Abstract
There is a need to identify mu opioid receptor (MOR) agonists that can preferentially activate certain signaling pathways over others in order to develop better analgesics with reduced side effects. The present study aimed to determine if single-particle tracking (SPT) of the MOR could provide a robust sensor for distinct signaling states without reliance on effector-based readouts. SPT of quantum dot labeled FLAG-tagged MORs in AtT20 cells was performed. Under basal conditions, mobile and immobile states of FLAG-MORs were found to coexist on the plasma membrane. After cells were treated acutely with the MOR agonist DAMGO, there was a higher fraction of mobile trajectories and free portions of the mobile tracks resulted, in part, from G-protein interaction. After 10 minutes of agonist exposure, the population of immobile receptors increased, as did co-localization with clathrin. Yet, no single mobility state could fully account for colocalization with clathrin. Prior discrepant reports of MOR mobility may be attributable to the limitations of population assessments and use of a single exposure duration, which the current approach overcomes. Despite the advantages, it appears that the SPT approach will be most informative when effectors are examined simultaneously with the receptors.