The aim of the present work was to study glucose production from non-carbohydrate precursors in cases of severe hypoglycemia. An in vivo method was developed, employing trace doses of 14C-alanine or 14C-glycerol. After an i.v. injection of these isotopes, there was a rapid appearance of label in circulating glucose and lactate. In particular 2 cases have been studied. Due to somewhat elevated plasma insulin levels these patients were initially treated with diazoxide and later on subjected to subtotal pancreatectomy. Histopathological examination (Prof. S. Falkmer, Umeå. Sweden) revealed islets of Langerhans that were increased in number as well as size. Although in both cases a decrease was observed in the levels of plasma insulin, there was only a slight clinical improvement after the operation. However, in one of the cases the operation apparently caused a markedly increased ability to produce glucose by the gluconeogenic pathway. This finding suggests that an important pathogenetic mechanism in some cases of hypoglycemia may be an inhibition of gluconeogenesis in liver caused by increased insulin levels.