Relative potencies of various opioid agonists to inhibit forskolin-stimulated production of intracellular cAMP in HEK293 cells expressing the wild-type and third intracellular loop mutant μ-opioid receptors
| The ability of various concentrations of DAMGO, morphine, or PL017 to inhibit 10 μM forskolin-stimulated intracellular cAMP production in HEK293 cells stably expressing the wild-type and the third intracellular loop deletion mutant μ-opioid receptor was determined as described under Materials and Methods. Ki represents the concentration of agonist that produced 50% of the maximal inhibition. Data represent mean ± S.E.M. of two (when the Ki values were similar to WT) to eight (when Ki values were different from WT) separate dose response experiments in triplicates. | ||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Receptor | DAMGO | Morphine | PL017 | |||||||||||||||
| KI | Max. Inh. | KI/KH | KI | Max. Inh. | KI/KH | KI | Max. Inh. | KI/KH | ||||||||||
| nM | % | nM | % | nM | % | |||||||||||||
| WT | 8.3 ± 2.6 | 87 ± 2 | 5.9 | 13 ± 2.8 | 85 ± 2 | 5.3 | 11 ± 2.1 | 85 ± 2 | 1.0 | |||||||||
| R276A | 29 ± 6.6* | 73 ± 2 | 20 | 22 ± 7.5 | 88 ± 3 | 12 | 10 ± 3.5 | 87 ± 3 | 1.5 | |||||||||
| R277A | 4.7 ± 1.2 | 60 ± 1.2* | 1.2 | 27 ± 5.4* | 68 ± 5* | 2.4 | 39 ± 5.4* | 76 ± 5 | 7.2 | |||||||||
| I278A | 21 ± 7.1 | 70 ± 3 | 80 | 19 ± 9.3 | 87 ± 3 | 3.6 | 4.5 ± 1.4* | 89 ± 5 | 0.8 | |||||||||
| T279A | 17 ± 9.0 | 73 ± 3* | 28 | 6.0 ± 0.6* | 85 ± 2 | 2.2 | 2.3 ± 0.5* | 83 ± 3 | 0.26 | |||||||||
| R280A | 4.8 ± 2.3 | 77 ± 9 | 5.4 | 11 ± 5.5 | 87 ± 2 | 2.4 | 3.6 ± 1.3* | 79 ± 7 | 0.38 | |||||||||
| R276A/R277A | 58 ± 25* | 72 ± 9 | 55 | 47 ± 8.5* | 78 ± 5 | 6.7 | 30 ± 6.7* | 67 ± 8 | 6.2 | |||||||||
↵* Signifies statistical significant differences (p < 0.05) as determined by comparing the values of the mutant receptors to that of wild-type receptor using student unpaired t test.