.2005
• 发表时间: 2005-08
• 期刊: Journal of neurophysiology
• 影响因子: 2.5
• 作者: T. Okamoto;T. Tamura;Kazuhiro Suzuki;Y. Kidokoro