MOLECULAR MECHANISMS OF AGONIST-INDUCED DESENSITIZATION OF THE CLONEDMOUSE KAPPA-OPIOID RECEPTOR

Prolonged exposure of opioid receptors to agonists can cause desensiti zation, a cellular event linked to tolerance. Although evidence exists for mu and delta opioid receptor desensitization, much less informati on is available concerning the in vitro regulation of kappa opioid rec eptors because no cell lines exist that specifically express this clas s of opioid receptor. Recently we have cloned the mouse kappa opioid r eceptor. After expression in COS-7 cells, this protein exhibits the ph armacological specificity of a kappa(1) receptor and mediates agonist inhibition of cAMP formation. Continuous exposure of COS-7 cells expre ssing the kappa receptor to the agonist trans-(+/-)-3, 4-dichloro-N-me thyl-N-[2-(1 -pyrrolidinyl)-cyclohexyl]-benzeneacetamide methanesulfon ate salt (U50,488) reduces the specific binding of the kappa-selective agonist [H-3]U69,593. Furthermore, the potency of U50,488 to inhibit the binding of the opiate antagonist [H-3] naloxone to the kappa recep tor is reduced. However, total specific binding of [H-3]naloxone is no t altered, indicating that short-term (2-4 hr) agonist treatment of th e kappa receptor reduces the affinity of the receptor for agonists but does not reduce the density of kappa receptors. The reduction in affi nity of the kappa receptor for agonists is dependent on the time of ag onist exposure and is reversible. The reduced affinity of the receptor for agonists is associated with kappa receptor desensitization, becau se kappa receptor-mediated inhibition of cAMP formation is lost in cel ls pretreated with U50,488. The desensitization of the kappa receptor is dependent on the time and concentration of agonist treatment, is bl ocked by the kappa-selective antagonist nor-binaltorphimine and is rev ersible. The desensitization is also induced by kappa agonists of diff ering structures such as dynorphin and bremazocine and is stereoselect ive. The enzyme beta adrenergic receptor kinase (BARK) appears to be i nvolved in kappa receptor desensitization since in COS-7 cells cotrans lected with the kappa receptor cDNA and a cDNA encoding a dominant neg ative mutant of BARK, agonist pretreatment does not desensitize the ka ppa receptor. These findings suggest that kappa receptor desensitizati on is likely to occur via a BARK-mediated mechanism.