Monolayer islet cells prepared from neonatal rat pancreases were cultured in media with 5.5 mM glucose alone or supplemented with 1 mM 2-deoxy-2-fluoroglucose for a total of 7 days. The recovery of insulin in the cells and media, and the insulin secretory responses to four secretagogues were determined on several occasions. Under culture conditions with a basal medium containing 5.5 mM glucose, the responses to insulin secretagogues tested were abolished after 7 days of culture. In contrast, the addition of 2-deoxy-2-fluoroglucose maintained the insulin secretory responses to glucose (2.8 approximately 33.4 mM), leucine (2.5 approximately 10 mM) and 2-ketoisocaproate (2.5 approximately 10 mM) at levels several times higher than at day 0. Moreover, in these monolayer islet cells the rates of 0.62 mM [U-14C]glutamine oxidation were significantly increased by leucine or 2-ketoisocaproate at 10 mM. From these results, it is suggested that a series of metabolic adaptations in response to change in nutritional status (possibly attributable to the action of 2-deoxy-2-fluoroglucose as an inhibitor of glycolysis) may facilitate the metabolism of amino acids in B cells. Such an activation may be associated with the survival and maturation of neonatal B cells in vitro.