A human codon-adapted PoXeR gene was synthesized by Gen Script (Piscataway, NJ, USA) and cloned into pEGFP vector between HindIII and BamHI sites. ND7/23 cells were purchased from DS Pharma Biomedical (Osaka, Japan) and cultured in high-glucose DMEM media (Wako) in a 37 °C, 5% CO2 incubator. Transfection of ND7/23 cells was performed by Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA). Cells were supplemented with 1 μM all-trans-retinal (Sigma-Aldrich) after transfection. Expression was confirmed by a fluorescence microscope (IX-73, Olympus, Tokyo, Japan).
Nd7 23 cells
Manufactured by Sumitomo Pharma
Sourced in Japan
The ND7/23 cells are a cell line derived from mouse dorsal root ganglia. They express voltage-gated sodium channels and are commonly used as a model system for studying neuronal function and excitability.
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4 protocols using nd7 23 cells
1
Heterologous Expression of PoXeR in ND7/23 Cells
2
Cloning and Expression of GtCCR4-eGFP
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The GtCCR4 gene was cloned into peGFP vector between HindIII and BamHI sites in such a way that the eGFP is tagged at the C-terminus of GtCCR4. ND7/23 cells were purchased from DS Pharma Biomedical (Osaka, Japan) and cultured in high-glucose DMEM media (Wako, Osaka, Japan) in a 37°C, 5% CO2 incubator. Transfection of ND7/23 cells was performed by Lipofectamine 2000 (Invitrogen, CA, USA). Cells were supplemented with 1 μM all-trans-retinal (Sigma-Aldrich) after transfection. Expression of GtCCR4 was confirmed by eGFP fluorescence for a whole-cell patch clamp recording.
3
Electrophysiological Assays of Sensory Receptors
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The electrophysiological assays of SzRs were performed using ND7/23 cells, that is, hybrid cell lines derived from neonatal rat dorsal root ganglia neurons fused with mouse neuroblastoma (42 (link)). ND7/23 cells were purchased from DS Pharma Biomedical Co. Ltd. (Osaka, Japan) and cultured in high-glucose Dulbecco’s modified Eagle’s medium (DMEM) (Wako, Osaka, Japan) supplemented with 5% fetal bovine serum at 37°C and incubated in 5% CO2. For the electrophysiological recordings, the expression plasmids were transiently transfected using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions. For cytochemistry, the expression plasmids were transiently transfected using Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions. Cells were supplemented with 1 μM all-trans retinal (Sigma-Aldrich, St. Louis, MO, USA) for the electrophysiological recordings or 2.5 μM all-trans retinal for the cytochemical assay after transfection. Electrophysiological recordings and cytochemical assay were then conducted 24 to 36 hours after the transfection. Successfully transfected cells were identified by eYFP fluorescence under a microscope.
4
Optogenetic Activation of GtCCR4 in ND7/23 Cells
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