The binding and subsequent fate of 125I-labeled bombesin was studied in rat pancreatic acini. At both 37 degrees C and 4 degrees C binding occurred to a single saturable site, with Kd equal to 1.7 nM. The amount of steady-state tracer binding was reduced at 4 degrees C (5.7%/mg protein) compared with 37 degrees C (8.7%/mg), with a similar relative change in calculated binding capacity. With the use of an acid wash procedure to remove surface bound ligand, 55% of cell-associated 125I-bombesin was internalized in the steady state at 37 degrees C but only 5% at 4 degrees C. Preincubation at 4 degrees C followed by an increase to 37 degrees C led to rapid internalization of bound bombesin, which was blocked by the metabolic inhibitor antimycin. 125I-bombesin was found to be degraded by two acinar systems. One was not related to receptor binding but could be inhibited with bacitracin. The other occurred after internalization and was partially blocked with chloroquine. Thus, after binding, bombesin is internalized, and the degradation products are released from the cell. Exposure to bombesin is also accompanied by a subsequent decrease in cell surface binding (54% after 1 h exposure to 100 nM bombesin), suggesting that the bombesin receptor may also be internalized in a ligand-dependent manner.