Single islet cells in monolayer cultures of neonatal rat pancreas were microinjected with the fluorescent dye Lucifer Yellow CH and the cultures were observed by combined phase contrast and fluorescent microscopy. The dye spread from an injected cell directly into neighboring islet cells, and successive microinjections of dye into different cells defined territories comprised of 2–6 communicating cells. The number of communicating cells could be modulated by addition to the cultures of different agents known to affect islet cell secretory activity. Cell communication was significantly increased by a high (16.7 mM) glucose concentration, by the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX, 0.1 mM), and by the calcium ionophore, A23187. The effect of A23187 was transient and dose-dependent. Somatostatin (1 μg/ml) significantly inhibited cell communication. These results demonstrate that cell-to-cell communication may participate in the regulation of islet cell secretory activity.