Overexpression of hepatocyte growth factor (HGF) in the β-cell of transgenic mice enhances β-cell proliferation, survival, and function. In the current studies, we have used conditional ablation of the c-met gene to uncover the physiological role of HGF in β-cell growth and function. Mice in which c-met is inactivated in the β-cell (MetCKO mice) display normal body weight, blood glucose, and plasma insulin compared with control littermates. In contrast, MetCKO mice displayed significantly diminished glucose tolerance and reduced plasma insulin after a glucose challenge in vivo. This impaired glucose tolerance in MetCKO mice was not caused by insulin resistance because sensitivity to exogenous insulin was similar in both groups. Importantly, in vitro glucose-stimulated insulin secretion in MetCKO islets was decreased by ∼50% at high glucose concentrations compared with control islets. Furthermore, whereas insulin and glucokinase expression in MetCKO islets were normal, GLUT-2 expression was decreased by ∼50%. These changes in β-cell function in MetCKO mice were not accompanied by changes in total β-cell mass, islet morphology, islet cell composition, and β-cell proliferation. Interestingly, however, MetCKO mice display an increased number of small islets, mainly single and doublet β-cells. We conclude that HGF/c-met signaling in the β-cell is not essential for β-cell growth, but it is essential for normal glucose-dependent insulin secretion.