Ability of delta-opioid receptors to interact with multiple G-proteins is independent of receptor density.
To determine whether the previously demonstrated ability of delta-opioid receptors to interact simultaneously with multiple G-proteins was a function of high receptor levels, this interaction was investigated in Chinese hamster ovary cells stably expressing 10 different levels of cloned delta-opioid receptors, ranging from 18,000 to 1.6 x 10(6) receptors/cell. The opioid agonist D-Ala2,D-Leu5-enkephalin (DADLE) inhibited forskolin-stimulated adenylyl cyclase activity in all 10 clones with variable maximal inhibitory levels. Furthermore, opioid agonists altered incorporation of [alpha-32P]azidoanilido-GTP into at least four G-protein alpha-subunits in all 10 clones, three of which were determined to be Gi3 alpha, Gi2 alpha and Go2 alpha. This effect was concentration-dependent, naloxone-reversible, and delta-opioid agonist-specific and was blocked by pretreatment with pertussis toxin. Although DADLE induced an increase in the incorporation of [alpha-32P]azidoanilido-GTP into three of the four G alpha proteins that was independent of receptor density, the magnitude of this response was greater as receptor density increased. In addition, concentrations of DADLE required to promote 50% maximal labeling were similar for all four G alpha proteins within each clone and did not appear to be affected by receptor density. Therefore, the ability of delta-opioid receptors to interact with multiple G-proteins is independent of receptor density and there is also no apparent correlation between the amount of G-protein activated and the maximal effect of an agonist.
Prather
P L
PL
Department of Pharmacology, University of Minnesota, School of Medicine, Minneapolis 55455.
McGinn
T M
TM
Erickson
L J
LJ
Evans
C J
CJ
Loh
H H
HH
Law
P Y
PY
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D016308:Enkephalin, Leucine-2-Alanine / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009270:Naloxone / Q000494:pharmacology; D017465:Receptors, Opioid, delta / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*
D017465:Receptors, Opioid, delta; D009270:Naloxone; D016308:Enkephalin, Leucine-2-Alanine; D019204:GTP-Binding Proteins