Specific Localization of b-Arrestin2 in Myenteric Plexus of Mouse Gastrointestinal Tract

b-arrestin2 is a key molecule involved in signaling and internalization of activated G protein-coupled receptors including m-opioid receptors (MOR). Previously we have shown that decreased expression of b-arrestin2 upon chronic morphine is associated with the development of opioid tolerance in the gastrointestinal tract. However, the localization of b-arrestin2 within the gastrointestinal wall is not known. In this study we found that b-arrestin2 is localized in the soma of a select group of neurons in the myenteric ganglia but not in smooth muscle. The density of b-arestin2 was significantly higher in the ileum than the colon. We identified four variants of b-arrestin2 in the ileum, with ARRB-005 and ARRB-013 being the most abundant. Further, the current study utilized multiple-labeling immunofluorescence to characterize the chemical coding of neurons expressing b-arrestin2 in the murine myenteric plexus and the co-localization of MOR1 and b-arrestin2. b-arrestin2 co-localized with choline acetyltransferase and calretinin. In contrast, b-arrestin2 neither co-localized with substance P, nitric oxide synthase nor calbindin. Genetic deletion of b-arrestin2 did not affect cholinergic neuron activation by nicotine in the isolated ileum (-log M EC50: wild type = 5.8 vs. b-arrestin2 knockout = 5.9). Our findings suggest specificity in the localization of b-arrestin2 in the myenteric plexus within MOR1-expressing neurons and provide a relation for direct intracellular crosstalk between MOR1 receptor activation and b-arrestin2 signaling in the myenteric neurons. b-arrestin