Pancreatic islets from BALB/c mice were transplanted to the kidney of syngenic hosts. After 1-40 weeks, the grafts were removed, perifused in vitro, and extracted. Fresh islets were similarly examined. The graft insulin content fell by 70% in 1 week and remained low throughout the observation period. In contrast, rates of basal or glucose-stimulated insulin release were not much, if at all, decreased. In fresh islets and grafts removed after 3 or 28 weeks, 2 consecutive pulses of glucose stimulation, 20-25 min long and separated by 20 min at basal glucose concentration (2 or 2.8 mmol/L), elicited the same insulin secretory response. When 10 mumol/L acetylcholine and 10 mumol/L eserine were present during the second pulse, the glucose-stimulated insulin release from fresh islets was potentiated as much as 11-fold. This potentiation was reduced by one half during the first week of transplantation, and subsequently by 80-90%. It is concluded that vagal deprivation rapidly induces a state of persistent cholinergic refractoriness in transplanted beta-cells, despite morphological signs of autonomic reinnervation of the grafts.