PEN is an abundant peptide in the brain that has been implicated in the regulation of feeding. We identified a receptor for PEN in mouse hypothalamus and Neuro2A cells. PEN bound to and activated GPR83, a G protein (heterotrimeric guanine nucleotide–binding protein)–coupled receptor (GPCR). Reduction of GPR83 expression in mouse brain and Neuro2A cells reduced PEN binding and signaling, consistent with GPR83 functioning as the major receptor for PEN. In some brain regions, GPR83 colocalized with GPR171, a GPCR that binds the neuropeptide bigLEN, another neuropeptide that is involved in feeding and is generated from the same precursor protein as is PEN. Coexpression of these two receptors in cell lines altered the signaling properties of each receptor, suggesting a functional interaction. Our data established PEN as a neuropeptide that binds GPR83 and suggested that these two ligand-receptor systems—PEN-GPR83 and bigLEN-GPR171—may be functionally coupled in the regulation of feeding.

Highlights d Exposure to the psychedelic drug DOI results in enduring molecular adaptations d Post-acute DOI unveils phenotypes akin to antidepressant adaptations d Concurrent occurrence of synaptic plasticity mediated via 5-HT

Opioids, such as morphine and fentanyl, are widely used for the treatment of severe pain; however, prolonged treatment with these drugs leads to the development of tolerance and can lead to opioid use disorder. The “Opioid Epidemic” has generated a drive for a deeper understanding of the fundamental signaling mechanisms of opioid receptors. It is generally thought that the three types of opioid receptors (μ, δ, κ) are activated by endogenous peptides derived from three different precursors: Proopiomelanocortin, proenkephalin, and prodynorphin. Posttranslational processing of these precursors generates >20 peptides with opioid receptor activity, leading to a long-standing question of the significance of this repertoire of peptides. Here, we address some aspects of this question using a technical tour de force approach to systematically evaluate ligand binding and signaling properties ([35S]GTPγS binding and β-arrestin recruitment) of 22 peptides at each of the three opioid receptors. We show that nearly all tested peptides are able to activate the three opioid receptors, and many of them exhibit agonist-directed receptor signaling (functional selectivity). Our data also challenge the dogma that shorter forms of β-endorphin do not exhibit receptor activity; we show that they exhibit robust signaling in cultured cells and in an acute brain slice preparation. Collectively, this information lays the groundwork for improved understanding of the endogenous opioid system that will help in developing more effective treatments for pain and addiction.

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