β‐arrestin2 expression is localized in cholinergic but not nitrergic motor neurons in the mouse longitudinal musclemyenteric plexus (LMMP)

Abstract

Morphine inhibits intestinal peristalsis via its action on cholinergic enteric neurons. Tolerance to morphine does not develop in the colon, except upon deletion of β‐arrestin2 (β‐arr2), a key molecule involved in G protein–coupled receptor desensitization (Maguma et al., 2012). However, the specific localization of β‐arr2 in the myenteric plexus is not known. In this study we utilized dual‐labeling immunofluorescence to characterize myenteric neurons expressing β‐arr2 and investigated the impact of β‐arr2 abrogation on cholinergic function. Our results reveal robust expression of β‐arr2 in the soma of select neurons that stained positive for choline acetyltransferase and calretinin, a motor neuron marker. β‐arr2 was neither co‐localized with substance P, nitric oxide synthase nor calbindin. The density of β‐arr2 neurons was higher in the ileum (10.8%) vs. colon (4.3%). Deletion of β‐arr2 did not affect cholinergic neuron activation by nicotine in the isolated ileum (− log EC50: wild type = 5.8 vs β‐arr2 knockout = 5.9). Our findings suggest specificity in the localization of β‐arr2 within cholinergic motor neurons of the myenteric plexus. β‐arr2 deletion does not directly alter cholinergic neuronal activation. Studying β‐arr2 signaling in the cholinergic motor neurons would be critical to assess mechanisms involved in reversal of opioid effect in the colon. Support: NIH grant DA024009