ChemiScreen™ δ Opioid Receptor Calcium-Optimized FLIPR Cell Line

ChemiScreen™ δ Opioid Receptor Calcium-Optimized FLIPR Cell Line

• ChemiScreen
• Chemicon (Millipore)

2 vials

SPECIFICATIONS: EC50 for calcium mobilization by DPDPE: ~ 6.6 nM

HOST CELLS: Chem-1, an adherent cell line expressing the promiscuous G-protein, Gα15.

TRANSFECTION: Full-length human OPRD1 cDNA encoding δ opioid receptor (Accession Number: NM_000911)

GROWTH MEDIA: DMEM containing 4.5 g/L glucose/10% heat inactivated fetal bovine serum/1x nonessential amino acids/10 mM HEPES/0.25 mg/ml Geneticin (G418)/100 U/ml each penicillin and streptomycin

Opiates derived from the opium poppy, Papaver somniferum, have been used in for millenia for their anti-diarrheal, analgesic and euphoric properties. More recently, endogenous peptides, enkephalins, dynorphins, and endorphins, were found to bind to the same sites as opiate alkaloids. The receptors for the classical opioids are three related GPCRs, delta, kappa, and mu, also known as OP₁, OP₂ and OP₃, resepctively), that activate Gi/o to reduce intracellular cAMP levels. Although most clinically used opioids function by activation of the µ opioid receptor, agonits of spinal  opioid receptors have antinociceptive activity that is independent of µ. In addition, activation of  increases locomotor activity, inhibits gastrointestinal motility, and decreases respiratory frequency (Dhawan et al., 1996). Agonists for  opioid receptors also exhibit antidepressant-like activity in animal models (Broom et al., 2002). Chemicon's cloned human δ opioid - expressing cell line is made in the Chem-1 host, which supports high levels of recombinant δ opioid expression on the cell surface and contains high levels of the promiscuous G protein Gα15 to couple the receptor to the calcium signaling pathway. Thus, the cell line is an ideal tool for screening for agonists and antagonists of the δ opioid receptor

Calcium flux assay, ligand binding assays

Human

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FUNCTION: SwissProt: P41143 # Inhibits neurotransmitter release by reducing calcium ion currents and increasing potassium ion conductance. Highly stereoselective. receptor for enkephalins.
SIZE: 372 amino acids; 40413 Da
SUBUNIT: Interacts with GPRASP1.
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
SIMILARITY: SwissProt: P41143 ## Belongs to the G-protein coupled receptor 1 family.

Human

Chem-1

GPCR Cell Lines

Chemicon

Chem-1

Cells are frozen at 2 x 10⁶ cells/mL in DMEM/20% fetal bovine serum/100 U/ml penicillin and streptomycin/10% DMSO. Cell line tests negative for mycoplasma.

Delta / OP1 / DOP / DOR

P41143

peptide

Place cells in liquid nitrogen immediately upon receipt. Maintain frozen in liquid nitrogen for up to 5 years. Thaw cells rapidly by removing from liquid nitrogen and immediately immersing in a 37ºC water bath. Immediately after ice has thawed, sterilize the exterior of the vial with 70% ethanol. Transfer contents of the vial to a T75 flask containing 20 mL growth media, and place in a humidified 37°C incubator with 5% CO2. After 8-24 h, cells will adhere to the plate, at which time the media should be replaced to remove residual DMSO. Cells are passaged by washing with Ca++ and Mg++-free HBSS (10 mL/T75), incubating with 0.05% trypsin/0.2 g/L EDTA (1 mL/T75) for 5-10 minutes at 37°C, and rapping the side of the flask to dislodge the cells. Neutralize the trypsin by addition of 4 volumes growth media. Cells are typically passaged 1:10 with every 3-4 days, and should be passaged at least once after thawing prior to use in calcium flux assays.

Opioid

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A

2 x 10⁶ cells/vial

• OPRD1
• OPRD
• DOR-1

GPCRs

ChemiScreen™ δ Opioid Receptor Calcium-Optimized FLIPR Cell Line

NM_000911.3