Characterization of the human oxytocin receptor stably expressed in 293 human embryonic kidney cells.
The human oxytocin (OT) receptor was stably expressed in 293 embryonic kidney cells (293/OTR), characterized pharmacologically and compared to human uterine myometrial receptors. The cloned receptor is expressed at a reasonably high density (0.82 fmole/microgram protein) and exhibits high affinity for [3H]OT (Kd = 0.32nM), similar to the value found in human myometrial tissue. The rank-order of potency for various antagonist and agonist ligands from several structural classes is also similar between the cloned and native receptor, as seen in a comparison of their inhibitory constants for [3H]OT binding. Agonist affinity at the cloned OT receptor is decreased by guanine nucleotide analogs, demonstrating functional G-protein-coupling. The OT receptor in 293 cells, like in human myometrium, is also coupled to the inositol phosphate pathway. In 293/OTR cells, OT stimulates inositol phosphate accumulation with an EC50 = 4.1 nM, an effect blocked by a potent and selective OT antagonist, L-366,948. Additionally, the cloned receptor in 293 cells desensitizes to high concentrations of OT, similar to the desensitization in myometrial tissue and also described for several other G-protein-coupled receptors. These results illustrate the utility of the 293 cell line for expressing human OT receptors in an environment quite comparable to the native myometrial tissue.
D001127:Arginine Vasopressin / Q000378:metabolism; D002460:Cell Line; D004361:Drug Tolerance; D004622:Embryo, Mammalian; D005260:Female; D015870:Gene Expression; D006150:Guanine Nucleotides / Q000494:pharmacology; D006801:Humans; D006868:Hydrolysis; D007668:Kidney; D009215:Myometrium / Q000737:chemistry; D010121:Oxytocin / Q000378:metabolism / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000378:metabolism; D018045:Receptors, Oxytocin / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins; D014162:Transfection; D014316:Tritium
D006150:Guanine Nucleotides; D010716:Phosphatidylinositols; D018045:Receptors, Oxytocin; D011994:Recombinant Proteins; D014316:Tritium; D001127:Arginine Vasopressin; D010121:Oxytocin