Significance We reveal that increased expression of Ca _V 3.1 channels in rat islets selectively impairs first-phase glucose-stimulated insulin secretion. This deterioration is recapitulated in human islets. Its causal role in diabetes development is clearly manifested in an in vivo diabetic model. Mechanistically, this is due to reduction of phosphorylated FoxO1 in the cytoplasm, elevated FoxO1 nuclear retention, and decreased syntaxin 1A, SNAP-25, and synaptotagmin III in a Ca _V 3.1 channel- and calcineurin-dependent manner. Our findings suggest that elevated expression of Ca _V 3.1 channels in pancreatic islets drives FoxO1-mediated down-regulation of exocytotic proteins resulting in the diabetic phenotypes of impaired insulin secretion and aberrant glucose homeostasis. This causal connection pinpoints β cell Ca _V 3.1 channels as a potential druggable target for antidiabetes therapy.