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Colocalization of synapsin and actin during synaptic vesicle recycling.

Bloom O, Evergren E, Tomilin N, Kjaerulff O, Löw P, Brodin L, Pieribone VA, Greengard P, Shupliakov O - J. Cell Biol. (2003)

Bottom Line: In addition, actin and synapsin were found colocalized in a dynamic filamentous cytomatrix at the sites of synaptic vesicle recycling, endocytic zones.Synapsin immunolabeling was not associated with clathrin-coated intermediates but was found on vesicles that appeared to be recycling back to the cluster.Disruption of synapsin function by microinjection of antisynapsin antibodies resulted in a prominent reduction of the cytomatrix at endocytic zones of active synapses.

View Article: PubMed Central - PubMed

Affiliation: The Rockefeller University, New York, NY 10021, USA. ona@chronos.med.yale.edu

ABSTRACT
It has been hypothesized that in the mature nerve terminal, interactions between synapsin and actin regulate the clustering of synaptic vesicles and the availability of vesicles for release during synaptic activity. Here, we have used immunogold electron microscopy to examine the subcellular localization of actin and synapsin in the giant synapse in lamprey at different states of synaptic activity. In agreement with earlier observations, in synapses at rest, synapsin immunoreactivity was preferentially localized to a portion of the vesicle cluster distal to the active zone. During synaptic activity, however, synapsin was detected in the pool of vesicles proximal to the active zone. In addition, actin and synapsin were found colocalized in a dynamic filamentous cytomatrix at the sites of synaptic vesicle recycling, endocytic zones. Synapsin immunolabeling was not associated with clathrin-coated intermediates but was found on vesicles that appeared to be recycling back to the cluster. Disruption of synapsin function by microinjection of antisynapsin antibodies resulted in a prominent reduction of the cytomatrix at endocytic zones of active synapses. Our data suggest that in addition to its known function in clustering of vesicles in the reserve pool, synapsin migrates from the synaptic vesicle cluster and participates in the organization of the actin-rich cytomatrix in the endocytic zone during synaptic activity.

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Distribution of synapsin and actin in active synapses as revealed by double labeling. (A) Electron micrograph of a synapse stimulated with high K+ for 30 min. Large gold particles (enhanced 10 nm) denote actin and small (enhanced 5 nm) denote synapsin immunoreactivity. (B–D) Regions of endocytic zones, containing filamentous cytomatrix in three synapses stimulated at 5 Hz for 20 min and labeled with immunogold as in A. (E and F) Electron micrographs of endocytic regions containing clathrin-coated intermediates in synapses stimulated with high K+ for 30 min. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars, 100 nm.
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fig5: Distribution of synapsin and actin in active synapses as revealed by double labeling. (A) Electron micrograph of a synapse stimulated with high K+ for 30 min. Large gold particles (enhanced 10 nm) denote actin and small (enhanced 5 nm) denote synapsin immunoreactivity. (B–D) Regions of endocytic zones, containing filamentous cytomatrix in three synapses stimulated at 5 Hz for 20 min and labeled with immunogold as in A. (E and F) Electron micrographs of endocytic regions containing clathrin-coated intermediates in synapses stimulated with high K+ for 30 min. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars, 100 nm.

Mentions: To directly test if synapsin and actin are colocalized around the synapse, double labeling experiments were performed. Sections were incubated in a mixture of rabbit anti–synapsin domain D antibodies and a mouse anti-actin antibody. The primary antibodies were detected with the corresponding secondary antibodies coupled to 5- and 10-nm gold particles, respectively. In stimulated synapses, labeling for both synapsin and actin was observed within the filamentous cytomatrix and on synaptic vesicles recycling back to the cluster (Fig. 5, A–D).


Colocalization of synapsin and actin during synaptic vesicle recycling.

Bloom O, Evergren E, Tomilin N, Kjaerulff O, Löw P, Brodin L, Pieribone VA, Greengard P, Shupliakov O - J. Cell Biol. (2003)

Distribution of synapsin and actin in active synapses as revealed by double labeling. (A) Electron micrograph of a synapse stimulated with high K+ for 30 min. Large gold particles (enhanced 10 nm) denote actin and small (enhanced 5 nm) denote synapsin immunoreactivity. (B–D) Regions of endocytic zones, containing filamentous cytomatrix in three synapses stimulated at 5 Hz for 20 min and labeled with immunogold as in A. (E and F) Electron micrographs of endocytic regions containing clathrin-coated intermediates in synapses stimulated with high K+ for 30 min. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars, 100 nm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199372&req=5

fig5: Distribution of synapsin and actin in active synapses as revealed by double labeling. (A) Electron micrograph of a synapse stimulated with high K+ for 30 min. Large gold particles (enhanced 10 nm) denote actin and small (enhanced 5 nm) denote synapsin immunoreactivity. (B–D) Regions of endocytic zones, containing filamentous cytomatrix in three synapses stimulated at 5 Hz for 20 min and labeled with immunogold as in A. (E and F) Electron micrographs of endocytic regions containing clathrin-coated intermediates in synapses stimulated with high K+ for 30 min. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars, 100 nm.
Mentions: To directly test if synapsin and actin are colocalized around the synapse, double labeling experiments were performed. Sections were incubated in a mixture of rabbit anti–synapsin domain D antibodies and a mouse anti-actin antibody. The primary antibodies were detected with the corresponding secondary antibodies coupled to 5- and 10-nm gold particles, respectively. In stimulated synapses, labeling for both synapsin and actin was observed within the filamentous cytomatrix and on synaptic vesicles recycling back to the cluster (Fig. 5, A–D).

Bottom Line: In addition, actin and synapsin were found colocalized in a dynamic filamentous cytomatrix at the sites of synaptic vesicle recycling, endocytic zones.Synapsin immunolabeling was not associated with clathrin-coated intermediates but was found on vesicles that appeared to be recycling back to the cluster.Disruption of synapsin function by microinjection of antisynapsin antibodies resulted in a prominent reduction of the cytomatrix at endocytic zones of active synapses.

View Article: PubMed Central - PubMed

Affiliation: The Rockefeller University, New York, NY 10021, USA. ona@chronos.med.yale.edu

ABSTRACT
It has been hypothesized that in the mature nerve terminal, interactions between synapsin and actin regulate the clustering of synaptic vesicles and the availability of vesicles for release during synaptic activity. Here, we have used immunogold electron microscopy to examine the subcellular localization of actin and synapsin in the giant synapse in lamprey at different states of synaptic activity. In agreement with earlier observations, in synapses at rest, synapsin immunoreactivity was preferentially localized to a portion of the vesicle cluster distal to the active zone. During synaptic activity, however, synapsin was detected in the pool of vesicles proximal to the active zone. In addition, actin and synapsin were found colocalized in a dynamic filamentous cytomatrix at the sites of synaptic vesicle recycling, endocytic zones. Synapsin immunolabeling was not associated with clathrin-coated intermediates but was found on vesicles that appeared to be recycling back to the cluster. Disruption of synapsin function by microinjection of antisynapsin antibodies resulted in a prominent reduction of the cytomatrix at endocytic zones of active synapses. Our data suggest that in addition to its known function in clustering of vesicles in the reserve pool, synapsin migrates from the synaptic vesicle cluster and participates in the organization of the actin-rich cytomatrix in the endocytic zone during synaptic activity.

Show MeSH
Related in: MedlinePlus