Neural and endocrine factors (i.e., Ach and GLP-1) restore defective glucose-stimulated insulin release in pancreatic islets lacking sulfonylurea type 1 receptors (SUR1^−/−) (Doliba NM, Qin W, Vatamaniuk MZ, Li C, Zelent D, Najafi H, Buettger CW, Collins HW, Carr RD, Magnuson MA, and Matschinsky FM. Am J Physiol Endocrinol Metab 286: E834–E843, 2004). The goal of the present study was to assess fuel-induced respiration in SUR1^−/− islets and to correlate it with changes in intracellular Ca²⁺, insulin, and glucagon secretion. By use of a method based on O₂ quenching of phosphorescence, the O₂ consumption rate (OCR) of isolated islets was measured online in a perifusion system. Basal insulin release (IR) was 7–10 times higher in SUR1^−/− compared with control (CON) islets, but the OCR was comparable. The effect of high glucose (16.7 mM) on IR and OCR was markedly reduced in SUR1^−/− islets compared with CON. Ach (0.5 μM) in the presence of 16.7 mM glucose caused a large burst of IR in CON and SUR1^−/− islets with minor changes in OCR in both groups of islets. In SUR1^−/− islets, high glucose failed to inhibit glucagon secretion during stimulation with amino acids or Ach. We conclude that 1) reduced glucose responsiveness of SUR1^−/− islets may be in part due to impaired energetics, as evidenced by significant decrease in glucose-stimulated OCR; 2) elevated intracellular Ca²⁺ levels may contribute to altered insulin and glucagon secretion in SUR1^−/− islets; and 3) The amplitudes of the changes in OCR during glucose and Ach stimulation do not correlate with IR in normal and SUR1^−/− islets suggesting that the energy requirements for exocytosis are minor compared with other ATP-consuming reactions.