Cyclosporine (CsA) prevented acute rejection of intrahepatic canine islet allografts (IHIA) in 5/5 pancreatectomized dogs. Normal fasting blood glucose levels were sustained in these dogs for 210 +/- 78 days (mean +/- SEM) following withdrawal of CsA. We tested whether combining islets from more than one pancreas would improve function and prolong islet allograft survival during and following administration of CsA. Areas under the glucose disappearance (GDA) and C-peptide response (CPA) curves following i.v. glucose (0.5 g/kg), 1 month posttransplant were not significantly different using islets from 1 or 2 pancreases, whereas GDA and CPA approached normal if islet yields from 3 or more pancreases were combined. Mean islet allograft survival following interruption of CsA decreased with an increase in the number of donor pancreases (one: 210 +/- 78 days, vs. two: 113 +/- 23 days, vs. greater than 2: 57 +/- 5 days). These studies demonstrate that: (1) IHIA uniformly resulted in fasting euglycemia in 36 of 38 diabetic dogs treated with CsA; (2) normal i.v. glucose metabolism required the combined islet yield of 3 or more donor pancreases, suggesting that a substantial number of intrahepatic islet cells are functionally lost despite effective CsA-induced immunosuppression; (3) use of multiple donors to accumulate an increased mass of islets may immunologically compromise allograft survival following discontinuation of CsA (these experiments, however, do not exclude a direct relationship between the duration of CsA therapy and the duration of immune unresponsiveness following interruption of CsA in multidonor islet allografts as an independent variable); and (4) unmodified islets obtained from multiple donors seem to require continuous immunosuppression to prevent rejection.