Background: GPR30 plays important roles in cardiometabolic regulation and cancer. Results: GPR30 forms a complex with a MAGUK and AKAP5 that constitutively inhibits cAMP production independently of G i/o and retains receptors in the plasma membrane.
Conclusion:The GPR30-MAGUK-AKAP5 complex mediates receptor signaling. Significance: These results present a new mechanism by which a receptor inhibits cAMP production.
Activation of kappa opioid receptors (KORs) expressed by peripheral sensory neurons that respond to noxious stimuli (nociceptors) can reduce neurotransmission of pain stimuli from the periphery to the central nervous system. We have previously shown that the antinociception dose-response curve for peripherally restricted doses of the KOR agonist (-)-(trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]benzeneacetamide (U50488) has an inverted U shape. Here, we found that the downward phase of the U50488 dose-response curve was blocked by an inhibitor of extracellular signal-regulated kinase (ERK) activation U0126. Local administration of the selective KOR agonist salvinorin A (Sal-A), also resulted in an inverted U-shaped curve; however, the downward phase was insensitive to U0126. By contrast, inhibition of c-Jun N-terminal kinase (JNK) partially blocked the downward phase of the doseresponse curve to Sal-A, suggesting a role for JNK. In cultures of peripheral sensory neurons, U50488 and Sal-A inhibited adenylyl cyclase activity with similar efficacies; however, their ability to activate ERK and JNK differed. Whereas U50488 activated ERK but not JNK, Sal-A activated JNK but not ERK. Moreover, although both U50488 and Sal-A produced homologous desensitization, desensitization to U50488 was blocked by inhibition of ERK activation, whereas desensitization to Sal-A was blocked by inhibition of JNK. Substitution of an ethoxymethyl ether for the C2 position acetyl group of Sal-A reduced stimulation of JNK, prevented desensitization by ethoxymethyl ether for the C2 position acetyl group of Sal-A, and resulted in a monotonic antinociception dose-response curve. Collectively, these data demonstrate the functional selectivity of KOR ligands for signaling in peripheral sensory neurons, which results in differential effects on behavioral responses in vivo.
Opioid overdose is a leading cause of death in the United States. The only treatment available currently is the competitive antagonist, naloxone (Narcan®). Although naloxone is very effective and has saved many lives, as a competitive antagonist it has limitations. Due to the short half‐life of naloxone, renarcotization can occur if the ingested opioid agonist remains in the body longer. Moreover, because antagonism by naloxone is surmountable, renarcotization can also occur in the presence of naloxone if a relatively larger dose of opioid agonist is taken. In such circumstances, a long‐lasting, non‐surmountable antagonist would offer an improvement in overdose treatment. Methocinnamox (MCAM) has been reported to have a long duration of antagonist action at mu opioid receptors in vivo. In HEK cells expressing the human mu opioid receptor, MCAM antagonism of mu agonist‐inhibition of cAMP production was time‐dependent, non‐surmountable and non‐reversible, consistent with (pseudo)‐irreversible binding. In vivo, MCAM injected locally into the rat hindpaw antagonized mu agonist‐mediated inhibition of thermal allodynia for up to 96 h. By contrast, antagonism by MCAM of delta or kappa agonists in HEK cells and in vivo was consistent with simple competitive antagonism. Surprisingly, MCAM also shifted the concentration‐response curves of mu agonists in HEK cells in the absence of receptor reserve in a ligand‐dependent manner. The shift in the [D‐Ala2,N‐MePhe4,Gly‐ol5]‐enkephalin (DAMGO) concentration‐response curve by MCAM was insensitive to naloxone, suggesting that in addition to (pseudo)‐irreversible orthosteric antagonism, MCAM acts allosterically to alter the affinity and/or intrinsic efficacy of mu agonists.
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