μ-Opioid receptor regulates CFTR coexpressed in Xenopus oocytes in a cAMP independent manner

Diane R. Wotta, Angela K Birnbaum, George L Wilcox, Robert Eide, Ping-Yee Law

Research output: Contribution to journalArticlepeer-review

8 Scopus citations

Abstract

The objective of this study was to characterize the signaling mechanisms of the μ-opioid receptor in its coupling to the cystic fibrosis transmembrane conductance regulator (CFTR) when coexpressed in Xenopus oocytes. Because oocytes do not contain endogenous cAMP-regulated ion channels, the cAMP-modulated CFTR was coexpressed with receptors as a 'reporter' channel. Agonist treatment of oocytes coexpressing μ-opioid receptors, β2-adrenergic receptors and CFTR produced Cl- currents in a dose-related manner and immunocytochemical analysis confirmed receptor expression. These data suggest that opioid agonists could activate adenylyl cyclase in this system to elevate cAMP levels. Heterotrimeric G protein βγ-subunits acting on adenylyl cyclase type II would increase cAMP levels. The probable presence of adenylyl cyclase type II and other components of opioid signal transduction such as G(iα2), were demonstrated by RT-PCR. However, measurement of cAMP levels in individual oocytes by radioimmunoassay showed that opioid agonist application to oocytes expressing μ-opioid receptors, β2-adrenergic receptors and CFTR did not increase cAMP levels, whereas application of the β2-adrenergic agonist, isoproterenol, or IBMX alone did increase cAMP levels. Opioid-induced CFTR activation was not affected by either application of the broad spectrum kinase inhibitor, H7, nor by application of the specific PKA inhibitor, KT5720, Injection of free βγ-subunit, which could activate the endogenous type II cyclase, was unable to produce measurable currents in oocytes expressing the CFTR. These studies indicate that opioid activation of the CFTR is not mediated through a cAMP/PKA pathway, by either βγ-subunit activation of an adenylyl cyclase type II or promiscuous coupling to G(sα).

Original languageEnglish (US)
Pages (from-to)55-65
Number of pages11
JournalMolecular Brain Research
Volume44
Issue number1
DOIs
StatePublished - Feb 1 1997

Keywords

  • Adenylyl cyclase
  • Cyclic AMP
  • Cysistic fibrosis
  • Cystic fibrosis transmembrane conductance regulator
  • G protein
  • Oocyte
  • Opioid
  • Xenopus

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