Ultrafast endocytosis at mouse hippocampal synapses.
Ultrafast endocytosis at mouse hippocampal synapses.
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为了维持神经传递，突触囊泡及其相关蛋白质必须在突触局部循环。突触囊泡被认为在融合后约20秒通过网格蛋白支架的组装或在约1秒内通过经由“吻和跑”内吞作用的融合孔的逆转而再生。在这里，我们使用光遗传学来刺激培养的海马神经元与一个单一的刺激，快速冷冻后，固定的时间间隔，并检查超微结构使用电子显微镜-“闪光和冷冻”电子显微镜。对接的囊泡在刺激的30 ms内融合并塌陷到膜中。代偿性内吞作用在活性区两侧的位点以50-100 ms发生。抑制肌动蛋白聚合可阻断内陷，抑制发动蛋白可阻断断裂。因为完整的突触囊泡不能被回收，这种形式的回收与吻和跑的内吞作用不相容;而且它比网格蛋白介导的内吞作用快200倍。很可能“超快内吞作用”是专门用于快速恢复膜的表面积。
To sustain neurotransmission, synaptic vesicles and their associated proteins must be recycled locally at synapses. Synaptic vesicles are thought to be regenerated ~20 s after fusion by the assembly of clathrin scaffolds or in ~1 s by the reversal of fusion pores via ‘kiss-and-run’ endocytosis. Here we use optogenetics to stimulate cultured hippocampal neurons with a single stimulus, rapidly freeze them after fixed intervals and examine the ultrastructure using electron microscopy – ‘flash-and-freeze’ electron microscopy. Docked vesicles fuse and collapse into the membrane within 30 ms of the stimulus. Compensatory endocytosis occurs with 50-100 ms at sites flanking the active zone. Invagination is blocked by inhibition of actin polymerization, and scission is blocked by inhibiting dynamin. Because intact synaptic vesicles are not recovered, this form of recycling is not compatible with kiss-and-run endocytosis; moreover it is 200-fold faster than clathrin-mediated endocytosis. It is likely that ‘ultrafast endocytosis’ is specialized to rapidly restore the surface area of the membrane.