The Na+/Ca2+ exchangers are key regulators for Ca2+ homeostasis in pancreatic β cells, but their exact role in insulin secretion is not fully understood. In the present study, we investigated the role of Na+/Ca2+ exchangers on cytosolic Ca2+ dynamics and exocytosis in INS-1 cells. We newly discovered that the Li-permeable Na+/Ca2+ exchangers (NCLX), which are known as mitochondrial Na+/Ca2+ exchangers, contribute to plasma membrane Na+-dependent Ca2+ transport, and confirmed the presence of NCLX in the plasma membrane using immunocytochemistry and cell surface biotinylation experiments. We further investigated the role of NCLX on exocytosis function by measuring depolarization-induced capacitance increase in siNCLX-transfected INS-1 cells. Downregulation of NCLX significantly suppressed the second phase capacitance increase without affecting the first phase, but inhibition of mitochondrial Na+/Ca2+ exchange alone by removing intracellular Na+ did not cause a significant effect. These results suggest that normal surface NCLX activity is required for vesicle recruitment in the second phase. In addition, we discovered that oligomycin inhibited the second phase capacitance increase in the presence of intracellular ATP, suggesting that local metabolic control by mitochondria is critical for this phase. Our data indicate that the activity of surface NCLX may regulate local Ca2+ to optimize mitochondrial local metabolic control and thus contributes to vesicle recruitment for the normal secretory function in pancreatic β cells.