trpm3  (Alomone Labs)

Journal: Headache

Article Title: TRPM3 activation causes CGRP release in trigeminal neurons: Implications for migraine mechanisms

doi: 10.1111/head.15082

Figure Lengend Snippet: TRPM3 and CGRP localization in male and female rat and human trigeminal system tissues. Immunofluorescence staining for TRPM3 (green), CGRP (red), and nuclei (blue) is shown in TG, dura mater, and MCA from male (A–C) and female (D–F) rats, as well as in human dura mater and MMA (G‐L). In rat TG (A, B), TRPM3 and CGRP coexpression is observed in distinct neuronal populations; neurons with diffuse CGRP labeling (white arrows, one fletching) likely represent C‐fiber neurons. And neurons with CGRP localized to Golgi‐like structures (white arrows, two fletchings) likely represents Aδ‐neurons. In the rat dura mater (B, E), CGRP‐positive fibers are observed alongside TRPM3‐expressing smooth muscle in the MMA (blue arrows) in single fibers (white arrow, one fletching) or bundle of fibers (white arrows, three fletchings). In the MCA (C, F), TRPM3 is localized to smooth muscle cells (blue arrows) and endothelial cells (pink arrows), whereas CGRP‐positive fibers (white arrows, one fletching), likely originating from the TG, are located near the adventitia. In human dura mater and MMA (G–L), TRPM3 is predominantly expressed in the smooth muscle layer (M) and the adjacent A, whereas CGRP‐positive fibers (white arrows) are observed primarily in the A and near vascular structures. Scale bars represent 50 μm in A–F and 25 μm in G–L. Images are representative of n = 3–4 immunohistochemistry experiments for each condition. A, adventitia; CGRP, calcitonin gene‐related peptide; I, intima; M, media; MCA, middle cerebral artery; MMA, middle meningeal artery; TG, trigeminal ganglia; TRPM3, transient receptor potential melastatin‐3. [Color figure can be viewed at wileyonlinelibrary.com ]

Article Snippet: Primary antibodies targeting CGRP (ab81887, mouse, mono‐clonal, Abcam, UK) and TRPM3 (ACC‐050‐200UL, rabbit, polyclonal, Alomone Labs, Isreal) were diluted 1:200 in antibody diluent (PBS‐T containing 1% BSA) and incubated with the tissues overnight at +4°C.

Techniques: Immunofluorescence, Staining, Labeling, Expressing, Immunohistochemistry

Journal: Headache

Article Title: TRPM3 activation causes CGRP release in trigeminal neurons: Implications for migraine mechanisms

doi: 10.1111/head.15082

Figure Lengend Snippet: Vasodilation induced by CIM0216 in male and female MCA in the presence of vehicle, isosakuranetin, or fremanezumab. Cumulative concentration‐response curves to CIM0216 are shown for male (A) and female (B) MCA precontracted with U46619 (10 −7 M). Responses were measured in the presence of vehicle, isosakuranetin (TRPM3 antagonist), or fremanezumab (CGRP receptor monoclonal antibody). No differences in vasodilatory responses were observed between conditions in either male or female arteries. Data are expressed as a percentage of precontraction (100%), with each point representing the mean ± SEM ( n = 5). CGRP, calcitonin gene‐related peptide; MCA, middle cerebral artery; SEM, standard error of the mean; TRPM3, transient receptor potential melastatin‐3. [Color figure can be viewed at wileyonlinelibrary.com ]

Article Snippet: Primary antibodies targeting CGRP (ab81887, mouse, mono‐clonal, Abcam, UK) and TRPM3 (ACC‐050‐200UL, rabbit, polyclonal, Alomone Labs, Isreal) were diluted 1:200 in antibody diluent (PBS‐T containing 1% BSA) and incubated with the tissues overnight at +4°C.

Techniques: Concentration Assay

Journal: Headache

Article Title: TRPM3 activation causes CGRP release in trigeminal neurons: Implications for migraine mechanisms

doi: 10.1111/head.15082

Figure Lengend Snippet: Calcium imaging of CGRP‐expressing neurons in female TG reveals TRPM3‐dependent activation by CIM0216 and inhibition by isosakuranetin. (A, B) Representative calcium imaging fields (top) and corresponding heatmaps (bottom) showing normalized fluorescence intensity (Δ F / F min ) over time in response to vehicle, CIM0216 (3 μM or 30 μM), isosakuranetin (10 μM), or their coapplication. Arrows indicate the time of compound application. (A) CIM0216 induced a concentration‐dependent increase in intracellular calcium, whereas vehicle had no effect. (B) Isosakuranetin alone did not induce calcium responses and reduced responses when coapplied with CIM0216. (C–F) Traces of individual neuron calcium responses (light lines) and the average response (black line) to 3 μM CIM0216 (C), 30 μM CIM0216 (D), 3 μM CIM0216 + 10 μM isosakuranetin (E), and 30 μM CIM0216 + 10 μM isosakuranetin (F). (G) Quantification of the proportion of CGRP‐expressing neurons responding to each treatment. Bars represent the percentage of responding (colored) versus nonresponding (white) neurons. Asterisks (*) indicate increase in responders compared to vehicle; dollar signs ($) indicate reduction with isosakuranetin cotreatment; number signs (#) indicate difference between 3 and 30 μM CIM0216 (Fisher's exact test, p < 0.05, n = 9). CGRP, calcitonin gene‐related peptide; TG, trigeminal ganglia; TRPM3, transient receptor potential melastatin‐3. [Color figure can be viewed at wileyonlinelibrary.com ]

Article Snippet: Primary antibodies targeting CGRP (ab81887, mouse, mono‐clonal, Abcam, UK) and TRPM3 (ACC‐050‐200UL, rabbit, polyclonal, Alomone Labs, Isreal) were diluted 1:200 in antibody diluent (PBS‐T containing 1% BSA) and incubated with the tissues overnight at +4°C.

Techniques: Imaging, Expressing, Activation Assay, Inhibition, Fluorescence, Concentration Assay