Journal: The Journal of Neuroscience

Article Title: Neuromuscular NMDA Receptors Modulate Developmental Synapse Elimination

doi: 10.1523/JNEUROSCI.1181-16.2016

Figure Lengend Snippet: Reduction of NMDA receptor activity slows synapse elimination. A, Pharmacological reduction of NMDA and AMPA receptor activity with AP5 and CNQX between P4 and P11. Left, Whole-mount NMJ immunolabeling in anterior tibialis (AT) muscle of P11 pups. Asterisks show polyinnervated end plates. Red, Rhodamine-α-bungarotoxin labels ACh receptors. Green, Antibodies SV2 and 2H3 label synaptic vesicles and neurofilaments. Right, Means and distributions of percentage of multiply innervated muscles. Open symbols indicate control; filled symbols, treated; horizontal lines, means. The treated muscles contained significantly more multiply innervated fibers than the contralateral internal control muscles (EDL: 22 ± 3% control limb vs 52 ± 4% treated limb, p = 0.00006, n = 12, 12; soleus: 35 ± 3% control limb vs 53 ± 2% treated limb, p = 0.00001, n = 11, 13). B, Vivo-morpholino downregulation of GluN1 expression between P4 and P11. A vivo-morpholino was pressure injected into the AT muscle end plate band in P4 pups. Conventions are as in A. The AT muscle demonstrated significant retention (9 ± 5% internal control limb vs 24 ± 8% treated limb, p = 0.001, n = 8, 8). Scale bars, 50 μm.

Article Snippet: Immunohistochemistry Muscles were immersion fixed for 15 min in 4% paraformaldehyde, rinsed 3× in PBS, stained with 10% rhodamine-conjugated α-bungarotoxin (catalog #T1175; Life Technologies), bathed overnight in primary antibodies SV2 (synaptic vesicles, 1:20, RRID:AB_2315390 and 2H3 neurofilament, 1:10, RRID:AB_531793; Developmental Studies Hybridoma Bank) and visualized by secondary antibody Alexa Fluor 488 donkey anti-mouse (RRID:AB_2556542, catalog # {"type":"entrez-nucleotide","attrs":{"text":"R37114","term_id":"794570","term_text":"R37114"}} R37114 ; Life Technologies).

Techniques: Activity Assay, Immunolabeling, Muscles, Control, Expressing, Injection

Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

Article Title: Nicotinamide Adenine Dinucleotide-Dependent Binding of the Neuronal Ca 2+ Sensor Protein GCAP2 to Photoreceptor Synaptic Ribbons

doi: 10.1523/JNEUROSCI.3701-09.2010

Figure Lengend Snippet: Antibodies and labeling reagents

Article Snippet: SV2 A; Synaptic Systems, cat. #119 00 2 , Rabbit polyclonal , 1:500 , GAR-Cy2; Rockland, cat. #611-111-122 , 1:1000.

Techniques: Labeling, Western Blot, Transduction, Immunolabeling, Immunostaining

Journal: Cells

Article Title: Emerging Role of miR-21-5p in Neuron–Glia Dysregulation and Exosome Transfer Using Multiple Models of Alzheimer’s Disease

doi: 10.3390/cells11213377

Figure Lengend Snippet: Characterization reveals successfully differentiated and maturated neurons, while Neu-AD cell lines show alterations in intracellular and exosomal miRNAs, which include the simultaneous elevation of miR-21 and miR-124. ( A ) Schematic representation of iPSC differentiation into induced neurons (Neu), as detailed in Materials and Methods. ( B ) iPSC-derived neurons express the typical neuronal markers synapsin-1, MAP-2, Tau, SV-2, PSD-95 and F-actin. ( C ) Representative images of synaptic puncta in neurons, with co-localization of synapsin-1 and PSD-95, and ( D ) quantification of individual and double-labeled puncta. Nuclei were counterstained with Hoechst. ( E ) Comparative number of cells expressing MAP-2, Tau, PSD-95 and SV-2 protein markers in neurons from AD and Ctrl donors. ( F ) Quantification of the selected inflamma-miRNAs in Neu-AD cell lines and ( G ) in their derived exosomes expressed as the binary logarithm of fold change vs. Neu-Ctrl cells (dotted line). Samples from a male patient (in blue dots) and a female patient (in pink dots) were distinguished. Scale bars correspond to 40 µm in ( A , B ) and to 20 µm in ( C ). Results are mean ± SEM from at least 7 independent experiments. Two-tailed Student’s t -test: * p < 0.05, Neu-AD vs. Neu-Ctrl; ## p < 0.01, male (n = 4) vs. female (n = 3) samples. Neu, iPSC-induced neurons; AD, Alzheimer’s disease; Ctrl, control; EXO, exosomes. SV-2, synaptic vesicle protein 2; PSD-95, postsynaptic density protein; MAP-2, microtubule-associated protein-2; F-actin, filamentous actin; NDM, neural differentiation media; LDN, LDN193189 dihydrochloride; SB, SB431542; FGF, fibroblast growth factor; EGF, epidermal growth factor; NIM, neural induction media; BDNF, brain-derived neurotrophic factor; GDNF, glial-cell-derived neurotrophic factor; ULA, ultra-low-attachment plates; PO, poly-L-ornithine; L, laminin; D, day.

Article Snippet: The following primary antibodies were used: rat anti-CD11b (1:50, Biolegend, San Diego, CA, USA), rabbit anti-TMEM119 (1:50, Abcam, Cambridge, UK), rabbit anti-Iba1 (1:100, FUJIFILM Wako, Osaka, Japan), rabbit anti-CX3CR1 (1:100, Santa Cruz Biotechnology, Dallas, TX, USA), rabbit anti-MFGE8 (1:100, Santa Cruz Biotechnology), mouse anti-iNOS (1:100, BD Biosciences, Franklin Lakes, NJ, USA), goat anti-Arg 1 (1:100, Santa Cruz Biotechnology), rabbit anti-TREM2 (1:100, Cell signaling, Danvers, MA, USA), rabbit anti-synapsin-1 (1:100, produced in house), mouse anti-MAP2 (1:100, Millipore, Burlington, MA, USA), rabbit anti-Tau (1:200, Synaptic Systems, Goettingen, Germany), rabbit anti-SV-2 (1:200, Synaptic Systems), mouse anti-PSD-95 (1:200, Millipore), rabbit anti-S100B (1:200, Abcam) and mouse anti-GFAP (1:200, Novocastra, Newcastle, UK).

Techniques: Derivative Assay, Labeling, Expressing, Two Tailed Test

Journal: Frontiers in Cellular Neuroscience

Article Title: Characterization of a Synaptic Vesicle Binding Motif on the Distal CaV2.2 Channel C-terminal

doi: 10.3389/fncel.2016.00145

Figure Lengend Snippet: Antibodies.

Article Snippet: SV2A (m) , SV2A (SV2) , Synaptic Systems, GMBH (clone 171G0) , 1:1000.

Techniques: FLAG-tag, Variant Assay