Journal: Neural Regeneration Research

Article Title: Nerve root magnetic stimulation regulates the synaptic plasticity of injured spinal cord by ascending sensory pathway

doi: 10.4103/NRR.NRR-D-24-00628

Figure Lengend Snippet: NRMS upregulates transcriptional levels of genes related to synaptic plasticity in damaged region of SCI rats. (A) The functional classification of different genes in the SCI + NRMS and SCI + SS groups by GO enrichment analysis. (B) The functional classification of different genes between the SCI + NRMS and SCI + SS groups by KEGG database. (C–F) The relative levels of Syn2 , Syp , GAP43 , and MAP2 mRNA normalized by the Sham group. The bold lines, upper boundaries, and lower boundaries represent the medians, 75 th percentiles, and 25 th percentiles, respectively. Whiskers extend 1.5 times interquartile range ( n = 4 per group). * P < 0.05, vs . Sham group; # P < 0.05, vs . SCI + SS group. GAP43: Growth-associated protein 43; GO: Gene Ontology; KEGG: Kyoto Encyclopedia of Genes and Genomes; MAP2: microtubule-associated protein 2; NRMS: nerve root magnetic stimulation; SCI: spinal cord injury; SS: sham stimulation; Syn2: synapsin II; Syp: synaptophysin.

Article Snippet: Moreover, the major functions of Synapsin II are focused on the regulation of Ca 2+ -dependent plasticity and neuronal outgrowth in glutamatergic neurons (Matos et al., 2019).

Techniques: Functional Assay

Journal: Neural Regeneration Research

Article Title: Nerve root magnetic stimulation regulates the synaptic plasticity of injured spinal cord by ascending sensory pathway

doi: 10.4103/NRR.NRR-D-24-00628

Figure Lengend Snippet: NRMS alters expression of proteins associated with synaptic plasticity in the injured spinal cord of rats. (A) Representative western blots show the expression levels of PSD95, GAP43, Synapsin I and Synapsin II collected from the damaged area of spinal cord on day 22 after SCI. (B–E) The semi-quantified results of PSD95, GAP43, Synapsin I, and Synapsin II protein levels. All experiments were repeated three times. Data are expressed as mean ± SD ( n = 5 per group). ** P < 0.01, *** P < 0.001, vs. Sham group; # P < 0.05, ## P < 0.01, vs . SCI + SS group. GAP43: Growth-associated protein 43; PSD95: postsynaptic density protein-95; NRMS: nerve root magnetic stimulation; SCI: spinal cord injury; SS: sham stimulation.

Article Snippet: Moreover, the major functions of Synapsin II are focused on the regulation of Ca 2+ -dependent plasticity and neuronal outgrowth in glutamatergic neurons (Matos et al., 2019).

Techniques: Expressing, Western Blot

Journal:

Article Title: Nerve Agent Exposure Elicits Site-Specific Changes in Protein Phosphorylation in Mouse Brain

doi: 10.1016/j.brainres.2010.04.034

Figure Lengend Snippet: Effect of DFP on a panel of neuronal phosphorylation sites comprising the CNSProfile technology platform as measured in striatum and hippocampus of male FVB mice.

Article Snippet: Immunoblotting was carried out using phosphorylation state-specific antibodies (and respective phosphorylation state insensitive antibodies for measuring the total protein levels) raised against pT34, pT75, pS102 or pS137 of DARPP-32 and pS94 of spinophilin (ITI); pS133 of CREB (Upstate Biotechnology, Charlottesville, VA); pT183 of ERK1/2 (Promega, Madison, WI); pS897 of the NMDA receptor NR1 subunit (Upstate); pS831 or pS845 of the AMPA receptor GluR1 subunit (Upstate); S9 of the GSK3β kinase (BD Biosciences); S473 or T308 of the protein kinase Akt (BD Biosciences); S549 or S603 of the synaptic vesicle protein, synapsin I/II (Dr. Angus Nairn, RU and BD Biosciences); or pS40 of tyrosine hydroxylase (Chemicon, Temecula, CA).

Techniques:

Journal:

Article Title: Nerve Agent Exposure Elicits Site-Specific Changes in Protein Phosphorylation in Mouse Brain

doi: 10.1016/j.brainres.2010.04.034

Figure Lengend Snippet: Sites of phosphorylation monitored in brain after DFP treatment using CNSProfile.

Article Snippet: Immunoblotting was carried out using phosphorylation state-specific antibodies (and respective phosphorylation state insensitive antibodies for measuring the total protein levels) raised against pT34, pT75, pS102 or pS137 of DARPP-32 and pS94 of spinophilin (ITI); pS133 of CREB (Upstate Biotechnology, Charlottesville, VA); pT183 of ERK1/2 (Promega, Madison, WI); pS897 of the NMDA receptor NR1 subunit (Upstate); pS831 or pS845 of the AMPA receptor GluR1 subunit (Upstate); S9 of the GSK3β kinase (BD Biosciences); S473 or T308 of the protein kinase Akt (BD Biosciences); S549 or S603 of the synaptic vesicle protein, synapsin I/II (Dr. Angus Nairn, RU and BD Biosciences); or pS40 of tyrosine hydroxylase (Chemicon, Temecula, CA).

Techniques: Activity Assay

Journal: The EMBO Journal

Article Title: Newly produced synaptic vesicle proteins are preferentially used in synaptic transmission

doi: 10.15252/embj.201798044

Figure Lengend Snippet: A, B Two‐color STED analysis of changes in synaptic vesicle protein levels during the transition from the releasable state to the inactive state. Living neurons were incubated with Atto647N‐conjugated Synaptotagmin 1 lumenal domain antibodies for 1 h at 37°C, to label the actively recycling vesicles, as in Fig A, and were then fixed and co‐immunostained for different proteins of interest directly, or were placed in a cell culture incubator for 3–4 days, to enable the antibody‐labeled molecules to enter the inactive pool, before fixation and co‐immunostaining. The samples were embedded in melamine and cut in ultrathin (50 nm) sections, before two‐color STED imaging, as in Fig B. Exemplary images are shown in (A) for SNAP25 and in (B) for Syntaxin 1, with the protein of interest signal next to the signal from the live tagging of Synaptotagmin 1, and a merged image of both, for day 0 (releasable vesicles) and day 4 (inactive vesicles). Scale bar: 1 μm. C We analyzed the amount of fluorescence corresponding to the protein of interest that overlapped with the Synaptotagmin 1 signal (i.e., the two signals were presented within the same voxels, which are substantially below the synaptic vesicle volume in this experiment). The only protein whose levels changed significantly is SNAP25 (SNAP25, n (day 0) = 4, n (day 4) = 3, * P = 0.0124, t (5) = 3.8200; Syntaxin 1, n (day 0) = 3, n (day 4) = 3, P = 0.8850, t (4) = 0.1541; VGlut 1/2, n (day 0) = 2, n (day 4) = 3, P = 0.1986, t (3) = 1.6447; vATPase, n (day 0) = 3, n (day 4) = 4, P = 0.7340, t (5) = 0.3594; VAMP2, n (day 0) = 4, n (day 4) = 4, P = 0.8837, t (6) = 0.1527; Synaptotagmin 1, n (day 0) = 3, n (day 4) = 3, P = 0.1604, t (4) = 1.7208; Syntaxin 16, n (day 0) = 4, n (day 4) = 4, P = 0.7406, t (6) = 0.3468; VAMP4, n (day 0) = 3, n (day 4) = 3, P = 0.9863, t (4) = 0.0183; Synapsin I/II, n (day 0) = 3, n (day 4) = 3, P = 0.6638, t (4) = 0.4685; at least 10 neurons sampled per experiment). Statistical significance was evaluated using unpaired t ‐tests. All data represent the mean ± SEM. Data information: Imaging was performed with a Leica SP5 STED microscope. Source data are available online for this figure.

Article Snippet: The primary antibodies used were as follows: Synaptophysin (guinea pig, # 101 004 or mouse, clone 7.2, # 101 011, both from Synaptic Systems, Göttingen, Germany), SNAP25 (rabbit, # 111 002; Synaptic Systems), Syntaxin 1 (rabbit, # 110 302; Synaptic Systems), VGlut 1/2 (rabbit, # 135 503; Synaptic Systems), vATPase (rabbit, # 109 002; Synaptic Systems), VAMP2 (rabbit, # 104 202; Synaptic Systems), Synaptotagmin 1 (mouse, clone 604.2, # 105 311, or mouse, clone 604.2, directly conjugated to Atto647N, # 105 311AT1, or rabbit, # 105 102, all from Synaptic Systems), Syntaxin 16 (rabbit, # 110 162; Synaptic Systems), VAMP4 (rabbit, # 136 002; Synaptic Systems), Synapsin (rabbit, # 106 002; Synaptic Systems), Rab7 (# 9367; Cell Signaling Technology, Cambridge, UK), and PSD95 (rabbit, #3450S; Cell Signaling).

Techniques: Incubation, Cell Culture, Labeling, Immunostaining, Imaging, Fluorescence, Microscopy