Functional Reconstitution of Purified Gi and Go with μ‐Opioid Receptors in Guinea Pig Striatal Membranes Pretreated with Micromolar Concentrations of N‐Ethylmaleimide

Hiroshi Ueda, Hidemi Misawa, Toshiaki Katada, Michio Ui, Hiroshi Takagi, Masamichi Satoh

Research output: Contribution to journalArticle

54 Citations (Scopus)

Abstract

Abstract: Functional coupling between μ‐opioid receptors and GTP‐binding regulatory proteins (G proteins) was investigated in reconstituted membranes of the guinea pig striatum. Selective μ‐opioid agonists stimulated low‐Km GTPase in striatal membranes, in a Na+‐dependent manner. The same μ‐opioid agonist {[D‐Ala2,N‐Me‐Phe4,Gly5‐ol]‐enkephalin (DAGO)} caused no stimulation when the membranes were exposed to islet‐activating protein (IAP; pertussis toxin). There was also no DAGO stimulation in preparations pretreated with a lower concentration (5 μM) of N‐ethylmaleimide (NEM), which abolished the ADP‐ribosylation of purified Gi (the G protein that mediates inhibition of adenylate cyclase) and Go (a G protein of unknown function purified from bovine brain) by IAP. In addition, as the NEM treatment caused no change in the μ‐agonist binding, NEM could probably substitute for IAP in inactivating native G proteins, without exhibiting effects on the receptor binding in membranes. The μ‐agonist stimulation of low‐Km GTPase activity in NEM‐treated membranes was recovered by reconstitution with purified Gi, or Go. The μ‐agonist stimulation of low‐Km GTPase was additive when Gi and Go were simultaneously reconstituted in NEM‐treated membranes in amounts of 0.5 pmol/assay, which was required for maximal recovery, in either reconstitution experiment. The present findings provide the first evidence that the μ‐opioid receptor may exist in at least two different forms, separately coupled to Gi or Go.

Original languageEnglish
Pages (from-to)841-848
Number of pages8
JournalJournal of Neurochemistry
Volume54
Issue number3
DOIs
Publication statusPublished - 1990 Mar
Externally publishedYes

    Fingerprint

Keywords

  • μ‐Opioid receptors–GTP‐binding regulatory proteins–Striatal membranes–[D‐Ala,N‐Me‐Phe,Gly‐ol]‐Enkephalin–Islet‐activating protein–Pertussis toxin–N‐Ethylmaleimide–G–G

ASJC Scopus subject areas

  • Biochemistry
  • Cellular and Molecular Neuroscience

Cite this