Fig. 6. The effect of EP₃ receptor stimulation on the localization of AQP2 in IMCD cells in the presence of high levels of cAMP. IMCD cells were left untreated (control) or incubated with AVP (100 nM; 15 minutes), Bt₂cAMP (500 µM; 15 minutes), forskolin (100 µM; 15 minutes), a strong, direct activator of adenylyl cyclase, and with or without sulprostone (1 µM; 30 minutes). If cells were incubated with both sulprostone and SC19220, SC19220 (10 µM) was added 10 minutes prior to sulprostone. AVP, Bt₂cAMP or forskolin were present during the final 15 minutes. After completion of the incubations, cells were fixed, permeabilized, and incubated with rabbit anti-AQP2 antibody and secondary Cy3-conjugated anti-rabbit antibodies. AQP2 immunofluorescence was detected by laser scanning microscopy. Scale bars: 20 µm.