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Two pathways of synaptic vesicle retrieval revealed by single-vesicle imaging.

ABSTRACT: Synaptic vesicle recycling is essential for maintaining efficient synaptic transmission. Detailed dissection of single-vesicle recycling still remains a major challenge. We have developed a fluorescent pH reporter that permits us to follow the fate of individual vesicles at hippocampal synapses after exocytosis. Here we show that, during low-frequency stimulation, single-vesicle fusion leads to two distinct vesicle internalizations, instead of one, as in general perception: one by a fast endocytosis pathway ( approximately 3 s), the other by a slow endocytosis pathway (after 10 s). The exocytosed vesicular proteins are preferentially recaptured in both pathways. RNAi knockdown of clathrin inhibits both pathways. As stimulation frequency increases, the number of endocytosed vesicles begins to match antecedent exocytosis. Meanwhile, the slow endocytosis is accelerated and becomes the predominant pathway. These results reveal that two pathways of endocytosis are orchestrated during neuronal activity, establishing a highly efficient endocytosis at central synapses.

SUBMITTER: Zhu Y

PROVIDER: S-EPMC2707496 | biostudies-literature | 2009 Feb

REPOSITORIES: biostudies-literature

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Two pathways of synaptic vesicle retrieval revealed by single-vesicle imaging.

Zhu Yongling Y Xu Jian J Heinemann Stephen F SF

Neuron 20090201 3

Synaptic vesicle recycling is essential for maintaining efficient synaptic transmission. Detailed dissection of single-vesicle recycling still remains a major challenge. We have developed a fluorescent pH reporter that permits us to follow the fate of individual vesicles at hippocampal synapses after exocytosis. Here we show that, during low-frequency stimulation, single-vesicle fusion leads to two distinct vesicle internalizations, instead of one, as in general perception: one by a fast endocyt ...[more]

PMID: 19217377

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Project description:BackgroundDuring nerve growth, cytoplasmic vesicles add new membrane preferentially to the growth cone located at the distal tip of extending axons. Growth cone membrane is also retrieved locally, and asymmetric retrieval facilitates membrane remodeling during growth cone repulsion by a chemorepellent gradient. Moreover, growth inhibitory factors can stimulate bulk membrane retrieval and induce growth cone collapse. Despite these functional insights, the processes mediating local membrane remodeling during axon extension remain poorly defined.ResultsTo investigate the spatial and temporal dynamics of membrane retrieval in actively extending growth cones, we have used a transient labeling and optical recording method that can resolve single vesicle events. Live-cell confocal imaging revealed rapid membrane retrieval by distinct endocytic modes based on spatial distribution in Xenopus spinal neuron growth cones. These modes include endocytic "hot-spots" triggered at the base of filopodia, at the lateral margins of lamellipodia, and along dorsal ridges of the growth cone. Additionally, waves of endocytosis were induced when individual filopodia detached from the substrate and fused with the growth cone dorsal surface or with other filopodia. Vesicle formation at sites of membrane remodeling by self-contact required F-actin polymerization. Moreover, bulk membrane retrieval by macroendocytosis correlated positively with the substrate-dependent rate of axon extension and required the function of Rho-family GTPases.ConclusionsThis study provides insight into the dynamic membrane remodeling processes essential for nerve growth by identifying several distinct modes of rapid membrane retrieval in the growth cone during axon extension. We found that endocytic membrane retrieval is intensified at specific subdomains and may drive the dynamic membrane ruffling and re-absorption of filopodia and lamellipodia in actively extending growth cones. The findings offer a platform for determining the molecular mechanisms of distinct endocytic processes that may remodel the surface distribution of receptors, ion channels and other membrane-associated proteins locally to drive growth cone extension and chemotactic guidance.

Dataset Information

Two pathways of synaptic vesicle retrieval revealed by single-vesicle imaging.

Publications

Two pathways of synaptic vesicle retrieval revealed by single-vesicle imaging.

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