Fig. 7.

Receptor recycling prevents S-phase induction triggered by brief treatment with forskolin. A, B, Timelines are shown. A, Pieces of epithelium were preincubated for 75 min (30/15/30 min) with 0.5 μm monensin (Mon) or 0.1 μmbafilomycin A1 (Baf). During the first 30 min, the medium did not contain BrdU. For the next 45 min, the medium contained FBS and BrdU. The pieces were then maintained in an FBS-containing medium with BrdU for the remainder of the 72 hr culture.B, Other pieces were pretreated with 0.5 μm monensin (Mon) or 0.1 μmbafilomycin A1 (Baf) for 30 min in a medium devoid of BrdU. The medium was then changed for a standard medium containing 1 μm forskolin (Forsk) for 15 min and replaced by a medium containing 0.5 μm monensin or 0.1 μm bafilomycin A1 and FBS and BrdU for 30 additional minutes. Finally, the monensin or the bafilomycin A1 was removed, and the pieces of utricle were cultured in a standard medium for the remainder of the 72 hr culture period. C, The 75 min (30/15/30 min) treatment with 0.5 μmmonensin or 0.1 μm bafilomycin A1 reduced the level of S-phase entry as compared with the control, but this was not significant. Treatment with monensin inhibited the 15 min forskolin effect up to 99% (Forsk 15 min + Mon), whereas treatment with bafilomycin A1 induced a 70.5% inhibition (Forsk 15 min + Baf). * Significant difference when compared with Forsk 15 min (p < 0.001). F, Fixed; S, start.