Journal: Physiological Reports

Article Title: The endocannabinoid systemʼs involvement in motor development relies on cannabinoid receptors, TRP channels, and Sonic Hedgehog signaling

doi: 10.14814/phy2.15565

Figure Lengend Snippet: Blocking TRP channels with AMG 9090 prevents the escape swimming deficits caused by dual FAAH/MAGL inhibition, but not deficits caused by singular MAGL inhibition. Escape swimming in response to an acute touch stimulus was assessed in zebrafish embryos at 2 days post‐fertilization (dpf). The effect of ~24‐h drug treatment on embryonic escape swimming distance (cm) and mean velocity (cm/s) was measured, and mean ranks of treatment groups were compared against each other. In each respective treatment group, the horizontal line within its boxplot represents the median value which is surrounded by the interquartile range, while the error bars represent the maximum and minimum values. (A, B) The effects of the vehicle control ( n = 35), 5 μM JZL 184 ( n = 33), 0.01 μM AMG 9090 ( n = 40), and 5 μM JZL 184 in the presence of 0.01 μM AMG 9090 ( n = 40) on escape swimming was compared. (C, D) The effects the vehicle control ( n = 35), 2 μM JZL 195 ( n = 38), 0.01 μM AMG 9090 ( n = 40), and 2 μM JZL 195 in the presence of 0.01 μM AMG 9090 ( n = 39) on escape swimming was compared. (E) Representative tracings of the swimming paths are shown for vehicle, 0.01 μM AMG 9090, 5 μM JZL 184, 5 μM JZL 184 + 0.01 μM AMG 9090, 2 μM JZL 195 and 2 μM JZL 195 + 0.01 μM AMG 9090. N = 4 experiments with 8–10 animals per experiment for each treatment. Columns which share the same letter(s) of the alphabet are not statistically different from one another (Kruskal–Wallis, followed by Dunn's multiple comparisons test—where statistical significance was determined as p < 0.05).

Article Snippet: Along with these compounds, animals were co‐treated with the CB1R antagonist AM 251 (Selleck Chemicals), the CB2R antagonist AM630 (Adooq Bioscience), the TRPV1/TRPA1/TRPM8 inhibitor AMG 9090 (Alomone Labs), or the SMO agonist Purmorphamine (Sigma‐Aldrich).

Techniques: Blocking Assay, Inhibition

Journal: PLoS ONE

Article Title: LIV-1 Promotes Prostate Cancer Epithelial-to-Mesenchymal Transition and Metastasis through HB-EGF Shedding and EGFR-Mediated ERK Signaling

doi: 10.1371/journal.pone.0027720

Figure Lengend Snippet: A, LIV-1 overexpressing cells (LIV#8) and control cells (con1) were treated with EGF and AG1478 for 2 hours. Western blotting showed that the EGF-elicited EGFR and ERK phosphorylation was blocked by the AG1478 inhibitor. B, RT-PCR showed increased HB-EGF, MMP2 and MMP9 expression in LIV-1 overexpressing cells. C, LIV-1 overexpressing cells (LIV#8 and LIV#14) were cultured in serum-free medium for 24 hours and the culture media were used to determine the MMP2 and MMP9 enzymatic activity by zymogram assay. D, the effect of MMP2/9 enzymatic activity on HB-EGF shedding was evaluated by ELISA. LIV-1 overexpressing clones (LIV#8 and LIV#14) secreted more HB-EGF than control clones (con1 and con2), and the secretion was reduced by MMP2/9 inhibition. ∗ indicates statistical significance compared to the control of the group ( P <0.05). E, Western blotting showed that inhibition of MMP2/9 activity suppressed EGFR and the downstream ERK phosphorylation. F, LIV-1 overexpressing cells were treated with MMP 2/9 inhibitor III for 24 hours in Transwell motility assays. Both migration and invasion of the treated cells were decreased. * indicates statistical significance compared to the control group ( P <0.05). G, diagram depicts the proposed role of LIV-1 in prostate cancer cell EMT and metastasis.

Article Snippet: Tyrphostin AG1478, U0126 and MMP2/9 inhibitor III were obtained from Alomone labs (Jerusalem, Israel), Cell Signaling Technology (Danvers, MA), and Calbiochem (Darmstadt, Germany), respectively.

Techniques: Western Blot, Reverse Transcription Polymerase Chain Reaction, Expressing, Cell Culture, Activity Assay, Enzyme-linked Immunosorbent Assay, Clone Assay, Inhibition, Migration

Journal: PLoS ONE

Article Title: A System for Performing High Throughput Assays of Synaptic Function

doi: 10.1371/journal.pone.0025999

Figure Lengend Snippet: ( A ) TTX concentration-response curve for response amplitudes to a 30 Hz, 10 sec pulse train from a single plate. Each point shows the mean ± SEM of 8 wells normalized to within-plate vehicle controls. Data were fit with a standard sigmoid concentration-response function (R 2 = 0.99). ( B ) SypHy fluorescence responses during a 30 Hz, 10 second stimulus train in the presence of the Cav2.1 inhibitor ω-agatoxin IVA (500 nM), the Cav2.2 inhibitor ω-conotoxin GVIA (1 µM), and the Cav2.3 inhibitor SNX-482 (1.2 µM). The waveforms depicted are an average of 24 wells for the vehicle and 8 wells for the treatment group. ( C,D ) Amplitudes (mean ± SEM) of the sypHy responses shown in (B) following (C) 30 pulses (1 sec) and (D) 300 pulses (10 sec) of the stimulus train (***: p<0.0001).

Article Snippet: Compounds, including TTX (Tocris) and the Ca ++ channel inhibitors ω-agatoxin IVA, ω-contoxin GVIA, and SNX-482 (Alomone Labs), were added with the final buffer addition of the plate wash process.

Techniques: Concentration Assay, Fluorescence