synthetic blocking peptide  (Alomone Labs)

Journal: Frontiers in Cellular Neuroscience

Article Title: Expression of Piezo1 in the Trigeminal Neurons and in the Axons That Innervate the Dental Pulp

doi: 10.3389/fncel.2022.945948

Figure Lengend Snippet: Light micrographs showing the specificity of Piezo1 immunostaining in the rat trigeminal ganglion [TG; (A,B) ] and the human dental pulp (C,D) , the size distribution (E,F) , and the neurochemical characterization (G–L) of Piezo1+ neurons in the rat TG. (A–D) Immunofluorescent staining for Piezo1 in the rat TG (A, B) and the human dental pulp (C,D) . The Piezo1 immunostaining in the TG neurons and pulpal axons is completely abolished by preadsorption with the corresponding blocking peptide, confirming the specificity of the Piezo1 antibody. (E,F) Light micrograph showing Piezo1+ neurons in the TG (E) and their size distribution (F) . Piezo1 is expressed mostly in medium-sized and large neurons. (G-L) Double immunofluorescent staining for Piezo1 and CGRP (G) , IB4 (H) , or NF200 (I) , and the quantitative analysis of colocalization of Piezo1 and CGRP (J) , IB4 (K) , or NF200 (L) . Piezo1+ neurons frequently co-stain NF200 but seldomly for CGRP or IB4. Scale bars = 50 μm in (A–E, G–I) .

Article Snippet: To control for the specificity of the Piezo1 antibody, we processed tissues according to the above protocols, except that either the anti-Piezo1 antibody was omitted or a Piezo1 blocking peptide (Piezo1: BLP-PC087, Alomone Labs) was added following the recommendation of the manufacturer.

Techniques: Immunostaining, Staining, Blocking Assay

Journal: Frontiers in Cellular Neuroscience

Article Title: Expression of Piezo1 in the Trigeminal Neurons and in the Axons That Innervate the Dental Pulp

doi: 10.3389/fncel.2022.945948

Figure Lengend Snippet: Axons that express Piezo1 in the sensory root of the rat trigeminal ganglion. (A,B) Electron micrographs showing immunoreaction products for Piezo1 (arrowheads) in small myelinated axons (asterisks), large myelinated axons (double asterisks), and unmyelinated axons (arrows). (C,D) Histograms showing the fraction (C) and the size distribution (D) of the Piezo1+ small myelinated, large myelinated, and unmyelinated axons. Asterisks in (C) indicate significant differences ( P < 0.05) between the fraction of small myelinated axons and that of large myelinated or unmyelinated axons. Scale bars = 500 nm.

Article Snippet: To control for the specificity of the Piezo1 antibody, we processed tissues according to the above protocols, except that either the anti-Piezo1 antibody was omitted or a Piezo1 blocking peptide (Piezo1: BLP-PC087, Alomone Labs) was added following the recommendation of the manufacturer.

Techniques:

Journal: Frontiers in Cellular Neuroscience

Article Title: Expression of Piezo1 in the Trigeminal Neurons and in the Axons That Innervate the Dental Pulp

doi: 10.3389/fncel.2022.945948

Figure Lengend Snippet: Light micrographs of immunofluorescent staining for Piezo1 in axons in the rat (A,B) and the human dental pulp (C–G) , and double immunofluorescent staining for Piezo1 and CGRP (H) or NF200 (I) in the human dental pulp. (A,B) Piezo1 is expressed in a few axons in the rat radicular (A) and coronal (B) pulp. (C–G) Piezo1 is expressed in a few axons in the human radicular pulp (C) and the core of the coronal pulp (D) , but in numerous axons (arrows) that branch extensively in the peripheral pulp (E) , and in many axons “ascending” toward dentinal tubules (arrowheads in E ). Piezo1 is also expressed in odontoblasts in the peripheral pulp [arrowheads in (G) , (G) is an enlargement of the boxed area in (F) ]. (H,I) Double immunofluorescent staining for Piezo1 and CGRP (H) or NF200 (I) in the peripheral region of the human dental pulp; colocalization is represented in yellow. Piezo1 is expressed mostly in NF200+ axons that form a plexus in the peripheral pulp (arrows) and “ascend” toward the dentin (arrowheads). Scale bars = 20 μm.

Article Snippet: To control for the specificity of the Piezo1 antibody, we processed tissues according to the above protocols, except that either the anti-Piezo1 antibody was omitted or a Piezo1 blocking peptide (Piezo1: BLP-PC087, Alomone Labs) was added following the recommendation of the manufacturer.

Techniques: Staining

Journal: Frontiers in Cellular Neuroscience

Article Title: Expression of Piezo1 in the Trigeminal Neurons and in the Axons That Innervate the Dental Pulp

doi: 10.3389/fncel.2022.945948

Figure Lengend Snippet: Electron micrographs showing immunostaining for Piezo1 (arrowheads) in myelinated (asterisks) and unmyelinated axons (arrows) in the radicular (A) , the core of the coronal (B) , and the peripheral (C) regions of the human dental pulp, and a histogram showing the fraction of Piezo1+ myelinated and unmyelinated axons of all Piezo1+ axons in each region of the human dental pulp (D) , and an electron micrograph of myelinated axon showing shedding of its myelin sheath during its course in the rat dental pulp [ (E) , modified from Bae and Yoshida, ]. Piezo1+ myelinated axons are more frequent in the radicular pulp than in the core of the coronal pulp, and virtually all Piezo1+ axons are unmyelinated in the peripheral pulp, suggesting that most Piezo1+ myelinated axons in the radicular pulp lose their myelin and become unmyelinated in the peripheral pulp. This assumption can be supported by our previous study showing the shedding of myelin sheath from myelinated axon within the rat dental pulp (E) . Asterisks in (D) indicate significant differences ( p < 0.05) among the fraction of Piezo1+ small myelinated or unmyelinated axons in each of the three regions of human dental pulp. Arrowheads in (A–C) indicate immunoreaction product of Piezo1. Arrowheads in (E) indicate unmyelinated portion of the myelinated axon (asterisk). Scale bar = 1 μm.

Article Snippet: To control for the specificity of the Piezo1 antibody, we processed tissues according to the above protocols, except that either the anti-Piezo1 antibody was omitted or a Piezo1 blocking peptide (Piezo1: BLP-PC087, Alomone Labs) was added following the recommendation of the manufacturer.

Techniques: Immunostaining, Modification