Abstract
The humanmuopioid receptor was expressed stably in Flp-In
T-REx HEK293 cells. Occupancy by the agonist DAMGO (Tyr-
D-Ala-Gly-N-methyl-Phe-Gly-ol) resulted in phosphorylation
of the ERK1/2 MAP kinases, which was blocked by the opioid
antagonist naloxone but not the cannabinoid CB1 receptor
inverse agonist SR141716A. Expression of the human cannabinoidCB1receptor
in these cells from the inducible Flp-In T-REx
locus did not alter expression levels of the mu opioid receptor.
This allowed the cannabinoidCB1agonist WIN55212-2 to stimulate
ERK1/2 phosphorylation but resulted in a large reduction
in the capacity of DAMGO to activate these kinases. Although
lacking affinity for the mu opioid receptor, co-addition of
SR141716A caused recovery of the effectiveness of DAMGO. In
contrast co-addition of the CB1 receptor neutral antagonist
O-2050 did not. Induction of the CB1 receptor also resulted in
an increase of basal [35S]guanosine 5 -3-O-(thio)triphosphate
(GTP S) binding and thereby a greatly reduced capacity of
DAMGO to further stimulate [35S]GTP S binding. CB1 inverse
agonists attenuated basal [35S]GTP S binding and restored the
capacity of DAMGO to stimulate. Flp-In T-REx HEK293 cells
were generated, which express the human mu opioid receptor
constitutively and harbor a modified D163N cannabinoid CB1
receptor that lacks constitutive activity. Induction of expression
of the modified cannabinoid CB1 receptor did not limit
DAMGO-mediated ERK1/2 MAP kinase phosphorylation and
did not allow SR141716A to enhance the function of DAMGO.
These data indicate that it is the constitutive activity inherent in
the cannabinoid CB1 receptor that reduces the capacity of coexpressed
mu opioid receptor to function.
Original language | English |
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Pages (from-to) | 11424 - 11434 |
Number of pages | 11 |
Journal | Journal of Biological Chemistry |
Volume | 283 |
Issue number | 17 |
DOIs | |
Publication status | Published - 2008 |