Figure 2

ER degradation limits Rho-mOREG functional expression. CNG cells transiently transfected with Rho-mOREG were assayed for increases in intracellular calcium in response to 10 uM eugenol following treatment with vehicle (control, 0.1% DMSO; A), 10 mM 3-MA (B) or 50 uM MG-132 (C) for 4 h. Images contain ~750–1000 confluent cells and represent responses 60 sec following odorant stimulation. Scale bar is 150 um. Dose-response curves were determined for eugenol (300 nM to 300 uM; D and F) or isoproterenol (100 pM to 100 nM; E and G) in cells treated with vehicle (control), 10 mM 3-MA (D and E) or 50 uM MG-132 (F and G) for 4 h. ~20–25% of total cells or ~40–50% of transfected cells (~50% transfection efficiency) responded to 100 uM eugenol under control conditions. Note that degradation inhibitors specifically increased Rho-mOREG functional responses and had no effect on β-AR function. The EC₅₀ value for eugenol was larger for control (17.1 +/- 3.0 uM) compared to 3-MA (10.8 +/- 2.6 uM) and MG-132 (8.1 +/- 2.6 uM) treatments, but these differences did not achieve statistical significance. The EC₅₀ values for isoproterenol for control (3.7 +/- 1.0 nM), 3-MA (4.2 +/- 2.5 nM), and MG-132 (2.6 +/- 0.9 nM) treatments were not significantly different.