The previous finding that intracellular transport of secretory proteins in the rat exocrine pancreas is accelerated by in vivo stimulation with a pancreatic secretagogue has been further analyzed. Using a radioassay for discharge of newly synthesized proteins, the rate of release was compared in control and prestimulated lobules. In control preparations discharge occurred with an initial lag period of 30 minutes and a maximum after two hours of incubation. After in vivo infusion of 5 x 10(-8) g/hr. caerulein for 24 h in vitro discharge started after 10 minutes of in vitro incubation and attained a maximal rate after one hour. Using the same radioassay and several inhibitors of intracellular transport and granule discharge, it could be demonstrated that both processes were reduced to the same extent in controls and in lobules with accelerated transport. To obtain direct evidence for the degree of acceleration of the different transport steps between rough endoplasmic reticulum, Golgi complex and zymogen granules, the respective subcellular fractions of these organelles prepared and characterized ultrastructurally and biochemically. The rate of disappearance of newly formed proteins from rough microsomes and the appearance in smooth microsomes and zymogen granules were significantly increased after in vivo stimulation. The data substantiate an acceleration of the regular transport steps by the secretagogue. There was no indication that a high level of secretory activity leads to a rerouting of secretory proteins or to an omission of one of the regular steps in intracellular transport.