MOP and NOP receptor interaction: Studies with a dual expression system and bivalent peptide ligands.

M F Bird,J Mcdonald,C L Gibson,G Caló,B Horley,R Guerrini,D G Lambert,B Fraser,J P O’Doherty,John M Streicher

doi:10.1371/journal.pone.0260880

M F Bird, J Mcdonald + Show 8 more

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https://doi.org/10.1371/journal.pone.0260880

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Abstract

Opioids targeting mu;μ (MOP) receptors produce analgesia in the peri-operative period and palliative care. They also produce side effects including respiratory depression, tolerance/dependence and addiction. The N/OFQ opioid receptor (NOP) also produces analgesia but is devoid of the major MOP side effects. Evidence exists for MOP-NOP interaction and mixed MOP-NOP ligands produce analgesia with reduced side effects. We have generated a HEKMOP/NOP human expression system and used bivalent MOP-NOP and fluorescent ligands to (i) probe for receptor interaction and (ii) consequences of that interaction. We used HEKMOP/NOP cells and two bivalent ligands; Dermorphin-N/OFQ (MOP agonist-NOP agonist; DeNO) and Dermorphin-UFP101 (MOP agonist-NOP antagonist; De101). We have determined receptor binding profiles, GTPγ[35S] binding, cAMP formation and ERK1/2 activation. We have also probed MOP and NOP receptor interactions in HEK cells and hippocampal neurones using the novel MOP fluorescent ligand, DermorphinATTO488 and the NOP fluorescent ligand N/OFQATTO594. In HEKMOP/NOP MOP ligands displaced NOP binding and NOP ligands displaced MOP binding. Using fluorescent probes in HEKMOP/NOP cells we demonstrated MOP-NOP probe overlap and a FRET signal indicating co-localisation. MOP-NOP were also co-localised in hippocampal tissue. In GTPγ[35S] and cAMP assays NOP stimulation shifted the response to MOP rightwards. At ERK1/2 the response to bivalent ligands generally peaked later. We provide evidence for MOP-NOP interaction in recombinant and native tissue. NOP activation reduces responsiveness of MOP activation; this was shown with conventional and bivalent ligands.

Highlights

Opioids targeting the mu opioid peptide (MOP) receptor are amongst the most widely used analgesics in both acute and chronic pain treatment
There is growing evidence to suggest that the potential pairing of the MOP and nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor systems may be beneficial in developing analgesics with reduced side effects [14,15,16,17]
Using our novel fluorescent ligands [14,39], we aimed to determine whether MOP and N/OFQ opioid receptor (NOP) receptors were expressed in close proximity to each other in our co-expression system

Summary

Introduction

Opioids targeting the mu opioid peptide (MOP) receptor are amongst the most widely used analgesics in both acute and chronic pain treatment Exclusively targeting this receptor leads to addiction and/or tolerance. Animal studies demonstrated that co-targeting an additional member of the opioid receptor family with either agonists or antagonists has been shown to be beneficial in longer-term antinociceptive paradigms, while reducing tolerance [1]. This synergistic effect is believed to be derived from the ability of opioid receptors to interact at a molecular and structural level [2]. There is growing evidence to suggest that the potential pairing of the MOP and nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor systems may be beneficial in developing analgesics with reduced side effects [14,15,16,17]

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