All electrophysiological experiments were performed as described previously23 (link). Agatoxin, isradipine, SNX 482, stromatoxin and iberiotoxin were purchased from Alomone (Jerusalem, Israel) and tetrodotoxin (TTX) from Sigma (Gillingham, UK).
Snx 482
Manufactured by Alomone
Sourced in Israel
SNX-482 is a synthetic, biologically active peptide that selectively blocks R-type calcium channels. It acts as a potent and selective antagonist of R-type calcium channels, with high affinity for Cav2.3 subunits. SNX-482 is commonly used in research applications to investigate the physiological roles of R-type calcium channels.
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Lab products found in correlation
ω conotoxin gvia, by Alomone (4 mentions)
ω agatoxin iva, by Alomone (3 mentions)
Isradipine, by Alomone (1 mentions)
Stromatoxin, by Alomone (1 mentions)
Iberiotoxin, by Alomone (1 mentions)
Tetrodotoxin ttx, by Merck Group (1 mentions)
Tta p2, by Alomone (1 mentions)
Papain, by Worthington (1 mentions)
Dispase, by Roche (1 mentions)
Ionomycin, by Alomone (1 mentions)
Baclofen, by Merck Group (1 mentions)
C 300, by Alomone (1 mentions)
Sta 500, by Alomone (1 mentions)
Antimycotics, by Thermo Fisher Scientific (1 mentions)
Papain and dnase, by Worthington (1 mentions)
Calcium green 1 am, by Thermo Fisher Scientific (1 mentions)
ω conotoxin, by Alomone (1 mentions)
ω agatoxin, by Alomone (1 mentions)
Dmem f12, by Thermo Fisher Scientific (1 mentions)
Fibronectin, by Merck Group (1 mentions)
7 protocols using snx 482
1
Electrophysiological Experiments Protocols
2
Drug Application Techniques for Dissociated Cell Recordings
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For dissociated cell recordings, drugs were applied with a gravity-fed system that positioned a glass capillary tube 100 μm from the recording cell in the direction of superfusion flow. Solution changes were performed with a D.C. controlled microvalve system (Lee; Essex, CT, USA). This method allowed reversible drug applications [26 (link), 33 (link)]. For current clamp recordings drugs were administered into the bath saline. Substances used were the DA receptor D1-like selective agonist SKF 81297 (Cat# S143), DA receptor D1-like antagonist SCH 23390 (Cat# 125941-87-9), Ca2+ CaV1 antagonist nicardipine (Cat# N7510) all from Sigma-Aldrich-RBI (St Louis, MO, USA); Ca2+ CaV2.2 blocker ω-conotoxin GVIA (Cat# C-300), Ca2+ CaV3 blocker TTA-P2 (Cat# T-155), Ca2+ CaV2.3 blocker SNX-482 (Cat# RTS-500), Na+ blocker tetrodotoxin (TTX) (Cat# T-550) from Alomone Laboratories (Israel) and Ca2+ CaV2.1 blocker ω-agatoxin TK (Cat# 4294-s) from Peptides International (Louisville, KY).
3
DRG Sensory Neuron Culture Reagents
4
Preparation and Use of Ion Channel Modulators
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Stock solutions of nimodipine (20 mM in DMSO), TTX (0.3 mM in acetic acid), and carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP; 1 mM in DMSO) were stored at –20 °C, and were diluted at least 1000 times to reach desired final concentrations. These chemicals were purchased from Tocris Cookson (Ellisville, MO, USA). The CaV2 channel blockers SNX-482, ω-agatoxin IVA, and ω-conotoxin GVIA were purchased from Alomone Labs (Jerusalem, Israel) and were added directly to the bath solution. K+-free solution was prepared without extracellular K+, Ca2+-free solution was prepared with omission of extracellular Ca2+ and the addition of 1 mM EGTA, and high (20, 35, and 50 mM) K+ solutions were prepared with equal molar substitution of K+ for Na+.
5
Calcium Channel Modulation in Neurons
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Ionomycin, ω-conotoxin GVIA, ω-agatoxin IVA, and SNX-482 were purchased from Alomone labs (catalog numbers I-700, C-300, STA-500, and RTS-500 respectively). The toxins were applied for 3 min in Tyrode’s solution at concentrations of 1 µM, 400 nM, and 500 nM for ω-conotoxin GVIA, ω-agatoxin IVA, and SNX-482, respectively (Ariel et al., 2012 (link); Ermolyuk et al., 2013 (link); Hoppa et al., 2012 (link)). Baclofen was purchased from Sigma-Aldrich (catalog number B 5399) and continuously perfused at 10 µM beginning 5 min before AP stimulation (Laviv et al., 2011 (link)).
6
Isolation and Culture of Neuronal Cells
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DMEM/F-12, antibiotics and antimycotics were obtained from Gibco Laboratories (Grand Island, NY, USA), papain and DNase from Worthington Biochemical Corporation (Lakewood, NJ, USA) and FBS from Lonza (Basel, Switzerland). Calcium Green-1 AM was purchased from Molecular Probes (Eugene, OR, USA), tetrodotoxin from EMD Chemicals (San Diego, CA, USA), and ω-conotoxin, ω-agatoxin and SNX-482 from Alomone Labs (Jerusalem, Israel). DMPP, poly-d-lysine, fibronectin, collagen and nifedipine were obtained from Millipore Sigma (St. Louis, MO, USA). All other chemicals were reagent grade and purchased from standard commercial sources.
7
SDS-PAGE Protein Analysis Reagents
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Sodium dodecyl sulfate (SDS), bovine serum albumin (BSA) and phenylmethane sulfonyl fluoride (PMSF) were purchased from Amresco (Solon, OH, USA). Phosphate-buffered saline (PBS), RIPA Lysis Buffer, SDS-PAGE kit and SDS loading buffer were obtained from Beyotime (Shanghai, China). Oligomycin A was purchased from Selleckchem (Houston, TX, USA). SNX482 was obtained from Alomone Labs (Jerusalem, Israel). Agarose, ethylenediaminetetraacetic acid (EDTA), methanol, paraformaldehyde, Tween 20, thenoyltrifluoroacetone (TTFA), polyformaldehyde, phosphatase inhibitor cocktail 3, dimethyl sulfoxide (DMSO), 4′,6-diamidino-2-phenylindole (DAPI) and N-formyl-Met-Leu-Phe (fMLP) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Fluo-4/AM, Ca2+-free HBSS, Ca2+-containing HBSS, digitonin and Coomassie blue G-250 were obtained from Invitrogen (Carlsbad, CA, USA). Unless otherwise stated, all concentrations shown are the final concentrations.
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