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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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Redistribution of synapsin in resting and stimulated reticulospinal synapses. (A) Transverse section of a reticulospinal axon (rs) stained with lamprey-specific synapsin antibodies using immunofluorescence. Punctate labeling (arrows) is localized close to the surface of the axon. (B) The ultrastructural localization of synapsin at rest. Gold particles are associated with the synaptic vesicle cluster. Thick arrows indicate active zone. (C–F) An increase in synapsin labeling in the area within 100 nm from the presynaptic membrane in the vesicle cluster at different states: (C) rest; (D) 5 Hz action potential stimulation; (E) high K+ stimulation; (F) control, SV2 immunolabeling at rest. (G–I) Electron micrographs of synapses stimulated by action potentials at 5 Hz for 20 min and labeled for synapsin. Thin arrow indicates filamentous matrix. Note that synapsin is associated with the filamentous cytomatrix and vesicles in the endocytic zone. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars: (A) 20 μm; (F) 100 nm, also for C–E; (G) 100 nm, also for B; (I) 100 nm, also for H.
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fig2: Redistribution of synapsin in resting and stimulated reticulospinal synapses. (A) Transverse section of a reticulospinal axon (rs) stained with lamprey-specific synapsin antibodies using immunofluorescence. Punctate labeling (arrows) is localized close to the surface of the axon. (B) The ultrastructural localization of synapsin at rest. Gold particles are associated with the synaptic vesicle cluster. Thick arrows indicate active zone. (C–F) An increase in synapsin labeling in the area within 100 nm from the presynaptic membrane in the vesicle cluster at different states: (C) rest; (D) 5 Hz action potential stimulation; (E) high K+ stimulation; (F) control, SV2 immunolabeling at rest. (G–I) Electron micrographs of synapses stimulated by action potentials at 5 Hz for 20 min and labeled for synapsin. Thin arrow indicates filamentous matrix. Note that synapsin is associated with the filamentous cytomatrix and vesicles in the endocytic zone. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars: (A) 20 μm; (F) 100 nm, also for C–E; (G) 100 nm, also for B; (I) 100 nm, also for H.

Mentions: The antisynapsin antibodies used in our experiments produced robust punctate immunoreactivity on cryostat sections of the lamprey spinal cord, reflecting the discrete localization of synapses in this tissue. Single puncta were detected consistently on the internal surface of reticulospinal axons, suggesting a synaptic labeling (Fig. 2 A).


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)

Redistribution of synapsin in resting and stimulated reticulospinal synapses. (A) Transverse section of a reticulospinal axon (rs) stained with lamprey-specific synapsin antibodies using immunofluorescence. Punctate labeling (arrows) is localized close to the surface of the axon. (B) The ultrastructural localization of synapsin at rest. Gold particles are associated with the synaptic vesicle cluster. Thick arrows indicate active zone. (C–F) An increase in synapsin labeling in the area within 100 nm from the presynaptic membrane in the vesicle cluster at different states: (C) rest; (D) 5 Hz action potential stimulation; (E) high K+ stimulation; (F) control, SV2 immunolabeling at rest. (G–I) Electron micrographs of synapses stimulated by action potentials at 5 Hz for 20 min and labeled for synapsin. Thin arrow indicates filamentous matrix. Note that synapsin is associated with the filamentous cytomatrix and vesicles in the endocytic zone. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars: (A) 20 μm; (F) 100 nm, also for C–E; (G) 100 nm, also for B; (I) 100 nm, also for H.
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Related In: Results  -  Collection

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

fig2: Redistribution of synapsin in resting and stimulated reticulospinal synapses. (A) Transverse section of a reticulospinal axon (rs) stained with lamprey-specific synapsin antibodies using immunofluorescence. Punctate labeling (arrows) is localized close to the surface of the axon. (B) The ultrastructural localization of synapsin at rest. Gold particles are associated with the synaptic vesicle cluster. Thick arrows indicate active zone. (C–F) An increase in synapsin labeling in the area within 100 nm from the presynaptic membrane in the vesicle cluster at different states: (C) rest; (D) 5 Hz action potential stimulation; (E) high K+ stimulation; (F) control, SV2 immunolabeling at rest. (G–I) Electron micrographs of synapses stimulated by action potentials at 5 Hz for 20 min and labeled for synapsin. Thin arrow indicates filamentous matrix. Note that synapsin is associated with the filamentous cytomatrix and vesicles in the endocytic zone. svc, synaptic vesicle cluster; v, vesicles in the filamentous matrix; d, dendrite; a, axoplasmic matrix. Bars: (A) 20 μm; (F) 100 nm, also for C–E; (G) 100 nm, also for B; (I) 100 nm, also for H.
Mentions: The antisynapsin antibodies used in our experiments produced robust punctate immunoreactivity on cryostat sections of the lamprey spinal cord, reflecting the discrete localization of synapses in this tissue. Single puncta were detected consistently on the internal surface of reticulospinal axons, suggesting a synaptic labeling (Fig. 2 A).

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