The role of pancreatic B cell dysfunction in the phase preceding clinical onset of insulin-dependent and non-insulin-dependent diabetes mellitus has been much debated. In this investigation, the impact of a prolonged diabetic environment on pancreatic islet B cells transplanted syngeneically under the kidney capsule of C57BL/6 (B6) and C57BL/Ks (BKs) mice was studied. Alloxan-diabetic mice bearing a subcapsular islet graft insufficient to normalize the blood glucose level were rendered normoglycemic by a second intrasplenic islet graft after various period of hyperglycemia to examine the reversibility of hyperglycemia-induced B cell dysfunction. Using a perfusion technique of the graft-bearing, it was found that both strains of mice exhibited a diminished glucose-induced insulin secretion after 6 wk of hyperglycemia, when compared with normoglycemic mice carrying islet grafts. When normoglycemia was restituted by the splenic graft after 4 or 12 wk, there was a normalization of glucose-stimulated insulin secretion in the renal islet grafts in B6 mice, whereas insulin secretion from the grafted BKs islets remained impaired. Morphometric measurements of the islet grafts demonstrated a 50% reduction in the graft volume in diabetic BKs mice after 12 wk, compared with normoglycemic animals, whereas no such decrease was observed in B6 mice. Islet grafts removed from hyperglycemic mice of both strains exhibited diminished insulin mRNA contents, and in the BKs mice there was also a reduced glucose oxidation rate in the islet grafts in vitro. This metabolic dysfunction can only partly be explained by a reduced graft size. The present findings emphasize the genetic constitution as a decisive factor for the survival and function during a period of sustained stress on a limited B cell mass.