Journal: Neuroscience

Article Title: Extracellular Protons Mediate Presynaptic Homeostatic Potentiation at the Mouse Neuromuscular Junction

doi: 10.1016/j.neuroscience.2021.01.036

Figure Lengend Snippet: ASIC1 and ASIC2a are located at the mouse NMJ. A. All of the images shown are maximum projections of 16 images collected at 0.5 μm increments vertically through the field containing the NMJ. α-bungarotoxin (α -BTX) is shown in Red. ASIC2a antibodies are in Green. Arrows point to clusters of ASIC2a staining that are outside the area defining the motor nerve terminal but near perisynaptic Schwann cell nuclei, indicated by the DAPI stain (Blue). Calibration bar, 10 μm. B. All of the images shown are maximum projections of 18 images collected at 0.5 μm increments vertically through the field containing the NMJ. α-bungarotoxin (α -BTX) is shown in Red. ASIC1 antibodies are in Green. Arrows point to clusters of ASIC2a staining that are outside the area defining the motor nerve terminal but near perisynaptic Schwann cell nuclei, indicated by the DAPI stain (Blue). Calibration bar, 10 μm.

Article Snippet: For ASIC1 staining we paired the guinea pig primary anti-ASIC1 antibody (AGP-053; Alomone Labs) with Goat Anti-Guinea pig Alexa Fluor 488 IgG (H±L) secondary antibody (A109FN).

Techniques: Staining

Journal: Stem Cells and Development

Article Title: Acid-Sensitive Ion Channels Are Expressed in Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes

doi: 10.1089/scd.2018.0234

Figure Lengend Snippet: mRNA expression of ASIC family in hiPSC differentiated cardiomyocytes. (A) Expression of ASIC1 and ASIC2 transcripts in human iPSC-derived cardiac clusters (IMR-90 line) and control cell lines. Beating hiPSC-derived cardiac clusters were collected on days 30 and 100 of differentiation using END2 protocol. hASIC1 mRNA was detected in undifferentiated hiPSCs and hiPSC-CMs (day 30), but was very low in day 100 hiPSC-CM. hiPSC-derived NPCs and hiPSC-NPC-derived neurons were used as positive controls. Human ASIC2 transcripts were detected only in hiPSC-NPC-derived neurons but not in any stage of hiPSC-CMs. (B) ASIC1a, ASIC2a, and ASIC2b mRNA were detected by using the second set of primers(see Materials and Methods section) in 55 days old hiPSC-CMs derived from iPSC-K3 line. (C, D) Quantitative polymerase chain reaction data confirmed the expression of ASIC1a, ASIC2a, and ASIC3 mRNA in hiPSC-CMs derived from both K3 and IMR-90 iPSC lines, n = 3 and n = 4, respectively. The **p < 0.01, *p < 0.05 compared with day 15 by one-way ANOVA followed by Tukey's test. NPC, neural progenitor cell.

Article Snippet: The following primary antibodies were used: guinea pig polyclonal anti-ASIC1 (1:200, Cat. No. AGP-053; Alomone Labs), mouse monoclonal anti-α-actinin (1:800, Cat. No. A-7811, Clone: EA-53; Sigma), and rabbit polyclonal anti-MAP2 (1:200, Cat. No. ab32454; Abcam).

Techniques: Expressing, Derivative Assay, Real-time Polymerase Chain Reaction

Journal: Stem Cells and Development

Article Title: Acid-Sensitive Ion Channels Are Expressed in Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes

doi: 10.1089/scd.2018.0234

Figure Lengend Snippet: Assessment of ASIC1 expression in human iPSC-CMs using immunocytochemistry. Representative immunofluorescence panels show colocalization of ASIC1 (red) with the cardiomyocyte marker a-actinin (green) in human iPSC-derived cardiomyocytes both at day 20 and at day 62 of directed differentiation. The neuroblastoma SH-SY5Y cells were used as positive control and exhibit more intense ASIC1 signal than CMs. Cells incubated only with fluorescently labeled secondary antibodies did not produce any specific signals. Nuclei were counterstained with DAPI (blue). CMs were differentiated from NP0040 iPSC line. Scale bars: 50 μm. Color images are available online.

Article Snippet: The following primary antibodies were used: guinea pig polyclonal anti-ASIC1 (1:200, Cat. No. AGP-053; Alomone Labs), mouse monoclonal anti-α-actinin (1:800, Cat. No. A-7811, Clone: EA-53; Sigma), and rabbit polyclonal anti-MAP2 (1:200, Cat. No. ab32454; Abcam).

Techniques: Expressing, Immunocytochemistry, Immunofluorescence, Marker, Derivative Assay, Positive Control, Incubation, Labeling

Journal: Stem Cells and Development

Article Title: Acid-Sensitive Ion Channels Are Expressed in Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes

doi: 10.1089/scd.2018.0234

Figure Lengend Snippet: Immunohistochemical detection of ASIC1 in adult rat brain and heart tissue cryoslices. Representative immunofluorescence panels show colocalization of ASIC1 with the neuronal marker microtubule-associated protein 2 (Map2) in the adult rat brain but no ASIC1 expression in the adult rat heart muscle cells, which stained positive for the cardiomyocyte marker sarcomeric α-actinin (Actn2). Tissue specimens incubated only with fluorescently labeled secondary antibodies gave no specific signals. Nuclei were counterstained with Hoechst 33342. Scale bars: 100 μm. Color images are available online.

Article Snippet: The following primary antibodies were used: guinea pig polyclonal anti-ASIC1 (1:200, Cat. No. AGP-053; Alomone Labs), mouse monoclonal anti-α-actinin (1:800, Cat. No. A-7811, Clone: EA-53; Sigma), and rabbit polyclonal anti-MAP2 (1:200, Cat. No. ab32454; Abcam).

Techniques: Immunohistochemical staining, Immunofluorescence, Marker, Expressing, Staining, Incubation, Labeling

Journal: The Journal of Neuroscience

Article Title: Merkel Cell-Driven BDNF Signaling Specifies SAI Neuron Molecular and Electrophysiological Phenotypes

doi: 10.1523/JNEUROSCI.3781-15.2016

Figure Lengend Snippet: Marker expression for T7 DRG neurons in the absence of Merkel cells

Article Snippet: The primary antibodies used were mouse anti-Islet1/2 (1:50; catalog #39.4D5, Developmental Studies Hybridoma Bank), rabbit anti-NF200 (1:1000; N4142, Sigma-Aldrich), goat anti-TrkB (1:200; AF1494, R&D Systems), goat anti-TrkC [AF1404 (1:100) and BAF1404 (1:20), R&D Systems], rabbit anti-Ret (1:50; catalog #18121, Immuno-Biological Laboratories), rabbit anti-parvalbumin (PV; 1:1000; PV25, Swant), rabbit anti-CGRP (1:1000; T-4032, Peninsula Laboratories), rabbit anti-phospho-SMAD1/5/8 (1:250; catalog 9511, Cell Signaling Technology), goat anti-TBX3 (1:100; sc-31656, Santa Cruz Biotechnology), rat anti-Keratin8 (1:20, TROMA-1, Developmental Studies Hybridoma Bank), guinea pig anti-Asic1 (1:250; Alomone Labs), rabbit anti-δ opioid receptor (DOR; 1:500; Alomone Labs), and rabbit anti-γENaC (1:500; StressMarq Biosciences).

Techniques: Marker, Expressing

Journal: The Journal of Neuroscience

Article Title: Merkel Cell-Driven BDNF Signaling Specifies SAI Neuron Molecular and Electrophysiological Phenotypes

doi: 10.1523/JNEUROSCI.3781-15.2016

Figure Lengend Snippet: Merkel cell deletion alters the expression of SAI ion channel components. A, qPCR analysis of ion channels in P21 thoracic DRGs (n ≥ 8 mice/genotype at each age). B–G, Immunostaining for NF200, γENaC, Asic1, and DOR in T7 DRG sections from P21 control littermate and K14; Atoh1CKO mice. Asterisks indicate double-positive cells. H, Percentages of Islet1/2+ T7 DRG neurons that were NF200+ γENaC+, NF200+Asic1, and NF200+DOR+ in P21 control and K14; Atoh1CKO mice (n ≥ 3 mice/genotype). Error bars in graphs represent the SEM, and asterisks indicate statistically significant differences between genotypes. *p < 0.05, ***p < 0.001.

Article Snippet: The primary antibodies used were mouse anti-Islet1/2 (1:50; catalog #39.4D5, Developmental Studies Hybridoma Bank), rabbit anti-NF200 (1:1000; N4142, Sigma-Aldrich), goat anti-TrkB (1:200; AF1494, R&D Systems), goat anti-TrkC [AF1404 (1:100) and BAF1404 (1:20), R&D Systems], rabbit anti-Ret (1:50; catalog #18121, Immuno-Biological Laboratories), rabbit anti-parvalbumin (PV; 1:1000; PV25, Swant), rabbit anti-CGRP (1:1000; T-4032, Peninsula Laboratories), rabbit anti-phospho-SMAD1/5/8 (1:250; catalog 9511, Cell Signaling Technology), goat anti-TBX3 (1:100; sc-31656, Santa Cruz Biotechnology), rat anti-Keratin8 (1:20, TROMA-1, Developmental Studies Hybridoma Bank), guinea pig anti-Asic1 (1:250; Alomone Labs), rabbit anti-δ opioid receptor (DOR; 1:500; Alomone Labs), and rabbit anti-γENaC (1:500; StressMarq Biosciences).

Techniques: Expressing, Immunostaining