Journal: Biology Open

Article Title: P2X 7 regulates ependymo-radial glial cell proliferation in adult Danio rerio following spinal cord injury

doi: 10.1242/bio.060270

Figure Lengend Snippet: P2X 7 receptors were expressed by GFAP + ependymo-radial glial cells. Spinal cord cross sections are shown (dorsal is up; asterisk indicates central canal; scale bar: 50 μm). Representative images of GFAP and P2X 7 labelling in naïve fish (A-C′), at 7 dpi (D-F′), and at 14 dpi (G-I′). Colocalization (arrows) was seen in radial processes and cell bodies (C′,F′,I′). GFAP + cells that did not colocalize with P2X 7 were also detected (open arrowheads) (C′,F′,I′). n =3/group. Created with BioRender.com.

Article Snippet: This included a P2X 7 control antigen (Alomone Labs, BLP-PR004, Q64663).

Techniques: Immunofluorescence

Journal: Biology Open

Article Title: P2X 7 regulates ependymo-radial glial cell proliferation in adult Danio rerio following spinal cord injury

doi: 10.1242/bio.060270

Figure Lengend Snippet: P2X 7 receptors were expressed by PCNA + proliferating cells. Spinal cord cross sections are shown (dorsal is up; asterisk indicates central canal; scale bar: 50 μm). Representative images of PCNA + and P2X 7 labelling in naïve fish (A-C′), at 7 dpi rostral (D-F′), and at 7 dpi caudal (G-I′). Colocalization (arrows) was seen in some cells (C′,F′,I′). PCNA + cells that did not colocalize with P2X 7 were also detected (open arrowheads) (C′,F′,I′). n =3/group. Created with BioRender.com.

Article Snippet: This included a P2X 7 control antigen (Alomone Labs, BLP-PR004, Q64663).

Techniques: Immunofluorescence

Journal: Biology Open

Article Title: P2X 7 regulates ependymo-radial glial cell proliferation in adult Danio rerio following spinal cord injury

doi: 10.1242/bio.060270

Figure Lengend Snippet: P2X 7 receptors were expressed by HuC/D + neurons. Spinal cord cross sections are shown (dorsal is up; asterisk indicates central canal; scale bar: 50 μm). Representative images of HuC/D + and P2X 7 labelling in naïve fish (A-C′), at 14 dpi rostral (D-F′), and at 14 dpi caudal (G-I′). Colocalization (arrows) was seen in larger mature neurons and smaller immature neurons (C′,F′,I′). n =3/group. Created with BioRender.com.

Article Snippet: This included a P2X 7 control antigen (Alomone Labs, BLP-PR004, Q64663).

Techniques: Immunofluorescence

Journal: Biology Open

Article Title: P2X 7 regulates ependymo-radial glial cell proliferation in adult Danio rerio following spinal cord injury

doi: 10.1242/bio.060270

Figure Lengend Snippet: Temporal changes in P2X 7 protein expression following SCI in adult zebrafish. Schematic representations of naïve and injured spinal cord areas of analysis (A-B), as well as tissue collection timeline (C). Representative western blot and corresponding total protein in naïve zebrafish (Zf) spinal cord tissue, mouse (Ms) hippocampal tissue as a positive control (+), and pre-absorption of naïve zebrafish spinal cord tissue with a blocking peptide as a negative control (-) (D). Representative western blot and corresponding total protein at 1 dpi (E), 7 dpi (G), and 14 dpi (I). Quantitative analysis of the 50 kDa isoform showed significant downregulation within the injury (In, n =10, one-way ANOVA with Dunnett's post hoc test, P =0.0079) and caudal to the lesion (Ca, n =9, P =0.0046) at 7 dpi when normalized to naïve tissue ( n =9) (H). Protein expression of the 50 kDa isoform showed basal levels of expression at 1 dpi (F) and 14 dpi (J). Data presented as means±s.d. **, significant differences, P <0.01. Created with BioRender.com.

Article Snippet: This included a P2X 7 control antigen (Alomone Labs, BLP-PR004, Q64663).

Techniques: Expressing, Analysis, Tissue, Western Blot, Positive Control, Blocking Assay, Negative Control

Journal: Scientific Reports

Article Title: Transient receptor potential vanilloid type 1 is expressed in the horizontal pathway of the vervet monkey retina

doi: 10.1038/s41598-020-68937-9

Figure Lengend Snippet: List of antibodies.

Article Snippet: For the control condition, the same protocol was ran simultaneously as described, excepting that the anti-TRPV1 primary antibody was preincubated with its blocking peptide (BP; #NB100-1617PEP, Novus Biologicals, Littleton, CO, USA) in a 1:10 dilution for 1 h.

Techniques: Purification, Muscles, Clinical Proteomics

Journal: Scientific Reports

Article Title: Transient receptor potential vanilloid type 1 is expressed in the horizontal pathway of the vervet monkey retina

doi: 10.1038/s41598-020-68937-9

Figure Lengend Snippet: Characterization of the TRPV1 antibody in the monkey retina. Western blot analysis of total protein samples from a vervet monkey retina, showing detection of the expected protein band at 100 kDa ( A ). The band was not detected when the antibody was pre-incubated with its corresponding TRPV1 blocking peptide (BP) at the ratio 1:10 ( A ). All lanes contained 10 μg of total protein. Immunohistochemistry on vervet retinal tissue with the antibody raised against TRPV1 revealed a unique staining profile ( B ). When the TRPV1 antibody was pre-incubated with its corresponding BP, it revealed an absence of staining in the vervet retinas ( C ). Quantitative analysis of the TRPV1 fluorescent signal through the six different retinal layers reveals a relatively higher signal intensity in the OPL and the GCL layers compared to the photoreceptor (PRL) and the outer nuclear (ONL) layers (P < 0.05) ( D ). TRPV1 transient receptor potential vanilloid type 1, BP blocking peptide, LC loading control, PRL photoreceptor layer, ONL outer nuclear layer, OPL outer plexiform layer, INL inner nuclear layer, IPL inner plexiform layer, GCL ganglion cell layer. Scale bar = 75 μm. Mean optical density ratios ± standard error of the mean (SEM) are depicted.

Article Snippet: For the control condition, the same protocol was ran simultaneously as described, excepting that the anti-TRPV1 primary antibody was preincubated with its blocking peptide (BP; #NB100-1617PEP, Novus Biologicals, Littleton, CO, USA) in a 1:10 dilution for 1 h.

Techniques: Western Blot, Incubation, Blocking Assay, Immunohistochemistry, Staining, Control

Journal: Scientific Reports

Article Title: Transient receptor potential vanilloid type 1 is expressed in the horizontal pathway of the vervet monkey retina

doi: 10.1038/s41598-020-68937-9

Figure Lengend Snippet: Labeling pattern of TRPV1-IR throughout the vervet retina. Confocal micrographs taken from the center ( A ), middle ( B ), and peripheral retina ( C ). Quantitative analysis of the TRPV1 fluorescent signal through the three distinct retinal eccentricities shows relatively higher central signal intensity in the IPL and the RGC layers, and greater peripherical labeling in the OPL and the ONL layers (P < 0.05) ( D ). PRL photoreceptor layer, ONL outer nuclear layer, OPL outer plexiform layer, INL inner nuclear layer, IPL inner plexiform layer, GCL ganglion cell layer, GCFL ganglion cell fiber layer. Scale bar = 75 µm. Mean optical density ratios ± standard error of the mean (SEM) are depicted.

Article Snippet: For the control condition, the same protocol was ran simultaneously as described, excepting that the anti-TRPV1 primary antibody was preincubated with its blocking peptide (BP; #NB100-1617PEP, Novus Biologicals, Littleton, CO, USA) in a 1:10 dilution for 1 h.

Techniques: Labeling

Journal: Scientific Reports

Article Title: Transient receptor potential vanilloid type 1 is expressed in the horizontal pathway of the vervet monkey retina

doi: 10.1038/s41598-020-68937-9

Figure Lengend Snippet: Double-label immunofluorescence illustrating immunolabeling of TRPV1 in ganglion cells, horizontal cells, and amacrine cells. ( A-C ) The antibody against Brn3a labels the nucleus of ganglion cells in the monkey retina, and these cells were strongly TRPV1 immunoreactive. Arrows indicate Brn3a-positive ganglion cells that are TRPV1 immunoreactive. Each protein expression is presented alone in grayscale: Brn3a ( A ) and TRPV1 ( B ); then, the two are presented merged ( C ), Brn3a in green and TRPV1 in red). ( D-I ) TRPV1 is expressed in horizontal cell synaptic terminals (arrows in OPL) and cell bodies (arrowheads in the INL). Each expression is presented alone in grayscale: PV ( D,G ), TRPV1 ( E,H ); then, the two are presented merged ( F,J ). ( J – L ) Arrows show the expression of TRPV1 in the amacrine cell bodies (INL). Each expression is presented alone in grayscale: syntaxin-positive amacrine cells ( J ) and TRPV1 ( K ); then, the two are presented merged ( L ). INL inner nuclear layer, IPL inner plexiform layer, OPL outer plexiform layer, GCL ganglion cell layer. Scale = 75 μm for ( A – C ) and ( J – L ), 30 μm for ( D – F ) and 10 μm for ( G – I ).

Article Snippet: For the control condition, the same protocol was ran simultaneously as described, excepting that the anti-TRPV1 primary antibody was preincubated with its blocking peptide (BP; #NB100-1617PEP, Novus Biologicals, Littleton, CO, USA) in a 1:10 dilution for 1 h.

Techniques: Immunofluorescence, Immunolabeling, Expressing

Journal: Scientific Reports

Article Title: Transient receptor potential vanilloid type 1 is expressed in the horizontal pathway of the vervet monkey retina

doi: 10.1038/s41598-020-68937-9

Figure Lengend Snippet: Double-label immunofluorescence showing no signal for TRPV1 in rod bipolar cells and Müller cells. TRPV1 is neither expressed in the cell bodies (small arrows in the upper INL) of bipolar cells nor their synaptic terminals at the edge of the OPL (arrowheads). Each protein expression is presented alone in grayscale: PKC ( A ) and TRPV1 ( B ); then, the two are presented merged ( C ), PKC in green and TRPV1 in red. Long arrows show specifically that the Müller cell bodies do not express TRPV1. Each protein expression is presented alone in grayscale: glutamine synthetase ( D ) and TRPV1 ( E ); then, the two are presented merged ( F ), glutamine synthetase in green and TRPV1 in red. Scale = 75 μm for ( A – F ).

Article Snippet: For the control condition, the same protocol was ran simultaneously as described, excepting that the anti-TRPV1 primary antibody was preincubated with its blocking peptide (BP; #NB100-1617PEP, Novus Biologicals, Littleton, CO, USA) in a 1:10 dilution for 1 h.

Techniques: Immunofluorescence, Expressing

Journal: Scientific Reports

Article Title: Transient receptor potential vanilloid type 1 is expressed in the horizontal pathway of the vervet monkey retina

doi: 10.1038/s41598-020-68937-9

Figure Lengend Snippet: Mapping of the receptors CB1R, CB2R, GPR55, and TRPV1 in the monkey retina. These receptors are differently expressed in the retina of monkeys. These results are compiled from several published articles , , . CB1R is represented in green, CB2R in magenta, GPR55 in blue, and TRPV1 in red. OS, photoreceptors outer segments; IS, photoreceptors inner segments; ONL, outer nuclear layer; ONL, outer plexiform layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer.

Article Snippet: For the control condition, the same protocol was ran simultaneously as described, excepting that the anti-TRPV1 primary antibody was preincubated with its blocking peptide (BP; #NB100-1617PEP, Novus Biologicals, Littleton, CO, USA) in a 1:10 dilution for 1 h.

Techniques:

Journal: Scientific Reports

Article Title: Transient receptor potential vanilloid type 1 is expressed in the horizontal pathway of the vervet monkey retina

doi: 10.1038/s41598-020-68937-9

Figure Lengend Snippet: Schematic illustration of the hypothetical mechanism of action of TRPV1 in horizontal cells of the monkey retina following stimulation of a cone. The different steps are highlighted by numbers. (1) When a cone is stimulated (C1), endocannabinoids (eCBs), particularly AEA and 2-AG, are produced in the synaptic cleft. (2) TRPV1 receptors are activated by eCBs in horizontal cells. (3) The calcium influx through the membrane causes depolarization of the horizontal cell. (4) GABA release in the synaptic cleft. (5) GABA binds to GABA receptors on the presynaptic neighboring cone pedicle (5). This mechanism of lateral inhibition could also potentially modulate the information transmitted to the bipolar cells and retinal ganglion cells. S spot stimulus, C1 stimulated cone 1, C2 adjacent cone 2, H horizontal cell, eCB endocannabinoid, GABAr Gamma-amino-butyric acid receptors, TRPV1 Transient receptor potential vanilloid type 1.

Article Snippet: For the control condition, the same protocol was ran simultaneously as described, excepting that the anti-TRPV1 primary antibody was preincubated with its blocking peptide (BP; #NB100-1617PEP, Novus Biologicals, Littleton, CO, USA) in a 1:10 dilution for 1 h.

Techniques: Produced, Membrane, Inhibition

Journal: Experimental gerontology

Article Title: Age-related impairment of autophagy in cervical motor neurons

doi: 10.1016/j.exger.2020.111193

Figure Lengend Snippet: A) Autophagy comprises a series of evolutionarily conserved phases by which cells target damaged intracellular components for degradation in lysosomes. During the initiation phase, damaged organelles trigger activation of the BCL-2-interacting protein Becn-1 (Becn1) whereas proteins targeted for degradation become ubiquitinated. During the elongation phase of autophagy, recruitment of ATG7 and the ATG5/12 complex results in the lipidation of LC3 (reflected in the conversion of LC3-I to LC3-II) and its incorporation into nascent autophagic vacuole membranes. Damaged organelles and ubiquitinated proteins are also incorporated into autophagic vacuoles via the specific autophagy receptor protein p62. Once the mature autophagic vacuole membrane envelops its cargo, it is then delivered to the lysosome, where all its contents are degraded, including p62. Accordingly, degradation of p62 is used to monitor the degradation phase. Figure modified from (Gonzalez Porras et al., 2018). B) Higher expression of Becn1, ATG7, the ATG5/12 complex and LC3 reflect increased production of autophagosomes, whereas an accumulation of p62 represents lack of corresponding autophagosome degradation. The figure represents the conceptual framework for the interpretation of changes in the expression of autophagy markers as a result of aging or other conditions.

Article Snippet: When multiple bands were evidenced in a blot (LC3B and ATG5 antibodies), the available blocking peptides for LC3B (NB100-2220PEP, 1:100; Novus) and ATG5 (NB110-53818PEP, 1:100; Novus) were used to detect the specific bands of interest.

Techniques: Activation Assay, Membrane, Modification, Expressing

Journal: Experimental gerontology

Article Title: Age-related impairment of autophagy in cervical motor neurons

doi: 10.1016/j.exger.2020.111193

Figure Lengend Snippet: Expression of the initiation and elongation autophagy markers Becn-1 (A), ATG 7 (B) and the ATG5/12 complex (C) as determined using Western blotting in the cervical spinal cord of mice across age groups. Bar graphs show mean ± 95% CI of individual measurements per animal (top) and corresponding representative blots (bottom). There were no significant differences in the protein expression levels of these autophagy markers across age groups.

Article Snippet: When multiple bands were evidenced in a blot (LC3B and ATG5 antibodies), the available blocking peptides for LC3B (NB100-2220PEP, 1:100; Novus) and ATG5 (NB110-53818PEP, 1:100; Novus) were used to detect the specific bands of interest.

Techniques: Expressing, Western Blot