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Dynamics of multiple trafficking behaviors of individual synaptic vesicles revealed by quantum-dot based presynaptic probe.

Lee S, Jung KJ, Jung HS, Chang S - PLoS ONE (2012)

Bottom Line: Actin disruption induced a dramatic decrease in the diffusive behaviors of SVs at synapses while microtubule disruption only reduced extrasynaptic mobility.Glycine-induced synaptic potentiation produced significant increases in synaptic and inter-boutonal trafficking of SVs, which were NMDA receptor- and actin-dependent while NMDA-induced synaptic depression decreased the mobility of the SVs at synapses.Together, our results show that sPH-AP-QD revealed previously unobserved trafficking properties of SVs around synapses, and the dynamic modulation of SV mobility could regulate presynaptic efficacy during synaptic activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biomedical Sciences, Seoul National University College of Medicine, Seoul, South Korea.

ABSTRACT
Although quantum dots (QDs) have provided invaluable information regarding the diffusive behaviors of postsynaptic receptors, their application in presynaptic terminals has been rather limited. In addition, the diffraction-limited nature of the presynaptic bouton has hampered detailed analyses of the behaviors of synaptic vesicles (SVs) at synapses. Here, we created a quantum-dot based presynaptic probe and characterized the dynamic behaviors of individual SVs. As previously reported, the SVs exhibited multiple exchanges between neighboring boutons. Actin disruption induced a dramatic decrease in the diffusive behaviors of SVs at synapses while microtubule disruption only reduced extrasynaptic mobility. Glycine-induced synaptic potentiation produced significant increases in synaptic and inter-boutonal trafficking of SVs, which were NMDA receptor- and actin-dependent while NMDA-induced synaptic depression decreased the mobility of the SVs at synapses. Together, our results show that sPH-AP-QD revealed previously unobserved trafficking properties of SVs around synapses, and the dynamic modulation of SV mobility could regulate presynaptic efficacy during synaptic activity.

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Effects of F-actin or microtubule on the diffusive behaviors of sPH-AP-QDs at synapses and extrasynaptic compartments.(A–B) Three representative sPH-AP-QDs displacement traces showing movement around their initial locations along the axon before and after cytochalasin B (A), or nocodazole (B) treatment. The upper colored bold line denotes the frames on which the sPH-AP-QDs are at synapses. (C–D) Reconstruction of a sPH-AP-QD in (A) or (B) mass center trajectories (white) for 60 s over the sPH fluorescence positive synapses (blue) images along the axons (green) before (left panel) and after (right panel) 20 min treatment with Cyto B (C) or Noco (D). The initial locations of sPH-AP-QDs were indicated by arrow. (E) Comparison of mobile fraction before and after Cyto B or Noco treatment. (F–G) Comparison of average diffusion coefficients at synapses (F) and at extrasynapses (G) before and after Cyto B or Noco treatment. (H–J) Comparison of influx frequency (H), dwell time (I), confinement area at synapses (J) before and after Cyto B or Noco treatment. Values are mean ± s.e. *p<0.01, paired t-test. (n = 11 neurons for Cyto B; n = 9 neurons for Noco, each neuron with at least 10 sPH-AP-QDs).
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pone-0038045-g002: Effects of F-actin or microtubule on the diffusive behaviors of sPH-AP-QDs at synapses and extrasynaptic compartments.(A–B) Three representative sPH-AP-QDs displacement traces showing movement around their initial locations along the axon before and after cytochalasin B (A), or nocodazole (B) treatment. The upper colored bold line denotes the frames on which the sPH-AP-QDs are at synapses. (C–D) Reconstruction of a sPH-AP-QD in (A) or (B) mass center trajectories (white) for 60 s over the sPH fluorescence positive synapses (blue) images along the axons (green) before (left panel) and after (right panel) 20 min treatment with Cyto B (C) or Noco (D). The initial locations of sPH-AP-QDs were indicated by arrow. (E) Comparison of mobile fraction before and after Cyto B or Noco treatment. (F–G) Comparison of average diffusion coefficients at synapses (F) and at extrasynapses (G) before and after Cyto B or Noco treatment. (H–J) Comparison of influx frequency (H), dwell time (I), confinement area at synapses (J) before and after Cyto B or Noco treatment. Values are mean ± s.e. *p<0.01, paired t-test. (n = 11 neurons for Cyto B; n = 9 neurons for Noco, each neuron with at least 10 sPH-AP-QDs).

Mentions: Presynaptic terminals are actin rich compartments and the SV mobility is predicted to be confined by the cytoskeleton [17]. We, however, found that actin disruption by cytochalasin B (Cyto B; 3 µg/ml) decreased the ratio of the mobile/immobile fraction significantly (Fig. 2A, C, E). This was accompanied by a 3.3-fold decrease in the diffusion coefficient at synapses (Fig. 2F, Fig. S6A and Table 1). The influx frequency (the number of entries of sPH-AP-QDs into the synapse per minute) was significantly decreased (Fig. 2A, C, H, Table 1). Cytochalasin B treatment also increased the mean dwell time (the duration that sPH-AP-QD stays at the synapse once it has entered) and decreased the confinement area (see Method) at synapses (Fig. 2I and Table 1) while it did not affect the diffusion coefficient at extrasynaptic areas (Fig. 2G).


Dynamics of multiple trafficking behaviors of individual synaptic vesicles revealed by quantum-dot based presynaptic probe.

Lee S, Jung KJ, Jung HS, Chang S - PLoS ONE (2012)

Effects of F-actin or microtubule on the diffusive behaviors of sPH-AP-QDs at synapses and extrasynaptic compartments.(A–B) Three representative sPH-AP-QDs displacement traces showing movement around their initial locations along the axon before and after cytochalasin B (A), or nocodazole (B) treatment. The upper colored bold line denotes the frames on which the sPH-AP-QDs are at synapses. (C–D) Reconstruction of a sPH-AP-QD in (A) or (B) mass center trajectories (white) for 60 s over the sPH fluorescence positive synapses (blue) images along the axons (green) before (left panel) and after (right panel) 20 min treatment with Cyto B (C) or Noco (D). The initial locations of sPH-AP-QDs were indicated by arrow. (E) Comparison of mobile fraction before and after Cyto B or Noco treatment. (F–G) Comparison of average diffusion coefficients at synapses (F) and at extrasynapses (G) before and after Cyto B or Noco treatment. (H–J) Comparison of influx frequency (H), dwell time (I), confinement area at synapses (J) before and after Cyto B or Noco treatment. Values are mean ± s.e. *p<0.01, paired t-test. (n = 11 neurons for Cyto B; n = 9 neurons for Noco, each neuron with at least 10 sPH-AP-QDs).
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Related In: Results  -  Collection

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pone-0038045-g002: Effects of F-actin or microtubule on the diffusive behaviors of sPH-AP-QDs at synapses and extrasynaptic compartments.(A–B) Three representative sPH-AP-QDs displacement traces showing movement around their initial locations along the axon before and after cytochalasin B (A), or nocodazole (B) treatment. The upper colored bold line denotes the frames on which the sPH-AP-QDs are at synapses. (C–D) Reconstruction of a sPH-AP-QD in (A) or (B) mass center trajectories (white) for 60 s over the sPH fluorescence positive synapses (blue) images along the axons (green) before (left panel) and after (right panel) 20 min treatment with Cyto B (C) or Noco (D). The initial locations of sPH-AP-QDs were indicated by arrow. (E) Comparison of mobile fraction before and after Cyto B or Noco treatment. (F–G) Comparison of average diffusion coefficients at synapses (F) and at extrasynapses (G) before and after Cyto B or Noco treatment. (H–J) Comparison of influx frequency (H), dwell time (I), confinement area at synapses (J) before and after Cyto B or Noco treatment. Values are mean ± s.e. *p<0.01, paired t-test. (n = 11 neurons for Cyto B; n = 9 neurons for Noco, each neuron with at least 10 sPH-AP-QDs).
Mentions: Presynaptic terminals are actin rich compartments and the SV mobility is predicted to be confined by the cytoskeleton [17]. We, however, found that actin disruption by cytochalasin B (Cyto B; 3 µg/ml) decreased the ratio of the mobile/immobile fraction significantly (Fig. 2A, C, E). This was accompanied by a 3.3-fold decrease in the diffusion coefficient at synapses (Fig. 2F, Fig. S6A and Table 1). The influx frequency (the number of entries of sPH-AP-QDs into the synapse per minute) was significantly decreased (Fig. 2A, C, H, Table 1). Cytochalasin B treatment also increased the mean dwell time (the duration that sPH-AP-QD stays at the synapse once it has entered) and decreased the confinement area (see Method) at synapses (Fig. 2I and Table 1) while it did not affect the diffusion coefficient at extrasynaptic areas (Fig. 2G).

Bottom Line: Actin disruption induced a dramatic decrease in the diffusive behaviors of SVs at synapses while microtubule disruption only reduced extrasynaptic mobility.Glycine-induced synaptic potentiation produced significant increases in synaptic and inter-boutonal trafficking of SVs, which were NMDA receptor- and actin-dependent while NMDA-induced synaptic depression decreased the mobility of the SVs at synapses.Together, our results show that sPH-AP-QD revealed previously unobserved trafficking properties of SVs around synapses, and the dynamic modulation of SV mobility could regulate presynaptic efficacy during synaptic activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biomedical Sciences, Seoul National University College of Medicine, Seoul, South Korea.

ABSTRACT
Although quantum dots (QDs) have provided invaluable information regarding the diffusive behaviors of postsynaptic receptors, their application in presynaptic terminals has been rather limited. In addition, the diffraction-limited nature of the presynaptic bouton has hampered detailed analyses of the behaviors of synaptic vesicles (SVs) at synapses. Here, we created a quantum-dot based presynaptic probe and characterized the dynamic behaviors of individual SVs. As previously reported, the SVs exhibited multiple exchanges between neighboring boutons. Actin disruption induced a dramatic decrease in the diffusive behaviors of SVs at synapses while microtubule disruption only reduced extrasynaptic mobility. Glycine-induced synaptic potentiation produced significant increases in synaptic and inter-boutonal trafficking of SVs, which were NMDA receptor- and actin-dependent while NMDA-induced synaptic depression decreased the mobility of the SVs at synapses. Together, our results show that sPH-AP-QD revealed previously unobserved trafficking properties of SVs around synapses, and the dynamic modulation of SV mobility could regulate presynaptic efficacy during synaptic activity.

Show MeSH
Related in: MedlinePlus