Somatostatin potently inhibits insulin secretion from pancreatic -cells. It does so via activation of ATP-sensitive K⁺-channels (K_ATP) and G protein regulated inwardly rectifying K⁺-channels (GIRK) which act to decrease voltage-gated Ca²⁺-influx, a process central to exocytosis. Since K_ATP channels, and indeed insulin secretion, is controlled by glucose oxidation, we have investigated whether somatostatin inhibits insulin secretion by direct effects on glucose metabolism. Oxidative metabolism in -cells was monitored by measuring changes in the O₂ consumption (O₂) of isolated mouse islets and MIN6 cells, a murine-derived -cell line. In both models, glucose-stimulated O_2, an effect closely associated with inhibition of K_ATP channel activity and induction of electrical activity (r > 0.98). At 100 nM, somatostatin abolished glucose-stimulated O₂ in mouse islets (n = 5, P < 0.05) and inhibited it by 80%± 28% (n = 17, P < 0.01) in MIN6 cells. Removal of extracellular Ca²⁺, 5 mM Co²⁺ or 20 µM nifedipine, conditions that inhibit voltage-gated Ca²⁺ influx did not mimic, but either blocked or reduced the effect of the peptide on O₂. The nutrient secretagogues, methylpyruvate (10 mM) and -ketoisocaproate (20 mM) also stimulated O₂ but this was unaffected by somatostatin. Somatostatin also reversed glucose-induced hyperpolarization of the mitochondrial membrane potential monitored using rhodamine-123. Application of somatostatin receptor selective agonists demonstrated that the peptide worked through activation of the type 5 somatostatin receptor (sstr₅). In conclusion, somatostatin inhibits glucose metabolism in murine -cells by an unidentified Ca²⁺-dependent mechanism. This represents a new signaling pathway by which somatostatin can inhibit cellular functions regulated by glucose metabolism.