Microdissected pancreatic islets of obese-hyperglycemic mice were used to study the uptake of mannoheptulose by β-cells in relation to the dynamics of mannoheptulose-inhibited insulin release. Mannoheptulose uptake was faster at 37° than at 8° and was inhibited by glucose, 3- O -methylglucose, or phlorizin. The transport rate for mannoheptulose was much lower than that previously observed for glucose. Nevertheless, a few minutes of exposure to 5 mM mannoheptulose were enough to establish intracellular concentrations of the same magnitude as the concentrations inhibiting insulin release and glucose oxidation when added to the medium. This observation makes it possible to explain the prompt inhibition of insulin release noted in microperfusion experiments as due to mannoheptulose interference with the β-cell metabolism of glucose. The mediated and glucose-sensitive transport of mannoheptulose, however, is also compatible with the idea that insulin release is governed by the binding of sugar to a receptor at the β-cell plasma membrane.