| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2005: ¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 2004: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Research Abstract |
When an action potential reaches the nerve terminal, Ca^<2+>-influx through open voltage-sensitive Ca channels triggers the exocytosis of synaptic vesicles, thereby releasing neurotransmitter molecules. Recent studies indicate that synaptic SNARE proteins (syntaxin, SNAP-25 and VAMP/synaptobrevin) are the minimal machinery for the synaptic vesicle fusion with the presynaptic membrane. Cytosolic proteins including NSF,SNAPs,nSec1/Munc18-1 and synaphin/complexin regulate the fusion. We have used the neuromuscular junction of Drosophila larvae to examine the localization of these proteins involved in the synaptic vesicle cycle and its regulation. Localization of these proteins was determined by immunofluorescence after fixation. In resting conditions, syntaxin was distributed along the internal surface of the nerve terminal with some densely stained regions. Synaphin localization was very similar to that of syntaxin. After exocytosis induced by high K^+-treatment, syntaxin localization did not significantly differ from the resting state. However, synaphin distribution became diffuse over the cytoplasm, suggesting its dynamic movement during or after exocytosis. Sibire^<ts> mutant also showed the dynamic movement of synaphin at a restrictive temperature where endocytosis of synaptic vesicles was completely suppressed without affecting exocytosis. Thus the dynamic movement of synaphin is related to synaptic vesicle exocytosis but not to endocytosis. The movement was blocked by the casein kinase II (CKII) inhibitors (5,6-dichlorobenzimidazole riboside and 4,5,6,7-tetrabromotriazole) but not by inhibitors for Ca^<2+>/CaM kinase II, A-kinase or C-kinase, suggesting the involvement of protein phosphorylation by CKII in the movement of synaphin.
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