Endogenous opioids activate opioid receptors (ORs) in the enteric nervous system to control intestinal motility and secretion. The μ-OR mediates the deleterious side effects of opioid analgesics, including constipation, respiratory depression, and addiction. Although the δ-OR (DOR) is a promising target for analgesia, the function and regulation of DOR in the colon are poorly understood. This study provides evidence that endogenous opioids activate DOR in myenteric neurons that may regulate colonic motility. The DOR agonists DADLE, deltorphin II, and SNC80 inhibited electrically evoked contractions and induced neurogenic contractions in the mouse colon. Electrical, chemical, and mechanical stimulation of the colon evoked the release of endogenous opioids, which stimulated endocytosis of DOR in the soma and proximal neurites of myenteric neurons of transgenic mice expressing DOR fused to enhanced green fluorescent protein. In contrast, DOR was not internalized in nerve fibers within the circular muscle. Administration of dextran sulfate sodium induced acute colitis, which was accompanied by DOR endocytosis and an increased density of DOR-positive nerve fibers within the circular muscle. The potency with which SNC80 inhibited neurogenic contractions was significantly enhanced in the inflamed colon. This study demonstrates that DOR-expressing neurons in the mouse colon can be activated by exogenous and endogenous opioids. Activated DOR traffics to endosomes and inhibits neurogenic motility of the colon. DOR signaling is enhanced during intestinal inflammation. This study demonstrates functional expression of DOR by myenteric neurons and supports the therapeutic targeting of DOR in the enteric nervous system. NEW & NOTEWORTHY DOR is activated during physiologically relevant reflex stimulation. Agonist-evoked DOR endocytosis is spatially and temporally regulated. A significant proportion of DOR is internalized in myenteric neurons during inflammation. The relative proportion of all myenteric neurons that expressed DOR and the overlap with the nNOS-positive population are increased in inflammation. DOR-specific innervation of the circular muscle is increased in inflammation, and this is consistent with enhanced responsiveness to the DOR agonist SNC80.

G protein-coupled receptors physically and functionally interact, leading to unique signaling. We report that delta and mu opioid receptors are coexpressed and functionally interact in myenteric neurons. Our findings have implications for opioid-based therapies for motility disorders and pain. BACKGROUND & AIMS:Functional interactions between the mu opioid receptor (MOR) and delta opioid receptor (DOR) represent a potential target for novel analgesics and may drive the effects of the clinically approved drug eluxadoline for the treatment of diarrhea-predominant irritable bowel syndrome. Although the enteric nervous system (ENS) is a likely site of action, the coexpression and potential interaction between MOR and DOR in the ENS are largely undefined. In the present study, we have characterized the distribution of MOR in the mouse ENS and examined MOR-DOR interactions by using pharmacologic and cell biology techniques.METHODS: MOR and DOR expression was defined by using MORmCherry and MORmCherry-DOR-eGFP knockin mice. MOR-DOR interactions were assessed by using DOR-eGFP internalization assays and by pharmacologic analysis of neurogenic contractions of the colon. RESULTS:Although MOR was expressed by approximately half of all myenteric neurons, MOR-positive submucosal neurons were rarely observed. There was extensive overlap between MOR and DOR in both excitatory and inhibitory pathways involved in the coordination of intestinal motility. MOR and DOR can functionally interact, as shown through heterologous desensitization of MORdependent responses by DOR agonists. Functional evidence suggests that MOR and DOR may not exist as heteromers in the ENS. Pharmacologic studies show no evidence of cooperativity between MOR and DOR. DOR internalizes independently of MOR in myenteric neurons, and MOR-evoked contractions are unaffected by the sequestration of DOR. CONCLUSIONS:Collectively, these findings demonstrate that although MOR and DOR are coexpressed in the ENS and functionally interact, they are unlikely to exist as heteromers under physiological conditions. (Cell Mol Gastroenterol Hepatol 2020;9:465-483; https://doi.

Ivacaftor–lumacaftor and ivacaftor are two new breakthrough cystic fibrosis transmembrane conductance modulators.The interactions of ivacaftor and its two metabolites hydroxymethylivacaftor (iva-M1) and ivacaftorcarboxylate (iva-M6) with neurotransmitter receptors were investigated in radioligand binding assays.Ivacaftor displayed significant affinity to the 5-hydroxytryptamine (5-HT; serotonin) 5-HT2C receptor (pKi=6.06±0.03), β3-adrenergic receptor (pKi=5.71±0.07), δ-opioid receptor (pKi=5.59±0.06) and the dopamine transporter (pKi=5.50±0.20); iva-M1 displayed significant affinity to the 5-HT2C receptor (pKi=5.81±0.04) and the muscarinic M3 receptor (pKi=5.70±0.10); iva-M6 displayed significant affinity to the 5-HT2A receptor (pKi=7.33±0.05). The in vivo central nervous system activity of ivacaftor (40 mg·kg−1 intraperitoneally for 21 days) was assessed in a chronic mouse model of depression. In the forced swim test, the ivacaftor-treated group displayed decreased immobility (52.8±7.6 s), similarly to fluoxetine (33.8±11.0 s), and increased climbing/swimming activity (181.5±9.2 s). In the open field test, ivacaftor produced higher locomotor activity than the fluoxetine group, measured both as mean number of paw touches (ivacaftor 81.1±9.6 versus fluoxetine 57.9±9.5) and total distance travelled (ivacaftor 120.6±16.8 cm versus fluoxetine 84.5±16.0 cm) in 600 s. Treatment of 23 cystic fibrosis patients with ivacaftor–lumacaftor resulted in significant improvements in quality of life (including anxiety) in all five domains of the AweScoreCF questionnaire (p=0.092–0.096).Our findings suggest ivacaftor displays potential clinical anxiolytic and stimulating properties, and may have beneficial effects on mood.