Journal: Frontiers in Physiology

Article Title: Plasticity of TRPV1-Expressing Sensory Neurons Mediating Autonomic Dysreflexia Following Spinal Cord Injury

doi: 10.3389/fphys.2012.00257

Figure Lengend Snippet: High-thoracic (T3) spinal cord injury-induced selective hypertrophy of sensory neurons expressing the capsaicin receptor (TRPV1) and the artemin receptor (GFRα3) in the L4/L5 DRG . (A) Substance P (SP) – positive DRG neurons and their size-frequency distributions. (B) IB4-binding DRG neurons. (C) P2X3-positive DRG neurons. (D) TRPV1-positive DRG neurons. (E) GFRα3-expressing DRG neurons, known to express TRPV1. The overall proportions of immunopositive neurons [insets (A–E) ] did not change for any subpopulation. Ganglia were harvested 3 months after sham-injury (gray) or complete T3 SCI (black). Scale bar = 50 μm. Asterisks indicate P < 0.05, K–S goodness-of-fit test.

Article Snippet: For immunohistochemistry, slides were incubated in 10% normal donkey serum in PBS plus Triton X-100 (0.1%) for 20 min. We used five antibodies to delineate different subsets of nociceptors: these included antibodies raised against TRPV1 (Neuromics, Edina, MN, USA; 1:2,000), the ionotropic ATP purinoceptor P2X 3 (Millipore, Billerica, MA, USA; 1:1,000), Substance P (Neuromics, 1:1,000), the glycoprotein isolectin B4 (IB4; Neuromics; 1:2,000), the glial cell line-derived neurotrophic factor family receptor α 3 (GFRα3; R&D Systems; 1:500).

Techniques: Expressing, Binding Assay

Journal: PloS one

Article Title: Chemoreception regulates chemical access to mouse vomeronasal organ: role of solitary chemosensory cells.

doi: 10.1371/journal.pone.0011924

Figure Lengend Snippet: Figure 1. SCCs preferentially locate at the entrance duct of the VNO and express chemosensory signaling components. A: A schematic drawing of a mouse hemi-nose. MOE: main olfactory epithelium; OB: olfactory bulb. VNO: vomeronasal organ (blue). B: Luminal view of the entire non- sensory epithelium and entrance duct of a VNO from a TRPM5-GFP mouse. Bright spots are GFP-positive SCCs. Arrow points to the anterior opening. Anterior to the VNO, the cartilaginous stenonii canal channels external fluids to the VNO opening. C: Plot of SCC density at different regions as determined from horizontal VNO sections of four mice (Mean 6 SEM), showing that the GFP-expressing SCCs preferentially reside at the entrance duct and adjacent 0.5 mm long anterior non-sensory epithelium. D: Confocal image of a typical GFP-expressing SCC. Arrowhead points to an apical microvillus. E: Immunolabeling of TRPM5 (red) in GFP-expressing cells (green) in a VNO section. F: Image taken from an epithelial strip from the entrance duct, showing that TRPM5 (GFP) expressing SCCs immunoreacted to an anti-a-gustducin antibody (red). Scales: B, 0.5 mm; D, 5 mm; E and F, 20 mm. doi:10.1371/journal.pone.0011924.g001

Article Snippet: Sections were then incubated 12 to 72 hours with primary antibodies against each of the following proteins: TRPM5 (1:250), c13 (1:500), both were provided kindly by Dr. RF Margolskee [61], a-gustducin (1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), substance P (1:1000, Chemicon, Temecula, CA), PGP 9.5 (ubiquitin carboxyl-terminal hydrolase; 1:500, Chemicon), ChAT (1:100; Chemicon), PLCb2 (1:200; Santa Cruz Biotechnology), and VAChT (1:200, Sigma).

Techniques: Expressing, Immunolabeling, Stripping Membranes

Journal: PloS one

Article Title: Chemoreception regulates chemical access to mouse vomeronasal organ: role of solitary chemosensory cells.

doi: 10.1371/journal.pone.0011924

Figure Lengend Snippet: Figure 2. Trigeminal innervation and immuno-expression of ChaT and VAChT for ACh synthesis and packaging in SCCs. A: Confocal image of a VNO epithelial strip from a TRPM5-GFP mouse, showing both PGP 9.5-labeled trigeminal nerve bundles (asterisks) at basal lamina and many fine intraepithelial fibers. Arrowheads point to varicosities found typically in peptidergic fibers. Arrow points to a branching nerve fiber. B: The GFP-fluorescence image overlaid with (A). All TRPM5-expressing SCCs are apposed or wrapped closely by one or a few intraepithelial nerve fibers. Arrows point to apexes of SCCs. C: immunolabeling of substance P in a section of the anterior non-sensory epithelium. Note that most of the labeled intraepithelial fibers appear to innervate SCCs. D: Percentages of intraepithelial fibers innervating SCCs. E and F: Confocal images of TRPM5- expressing SCCs (green) immunoreacted to antibodies against the ChAT and VAChT respectively (red). G: Whole-mount fluorescence image of the ChAT (GFP)-expressing cells taken from a VNO entrance duct of a ChAT(BAC)-eGFP mouse. H: CHAT (GFP)-expressing cells of the VNO immunolabeled by the anti-a-gustducin antibody (red). Scales: B, F, G, and H, 10 mm; C, 50 mm; E, 20 mm. doi:10.1371/journal.pone.0011924.g002

Article Snippet: Sections were then incubated 12 to 72 hours with primary antibodies against each of the following proteins: TRPM5 (1:250), c13 (1:500), both were provided kindly by Dr. RF Margolskee [61], a-gustducin (1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), substance P (1:1000, Chemicon, Temecula, CA), PGP 9.5 (ubiquitin carboxyl-terminal hydrolase; 1:500, Chemicon), ChAT (1:100; Chemicon), PLCb2 (1:200; Santa Cruz Biotechnology), and VAChT (1:200, Sigma).

Techniques: Expressing, Stripping Membranes, Labeling, Fluorescence, Immunolabeling