The extracellular matrix (ECM) critically affects β cell functions via integrin activation. But whether these ECM actions drive the spatial organization of β cells, as they do in epithelial cells, is unknown. Here, we show that within islets of Langerhans, focal adhesion activation in β cells occurs exclusively where they contact the capillary ECM (vascular face). In cultured β cells, 3D mapping shows enriched insulin granule fusion where the cells contact ECM-coated coverslips, which depends on β1 integrin receptor activation. Culture on micro-contact printed stripes of E-cadherin and fibronectin shows that β cell contact at the fibronectin stripe selectively activates focal adhesions and enriches exocytic machinery and insulin granule fusion. Culture of cells in high glucose, as a model of glucotoxicity, abolishes granule targeting. We conclude that local integrin activation targets insulin secretion to the islet capillaries. This mechanism might be important for islet function and may change in disease.