Internalization and recycling of human mu opioid receptors expressed in Sf9 insect cells

Life Sci. 2003 May 23;73(1):115-28. doi: 10.1016/s0024-3205(03)00250-9.


Internalization and recycling of G protein-coupled receptors (GPCRs), such as the mu-opioid receptor, largely depend on agonist stimulation. Agonist-promoted internalization of some GPCRs has been shown to mediate receptor desensitization, resensitization, and down-regulation. In this study, we investigated whether different mu opioid agonists displayed different effects in receptor internalization and recycling, the potential mechanisms involved in ohmefentanyl-induced internalization process. In transfected Sf9 insect cells expressing 6His-tagged wild type mu opioid receptor, exposure to 100 nM ohmefentanyl caused a maximum internalization of the receptor at 30 min and receptors seemed to reappear at the cell membrane after 60 min as determined by radioligand binding assay. Ohmefentanyl-induced human mu opioid receptor internalization was concentration-dependent, with about 40% of the receptors internalized following a 30-min exposure to 1 microM ohmefentanyl. 10 microM morphine and 1 microM DAMGO could also induce about 40% internalization. The antagonist naloxone and pretreatment with pertussis toxin both blocked ohmefentanyl-induced internalization without affecting internalization themselves. Incubation with sucrose 0.45 M significantly inhibited ohmefentanyl-induced internalization of the mu receptor. The removal of agonists ohmefentanyl and morphine resulted in the receptors gradually returning to the cell surface over a 60 min period, while the removal of agonist DAMGO only partly resulted in the receptor recycling. The results of this study suggest that ohmefentanyl-induced internalization of human mu opioid receptor in Sf9 insect cells occurs via Gi/o protein-dependent process that likely involves clathrin-coated pits. In addition, the recycling process displays the differential modes of action of different agonists.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Analgesics, Opioid / pharmacology
  • Animals
  • Cell Line
  • Cyclic AMP / metabolism
  • Dose-Response Relationship, Drug
  • Down-Regulation / drug effects
  • Enkephalin, Ala(2)-MePhe(4)-Gly(5)- / pharmacology
  • Fentanyl / pharmacology
  • Humans
  • Hypertonic Solutions
  • Insecta / metabolism*
  • Microscopy, Confocal
  • Morphine / pharmacology
  • Naloxone / pharmacology
  • Narcotic Antagonists / pharmacology
  • Pertussis Toxin / pharmacology
  • Receptors, Cell Surface / drug effects*
  • Receptors, Opioid, mu / agonists
  • Receptors, Opioid, mu / metabolism*
  • Transfection


  • Analgesics, Opioid
  • Hypertonic Solutions
  • Narcotic Antagonists
  • Receptors, Cell Surface
  • Receptors, Opioid, mu
  • Enkephalin, Ala(2)-MePhe(4)-Gly(5)-
  • Naloxone
  • Morphine
  • Cyclic AMP
  • Pertussis Toxin
  • Fentanyl