Fig. 6: Active-state selective agonists have different efficacy for ³⁵S-GTPγS binding and release.

MOR receptor stimulation with different agonists in both response paradigms. In the release reaction, membranes are incubated without NaCl for ³⁵S-GTPγS loading and the release reaction is performed in the presence of 100 mM NaCl. Stephenson’s Efficacy (ε), Two-state Efficacy (α), and Operational Efficacy (τ) are presented for each fit of the data. Where efficacy could not be estimates, maximal bars are presented as without error. Although efficacy cannot be measured directly for full agonists, loss of efficacy can be estimated. All parameter estimates not adequately determined by nonlinear regression are presented as the undefined variable (θ) on the Y-axis. For Stephen’s efficacy, an unpaired two-tailed t-test was used (*p < 0.05, **p < 0.01); for two-state and operation efficacy, an extra sum-of-squares F-test was applied to determine changes in efficacy (*p < 0.05, ****p < 0.0001). DAMGO curves are the average multiple experiments (binding n = 5; release n = 6). All other agonists are the average of three independent experiments run with DAMGO control. Data are plotted as the mean with s.e.m.