Identification of dynorphins as endogenous ligands for an opioid receptor-like orphan receptor.
To identify the endogenous ligands for a cloned orphan receptor that shares high degrees of sequence homology with opioid receptors, this orphan receptor was expressed in Xenopus oocytes and in mammalian cell lines CHO-K1 and HEK-293. The coupling of the receptor to a G protein-activated K+ channel was used as a functional assay in oocytes. Endogenous opioid peptide dynorphins were found to activate the K+ channel by stimulating the orphan receptor. This activation was dose-dependent, with EC50 values at 45 and 37 nM for dynorphin A and dynorphin A-(1-13), respectively. The dynorphin effect was antagonized by the non-selective opioid antagonist naloxone but at rather high concentrations in the micromolar range. Naloxone also caused a rightward shift of the dose-response curve for dynorphin A, suggesting a competitive antagonism mechanism. In transiently transfected cells, 5 microM dynorphin A-(1-13) inhibited the forskolin-stimulated cyclic AMP increase by 51 and 35% in CHO-K1 and HEK-293 cells, respectively. Other classes of endogenous opioids, i.e. enkephalins and endorphins, caused very little activation of this receptor. These results suggest that this orphan receptor is a member of the opioid receptor family and that dynorphins are endogenous ligands for this receptor.
Zhang
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Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis 46202, USA.
Yu
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D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D003001:Cloning, Molecular; D005576:Colforsin / Q000494:pharmacology; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism; D004399:Dynorphins / Q000378:metabolism* / Q000494:pharmacology*; D005071:Evoked Potentials / Q000187:drug effects*; D005260:Female; D006801:Humans; D007700:Kinetics; D008024:Ligands; D009270:Naloxone / Q000494:pharmacology; D009292:Narcotic Antagonists / Q000494:pharmacology; D009294:Narcotics / Q000494:pharmacology*; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D011957:Receptors, Opioid / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D014982:Xenopus laevis
D008024:Ligands; D009292:Narcotic Antagonists; D009294:Narcotics; D011957:Receptors, Opioid; D011994:Recombinant Proteins; D005576:Colforsin; D009270:Naloxone; D004399:Dynorphins; D000242:Cyclic AMP