Journal: Nature medicine

Article Title: Spinal cord reconstitution with homologous neural grafts enables robust corticospinal regeneration

doi: 10.1038/nm.4066

Figure Lengend Snippet: ( a – b ) Rostral aspect of a GFP-expressing multipotent NPC graft in the site of T3 complete spinal cord transection, shown in horizontal section. Rostral is left, caudal is right. CST axons are labeled with biotinylated dextran amine (BDA, shown in red). Inset shows overview of the graft. Scale bar, 240 μm; 1 mm (inset). ( b ) The CST approaches and regenerates into an NPC graft in the lesion site. Scale bar, 240 μm. ( c – e ) Higher magnification views of boxed areas in b show the density, varicosities, and tortuosities of regenerating corticospinal axons that extend into the graft. Scale bars c, 60 μm; d, 30 μm; e , 20 μm. ( f ) Regenerating CST axons surround neurons in the center of the graft (* in the inset), 3mm from the rostral host–graft border. Inset shows overview of the graft. Scale bar, 20 μm; 1 mm (inset). ( g ) Quantification of CST axons in NPC grafts in six rats (left). Quantification of the proportion of CST axons in NPC grafts, normalized to the total number of CST axons 0.5mm rostral to the lesion site in six rats (right). Data are presented as mean ± SEM. Circles indicate data from individual rats. ( h ) Triple labeling for GFP, CST, and microtubule-associated protein 2 (MAP2) demonstrates a CST axon in the graft that exhibits a bouton-like swelling in close apposition to a dendrite of a grafted neuron, labeled with MAP2 (arrowheads; upper left, scale bar, 2 μm). Triple labeling for GFP, CST, and synaptophysin (Syn; upper right, scale bar, 2 μm) or vesicular glutamate transporter 1 (vGlut1; lower left, scale bar, 2 μm) in the graft indicates that the corticospinal axon in the graft is co-localized with presynaptic and excitatory synaptic markers, respectively (arrowheads). Electron microscopy image of a 3,3′-diaminobenzidine (DAB)–labeled CST axon terminal (black) forming a synapse (arrows) with a neuronal process within the graft (lower right). Arrowheads indicate presynaptic vesicles. Scale bar, 100 nm. ( i ) Sagittal images of GPF-expressing syngenic bone marrow stromal cell (MSC) grafts in the site of C4 spinal cord transection. Left, double labeling for CST axons (red) and GFP expressing MSCs (green); right, double labeling for CST axons and glial fibrillary acidic protein (GFAP, blue). Dashed lines indicate rostral host/graft interface. Scale bar, 200 μm. ( j ) Sagittal image of CST axons in a C4 CST transection lesion in the absence of any graft. Scale bar, 200 μm.

Article Snippet: Sections were incubated with primary antibodies against GFP (rabbit from Invitrogen at 1:1000 or chicken from Abcam at 1:1000); GFAP (mouse from Chemicon at 1:1000 or rabbit from Dako at 1:750 to label astrocytes); human GFAP (rabbit from Origene at 1:500 to label human specific astrocytes); NeuN (mouse from Chemicon at 1:500 to label mature neurons); ChAT (goat from Chemicon at 1:250 to label spinal cord motor neurons); MAP2 (mouse from Chemicon at 1:2,000 to label dendrites); vesicular glutamate transporters 1 (vGlut1, mouse from Chemicon at 1:1,000 to label glutamatergic terminals); synaptophysin (mouse from Chemicon at 1:1,000 to label presynaptic terminals); adenomatous polyposis coli (APC, mouse from Oncogene at 1:400 to label oligodendrocytes); NG2 (rabbit from Millipore at 1:400 to label oligodendrocyte precursors); Hu (human at 1:1000 to label neurons, generous gift from Robert Darnell, The Rockefeller University, NY); doublecortin (DCX, goat from Santa Cruz at 1: 250 to label immature neurons); nestin (mouse from BD at 1:200 to label neural progenitor cells); neurofilament (NF200; mouse from Millipore at 1:250 to label axons); 27C7 (mouse at 1:200 to label Schwann cells, generous gift from K. Wewetzer, University of Freiburg, Freiburg, Germanyat); RFP (mouse from Abcam at 1:200); human nuclei (mouse from Millipore at 1:200); PAX6 (rabbit from Covance at 1:1000); Pax6 (mouse from DHSB at 1:5000); Sox1 (Goat from R&B at 1:1000); HoxB4 (Rat from DHSB at 1:50); FoxG1 (rabbit from Abcam at 1:100); OTX2 (goat from R and D at 1:2000); EN1 (mouse from DHSB at 1:800); Camk2 (rabbit from Genetex at 1:200), and Dyelight 405 or 594-conjugated streptavidin (from Invitrogen at 1:250 to label BDA traced CST axons).

Techniques: Expressing, Labeling, Electron Microscopy

Journal: Journal of Neurophysiology

Article Title: Similarities and differences in circuit responses to applied Gly 1 -SIFamide and peptidergic (Gly 1 -SIFamide) neuron stimulation

doi: 10.1152/jn.00567.2018

Figure Lengend Snippet: One of the Gly1-SIFamide-IR neurons projecting from the CoGs to the STG is the identified projection neuron MCN5. Ai: Gly1-SIFamide-IR in the stn is limited to two pairs of closely bound axons which project between the anterior ganglia (CoGs and OG) and the STG. Aii: CabTRP Ia immunolabeling (α-substance P) in the stn labels two axons, shown previously to be the axons of the paired CoG projection neuron MCN1 (modulatory commissural neuron 1; Blitz et al. 1999). Aiii: a composite of the Gly1-SIFamide-IR and substance P-IR shows that these two labels are in distinct axons. As MCN5 and MCN1 are the only neurons that travel through the ion, these observations strongly suggest that the Gly1-SIFamide-IR CoG neuron that projects through the ion and stn is MCN5. Scale bar in Aiii = 25 µm and applies to A, i–iii. Bi: a Neurobiotin backfill of the stn with the ions intact and sons bisected labeled two neurons. The same preparation was subsequently labeled with antibodies against Gly1-SIFamide (Bii) and substance P (Biii). Biv: an overlay of the Neurobiotin labeling with Gly1-SIFamide-IR and substance P-IR demonstrates that one Neurobiotin labeled neuron colabels with α-SIFamide and the other colabels with α-substance P. The unfilled arrow points to the same neuron in all images, identified as MCN1 (Blitz et al. 1999; Christie et al. 1997b; Coleman and Nusbaum 1994). The filled arrow points to the same neuron in all images, identified as MCN5 (Norris et al. 1996). Scale bar in Biv = 100 µm and applies to Bi–iv.

Article Snippet: However, this latter rhythm does not include prolonged IC neuron bursting and the slowed pyloric rhythm results instead from the gastric mill protractor neuron LG inhibition of the MCN1 axon terminals ( Bartos and Nusbaum 1997 ; Coleman and Nusbaum 1994 ).

Techniques: Immunolabeling, Labeling

Journal: The Journal of Neuroscience

Article Title: Neurotransmitters and Motoneuron Contacts of Multifunctional and Behaviorally Specialized Turtle Spinal Cord Interneurons

doi: 10.1523/JNEUROSCI.2200-19.2020

Figure Lengend Snippet: Antibody combinations used in the study

Article Snippet: Series of four-color confocal images were gathered at intervals of 0.3 μm to determine the neurotransmitter type associated with each cell and if contacts were made with motoneurons. table ft1 table-wrap mode="anchored" t5 Table 1. caption a7 Section content Primary antibody combination Primary antibody concentration Source Secondary antibodies Axon terminals Rabbit anti-Homer 1:500 M. Watanabe AlexaFluor 488 Mouse anti-gephyrin 1:500 Synaptic Systems AlexaFluor 647 Goat anti-ChAT 1:100 Millipore Pacific Blue Axon terminals Guinea pig anti-VGLUT2 1:5000 Millipore AlexaFluor 647 Rabbit anti-GAD OR 1:1000 Sigma-Aldrich AlexaFluor 488 Sheep anti-GlyT2 1:1000 Chemicon AlexaFluor 488 Goat anti-ChAT 1:100 Millipore Pacific Blue Motoneurons Goat anti-ChAT 1:100 Millipore AlexaFluor 647 Rabbit anti-FG 1:2000 Millipore Pacific Blue Open in a separate window Antibody combinations used in the study

Techniques: Concentration Assay

Journal: Journal of neuroscience research

Article Title: Polyethylene glycol treated allografts not tissue matched nor immunosuppressed rapidly repair sciatic nerve gaps, maintain neuromuscular functions, and restore voluntary behaviors in female rats

doi: 10.1002/jnr.24227

Figure Lengend Snippet: Primary antibodies used in this study.

Article Snippet: Target/Antigen Antibody Antigen Species Source Immunogen dilution RRID nerve terminals/Synaptic vesicle glycoprotein 2A SV2- mouse monoclonal Ommata (electric fish) DSHB, cat# SV2 purified synaptic vesicles 1:400 AB_2315385 neurofilament (NF-M) 2H3- mouse monoclonal Rat DSHB, cat# 2H3 neonate rat brain cytoskeletal preparations 1:400 AB_531793 Open in a separate window Primary antibodies used in this study.

Techniques: Purification

Journal: Journal of neuroscience research

Article Title: Polyethylene glycol treated allografts not tissue matched nor immunosuppressed rapidly repair sciatic nerve gaps, maintain neuromuscular functions, and restore voluntary behaviors in female rats

doi: 10.1002/jnr.24227

Figure Lengend Snippet: Primary antibodies used in this study.

Article Snippet: ​ Target/Antigen Antibody Antigen Species Source Immunogen dilution RRID nerve terminals/Synaptic vesicle glycoprotein 2A SV2- mouse monoclonal Ommata (electric fish) DSHB, cat# SV2 purified synaptic vesicles 1:400 AB_2315385 neurofilament (NF-M) 2H3- mouse monoclonal Rat DSHB, cat# 2H3 neonate rat brain cytoskeletal preparations 1:400 AB_531793 Open in a separate window Primary antibodies used in this study.

Techniques: Purification

Journal: bioRxiv

Article Title: Astrocytic synapse elimination controls ocular dominance plasticity

doi: 10.1101/2022.09.23.509193

Figure Lengend Snippet: (A, C) Schematic illustration of the experimental concept and schedule. At P14, all mice were injected with AAV9::EF1a-DIO-eYFP into the LGN and AAV1::hSyn-Cre into the right (contralateral, A) or left (ipsilateral, C) eye. To label contralateral retinogeniculate synapses in the left LGN, CTB-594 was injected into the right eye at P32. The mice were sacrificed at P35. (B, D) Representative z-stacked images of eYFP expression in the left LGN of right eye-injected (B) or left eye-injected (D) mice with AAV1::hSyn-Cre. Scale bars = 5 µm.

Article Snippet: To label the right eye-derived retinogeniculate axon terminals, CTB-594 (1 mg/ml, Thermo Fisher) was intraorbitally injected into the right eye of the injected mice at P32.

Techniques: Injection, Expressing