title: Adenosine A(1) receptor agonist N(6)-cyclohexyl-adenosine induced phosphorylation of delta opioid receptor and desensitization of its signaling.
abstract: OBJECTIVE
  To define the effect of adenosine A(1) receptor (A(1)R) on delta opioid receptor (DOR)-mediated signal transduction.
  
  
  METHODS
  CHO cells stably expressing HA-tagged A(1)R and DOR-CFP fusion protein were used. The localization of receptors was observed using confocal microscope. DOR-mediated inhibition of adenylyl cyclase was measured using cyclic AMP assay. Western blots were employed to detect the phosphorylation of Akt and the DOR. The effect of A(1)R agonist N(6)-cyclohexyladenosine (CHA) on DOR down-regulation was assessed using radioligand binding assay.
  
  
  RESULTS
  CHA 1 micromol/L time-dependently attenuated DOR agonist [D-Pen(2,5)]enkephalin (DPDPE)-induced inhibition of intracellular cAMP accumulation with a t(1/2)=2.56 (2.09-3.31) h. Pretreatment with 1 micromol/L CHA for 24 h caused a right shift of the dose-response curve of DPDPE-mediated inhibition of cAMP accumulation, with a significant increase in EC(50) but no change in E(max). Pretreatment with 1 micromol/L CHA for 1 h also induced a significant attenuation of DPDPE-stimulated phosphorylation of Akt. Moreover, CHA time-dependently phosphorylated DOR (Ser363), and this effect was inhibited by A(1)R antagonist 1,3-Dipropyl-8-cyclopentylxanthine (DPCPX) but not by DOR antagonist naloxone. However, CHA failed to produce the down-regulation of DOR, as neither receptor affinity (K(d)) nor receptor density (B(max)) of DOR showed significant change after chronic CHA exposure.
  
  
  CONCLUSIONS
  Activation of A(1)R by its agonist caused heterologous desensitization of DOR-mediated inhibition of intracellular cAMP accumulation and phosphorylation of Akt. Activation of A(1)R by its agonist also induced heterologous phosphorylation but not down-regulation of DOR.
authors.affiliation: Shanghai Institute of Materia Medica, Chinese Academy of Sciences, China.
authors.forename: Yun
authors.initials: Y
authors.lastname: Cheng
authors.forename: Yi-min
authors.initials: YM
authors.lastname: Tao
authors.forename: Jian-feng
authors.initials: JF
authors.lastname: Sun
authors.forename: Yu-hua
authors.initials: YH
authors.lastname: Wang
authors.forename: Xue-jun
authors.initials: XJ
authors.lastname: Xu
authors.forename: Jie
authors.initials: J
authors.lastname: Chen
authors.forename: Zhi-qiang
authors.initials: ZQ
authors.lastname: Chi
authors.forename: Jing-gen
authors.initials: JG
authors.lastname: Liu
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D058907:Adenosine A1 Receptor Agonists; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D006207:Half-Life; D006801:Humans; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011869:Radioligand Assay; D017465:Receptors, Opioid, delta / Q000187:drug effects* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors
chemical_list: D058907:Adenosine A1 Receptor Agonists; D017465:Receptors, Opioid, delta; C027513:N(6)-cyclohexyladenosine; D000242:Cyclic AMP; D051057:Proto-Oncogene Proteins c-akt; D000241:Adenosine