Similar to other endocrine glands, the endocrine pancreas displays a characteristic topography of its constituent cells. The functional significance of this structural organization was examined by measuring the secretory activity of the B cell in rat islet cell preparations of different composition. Glucose released 30-fold more insulin from B cells lodged within intact islets as from purified single B cells; structurally coupled B cells and single B cells isolated with A cells or incubated with glucagon responded 4- and 2-fold, respectively, more effectively to glucose than single B cells alone. Glucose homeostasis is thus dependent not only on the number and integrity of the insulin-containing B cells but also on their interactions with neighboring B and non-B cells. This study provides direct support for the concept that the microanatomy of the islet creates the anatomical basis for functional cooperation between islet cells and hence for an appropriate glucose-induced insulin release.