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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional <t>deconvolution</t> software (MetaMorph) was used.
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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional <t>deconvolution</t> software (MetaMorph) was used.
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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional <t>deconvolution</t> software (MetaMorph) was used.
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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional <t>deconvolution</t> software (MetaMorph) was used.
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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional <t>deconvolution</t> software (MetaMorph) was used.
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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional <t>deconvolution</t> software (MetaMorph) was used.
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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional <t>deconvolution</t> software (MetaMorph) was used.
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cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional deconvolution software (MetaMorph) was used.

Journal: The Journal of Neuroscience

Article Title: BDNF-Dependent Recycling Facilitates TrkB Translocation to Postsynaptic Density during LTP via a Rab11-Dependent Pathway

doi: 10.1523/JNEUROSCI.3256-12.2013

Figure Lengend Snippet: cLTP-enhanced recycling facilitates TrkB-FL translocation into postsynaptic densities. A, Hippocampal neurons (18 DIV) were transfected with GFP and treated with various agents as indicated. Endogenous TrkB-FL stained by the Trk (C-14) antibody that recognizes the C terminus of Trk (catalog# sc-11; Santa Cruz Biotechnology) was labeled red. Arrowheads and arrows denote protrusions with and without TrkB in the spine head, respectively. Scale bars, 1 μm. B, Three types of TrkB-FL distribution in spines. Scale bars, 2 μm. C, Proportions of spines containing TrkB-FL in the indicated locations. Graphs represent means ± SEM (n = 116, 169, and 135 spines analyzed from 8, 8, and 10 neurons, respectively, *p < 0.05 relative to no BDNF control of each group, #p < 0.05, χ2 test. D, Coimmunofluorescence labeling of recycled TrkB and PSD-95 in neurons (9 DIV) transfected with pcDNA3.1 or pCAGIG-GFP-Rab11S25N. Hippocampal neurons were stained for recycled TrkB (green) and PSD-95 (red) with (left) or without (right) glycine stimulation. Blue indicates neurons expressing pcAGIG-GFP-Rab11S25N. Colocalization is indicated by white arrows. Scale bars, 10 μm. E, Quantification analysis of the colocalization of recycled TrkB-FL or TrkB.T1 with PSD-95 in D. Graphs represent means ± SEM (n ≥ 30 cells for each condition per experiment, ***p < 0.001 relative to its baseline, #p < 0.05, Student's t test). F, Quantification analysis of the clusters of PSD95. Images were taken using a 100× objective and two-dimensional deconvolution software (MetaMorph) was used.

Article Snippet: Images were taken using a 100× objective and two-dimensional deconvolution software (MetaMorph) was used.

Techniques: Translocation Assay, Transfection, Staining, Labeling, Control, Expressing, Software