Hirokawa N, Sobue K, Kanda K, Harada A, Yorifuji H
Department of Anatomy and Cell Biology, School of Medicine, University of Tokyo, Japan.
J Cell Biol. 1989 Jan;108(1):111-26. doi: 10.1083/jcb.108.1.111.
We have examined the cytoskeletal architecture and its relationship with synaptic vesicles in synapses by quick-freeze deep-etch electron microscopy (QF.DE). The main cytoskeletal elements in the presynaptic terminals (neuromuscular junction, electric organ, and cerebellar cortex) were actin filaments and microtubules. The actin filaments formed a network and frequently were associated closely with the presynaptic plasma membranes and active zones. Short, linking strands approximately 30 nm long were found between actin and synaptic vesicles, between microtubules and synaptic vesicles. Fine strands (30-60 nm) were also found between synaptic vesicles. Frequently spherical structures existed in the middle of the strands between synaptic vesicles. Another kind of strand (approximately 100 nm long, thinner than the actin filaments) between synaptic vesicles and plasma membranes was also observed. We have examined the molecular structure of synapsin 1 and its relationship with actin filaments, microtubules, and synaptic vesicles in vitro using the low angle rotary shadowing technique and QF.DE. The synapsin 1, approximately 47 nm long, was composed of a head (approximately 14 nm diam) and a tail (approximately 33 nm long), having a tadpole-like appearance. The high resolution provided by QF.DE revealed that a single synapsin 1 cross-linked actin filaments and linked actin filaments with synaptic vesicles, forming approximately 30-nm short strands. The head was on the actin and the tail was attached to the synaptic vesicle or actin filament. Microtubules were also cross-linked by a single synapsin 1, which also connected a microtubule to synaptic vesicles, forming approximately 30 nm strands. The spherical head was on the microtubules and the tail was attached to the synaptic vesicles or to microtubules. Synaptic vesicles incubated with synapsin 1 were linked with each other via fine short fibrils and frequently we identified spherical structures from which two or three fibril radiated and cross-linked synaptic vesicles. We have examined the localization of synapsin 1 using ultracryomicrotomy and colloidal gold-immunocytochemistry of anti-synapsin 1 IgG. Synapsin 1 was exclusively localized in the regions occupied by synaptic vesicles. Statistical analyses indicated that synapsin 1 is located mostly at least approximately 30 nm away from the presynaptic membrane. These data derived via three different approaches suggest that synapsin 1 could be a main element of short linkages between actin filaments and synaptic vesicles, and between microtubules and synaptic vesicles, and between synaptic vesicles in the nerve terminals.(ABSTRACT TRUNCATED AT 400 WORDS)
我们通过快速冷冻深度蚀刻电子显微镜（QF.DE）研究了突触中的细胞骨架结构及其与突触小泡的关系。突触前终末（神经肌肉接头、电器官和小脑皮质）中的主要细胞骨架成分是肌动蛋白丝和微管。肌动蛋白丝形成网络，并且经常与突触前质膜和活性区紧密相连。在肌动蛋白与突触小泡之间、微管与突触小泡之间发现了约30纳米长的短连接链。在突触小泡之间也发现了细链（30 - 60纳米）。在突触小泡之间的链的中间经常存在球形结构。还观察到突触小泡与质膜之间的另一种链（约100纳米长，比肌动蛋白丝细）。我们使用低角度旋转阴影技术和QF.DE在体外研究了突触素1的分子结构及其与肌动蛋白丝、微管和突触小泡的关系。突触素1长约47纳米，由一个头部（直径约14纳米）和一个尾部（长约33纳米）组成，呈蝌蚪状外观。QF.DE提供的高分辨率显示，单个突触素1交联肌动蛋白丝，并将肌动蛋白丝与突触小泡相连，形成约30纳米的短链。头部在肌动蛋白上，尾部附着在突触小泡或肌动蛋白丝上。微管也由单个突触素1交联，突触素1还将微管与突触小泡相连，形成约30纳米的链。球形头部在微管上，尾部附着在突触小泡或微管上。用突触素1孵育的突触小泡通过细短纤维相互连接，并且我们经常识别出有两三条纤维辐射并交联突触小泡的球形结构。我们使用抗突触素1 IgG的超薄冷冻切片术和胶体金免疫细胞化学研究了突触素1的定位。突触素1仅定位于突触小泡占据的区域。统计分析表明，突触素1大多位于距突触前膜至少约30纳米处。通过三种不同方法获得的数据表明，突触素1可能是神经末梢中肌动蛋白丝与突触小泡之间、微管与突触小泡之间以及突触小泡之间短连接的主要成分。（摘要截短至400字）
