THEMED ISSUE: GPCR RESEARCH PAPER Involvement of PKCa and G-protein-coupled receptor kinase 2 in agonist-selective desensitization of m-opioid receptors in mature brain neurons

Background and purpose: The ability of an agonist to induce desensitization of the m-opioid receptor (MOR) depends upon the agonist used. Furthermore, previous data suggest that the intracellular mechanisms underlying desensitization may be agonist-specific. We investigated the mechanisms underlying MOR desensitization, in adult mammalian neurons, caused by morphine (a partial agonist in this system) and DAMGO (a high-efficacy agonist). Experimental approach: MOR function was measured in locus coeruleus neurons, by using whole-cell patch-clamp electro- physiology, in rat and mouse brain slices (both wild-type and protein kinase C (PKC)a knockout mice). Specific isoforms of PKC were inhibited by using inhibitors of the receptors for activated C-kinase (RACK), and in vivo viral-mediated gene-transfer was used to transfect neurons with dominant negative mutants (DNMs) of specific G-protein-coupled receptor kinases (GRKs). Key results: Morphine-induced desensitization was attenuated by using RACK inhibitors that inhibit PKCa, but not by other isoform-specific inhibitors. Further, the PKC component of morphine-induced desensitization was absent in locus coeruleus neurons from PKCa knockout mice. The PKC-enhanced morphine-induced desensitization was not affected by over-expression of a GRK2 dominant negative mutant (GRK2 DNM). In contrast, DAMGO-induced MOR desensitization was independent of PKC activity but was reduced by over-expression of the GRK2 DNM but not by that of a GRK6 DNM. Conclusions and implications: In mature mammalian neurons, different MOR agonists can induce MOR desensitization by different mechanisms, morphine by a PKCa-mediated, heterologous mechanism and DAMGO by a GRK-mediated, homolo- gous mechanism. These data represent functional selectivity at the level of receptor desensitization.