Journal: International Journal of Molecular Sciences

Article Title: Mechanical Stretch Activates TRPV4 and Hemichannel Responses in the Nonpigmented Ciliary Epithelium

doi: 10.3390/ijms24021673

Figure Lengend Snippet: Immunolocalization of TRPV4 in the mouse ciliary body. TRPV4 (green) is abundant in the NPE (nonpigmented ciliary epithelium) cell layer at the surface but not the underlying pigmented ciliary epithelium (PE) cell layer or stroma. The lower panel shows a negative control in which no primary antibody was used.

Article Snippet: The knockout validated, rabbit polyclonal anti-TRPV4 antibody was purchased from Alomone Lab (Catalog # ACC-034; Jerusalem, Israel).

Techniques: Negative Control

Journal: International Journal of Molecular Sciences

Article Title: Mechanical Stretch Activates TRPV4 and Hemichannel Responses in the Nonpigmented Ciliary Epithelium

doi: 10.3390/ijms24021673

Figure Lengend Snippet: The ATP release response to cyclic stretch was inhibited by the selective TRPV4 antagonists HC 067047 and RN-1734 but not by the TRPV1 antagonist A889425. The amount of ATP in the bathing medium was measured after subjecting monolayers of cultured NPE cells to 10% cyclic stretch (0.5 Hz) for 2 min in the presence of either HC 067047 (10 µM) ( A ) or RN-1734 (10 µM) ( B ) or A889425 (1 µM) ( C ) added 20 min beforehand. The data are the mean ± SE of results from six independent experiments. ** ( p < 0.01) and *** ( p < 0.001) indicate significant differences from control and ### ( p < 0.001) indicates significant differences from a stretch stimulus.

Article Snippet: The knockout validated, rabbit polyclonal anti-TRPV4 antibody was purchased from Alomone Lab (Catalog # ACC-034; Jerusalem, Israel).

Techniques: Cell Culture

Journal: International Journal of Molecular Sciences

Article Title: Mechanical Stretch Activates TRPV4 and Hemichannel Responses in the Nonpigmented Ciliary Epithelium

doi: 10.3390/ijms24021673

Figure Lengend Snippet: The connexin mimetic peptide Gap 27 prevents ATP release and PI uptake responses to TRPV4 activation by a selective agonist, GSK1016790A. The amount of ATP in the bathing medium ( A ) or cell PI fluorescence ( B ) was measured after exposing the monolayers of cultured NPE cells to GSK1016790A (10 nM) in the presence or absence of Gap 27 (200 µM) added 60 min beforehand. The data are the mean ± SE of results from six independent experiments. *** ( p < 0.001) indicates significant differences from control.

Article Snippet: The knockout validated, rabbit polyclonal anti-TRPV4 antibody was purchased from Alomone Lab (Catalog # ACC-034; Jerusalem, Israel).

Techniques: Activation Assay, Fluorescence, Cell Culture

Journal: International Journal of Molecular Sciences

Article Title: Mechanical Stretch Activates TRPV4 and Hemichannel Responses in the Nonpigmented Ciliary Epithelium

doi: 10.3390/ijms24021673

Figure Lengend Snippet: The TRPV4 antagonist HC067047 prevents ATP release and PI uptake responses to an osmotic swelling stimulus. The amount of ATP in the bathing medium ( A ) or cell PI fluorescence ( B ) was measured after exposing the monolayers of cultured NPE cells to hypoosmotic Kreb’s solution (200 mOsm) for 2 min in the presence or absence of HC067047 (10 µM) added 20 min beforehand. The data are the mean ± SE of results from twelve ( A ) or six ( B ) independent experiments. *** ( p < 0.001) indicates significant differences from control.

Article Snippet: The knockout validated, rabbit polyclonal anti-TRPV4 antibody was purchased from Alomone Lab (Catalog # ACC-034; Jerusalem, Israel).

Techniques: Fluorescence, Cell Culture

Journal: PLoS ONE

Article Title: Selective Targeting of TRPV1 Expressing Sensory Nerve Terminals in the Spinal Cord for Long Lasting Analgesia

doi: 10.1371/journal.pone.0007021

Figure Lengend Snippet: Immunofluroscent pictures from control animals (top panel) and animals treated with RTX (1.9 µg/kg) for 20 days (bottom panel). Representative pictures shown are from 3–5 sections stained at least from 3 rats for each group. A. Left panel shows complete loss of TRPV1 staining in spinal dorsal horn after RTX treatment. Right panel shows that there was no change in staining for NeuN, a neuronal marker. B. RTX treatment did not affect the staining or the number of DRG neurons stained. Immunofluroscent pictures of L4 DRG in control animals (top panel) and animals treated with RTX (bottom panel) show no difference in TRPV1 labeling between these two groups. C. TRPV1 staining of paw skin sections did not show a change between controls and following RTX treatment. Sections from another rat show intrathecal RTX eliminated TRPV1 staining (D), reduced CGRP staining significantly (E) and reduced SP staining significantly (F), but did not alter TRPV4 staining in spinal cord (G). The corresponding histograms of analysis of gray value of the stained region are shown below. The scale bar is 100 µm except for C it is 50 µm.

Article Snippet: The sections were incubated with polyclonal rabbit anti-TRPV1 antibody (Affinity BioReagents, PA1-747, 1∶500), or polyclonal rabbit anti-TRPV4 antibody (Alomone, ACC-034, 1∶200), or monoclonal mouse anti-CGRP antibody (Sigma, C-7113, 1∶2500), or polyclonal guinea pig anti-SP antibody (abcam, ab10353, 1∶1000), or monoclonal mouse anti-NeuN antibody (Chemicon, MAB377, 1∶100) for 1 hour at room temperature, then incubated with Rhodamine Red (TM)-X donkey anti-rabbit IgG (Jackson 711-295-152, 1∶100), or FITC donkey anti-mouse IgG (Jackson, 715-095-151, 1∶100), or Rhodamine Red (TM)-X donkey anti-guinea pig IgG (Jackson, 706-295-148, 1∶100) for 1 hour at room temperature.

Techniques: Staining, Marker, Labeling