Pancreatic islet cells from normal adult rats were purified by autofluorescence-activated cell sorting (FACS). A purification of islet beta-cells (greater than 90%) and alpha-cells (greater than 70%) was obtained after FACS separation. Dual-wavelength microfluorimetry was employed for measurements of cytoplasmic Ca2+ concentration (Ca2+i) in single beta- and alpha-cells. In FACS-purified beta-cells, 11.8 mM glucose induced oscillations of Ca2+i with a frequency of 1 cycle per 2-4 min, whereas 20 mM glucose induced a rapid rise in Ca2+i followed by a sustained elevation in Ca2+i. In contrast, alpha-cells exhibited a decrease in Ca2+i following exposure to 20 mM glucose, and subsequent exposure to 20 mM arginine induced a large increase in Ca2+i with periodic fluctuations. FACS purification of a heterogeneous islet-cell population provides a unique model for characterizing secretagogue-induced changes in Ca2+i in single beta-and alpha-cells.