Single B-cells have been previously shown to respond poorly to glucose, in contrast to B-cells lodged in intact islets or in small groups of structurally coupled B-cells, isolated as such from islets. To analyze the role of cell coupling in glucose-induced insulin release, single B-cells were reaggregated in vitro and then tested for their secretory capability. Glucose as well as (Bu)2cAMP stimulated the degree of reaggregation during short shaking incubations (up to 180 min); onset of this process was most rapidly observed with (Bu)2cAMP (within 20 min), but after 180 min a comparable extent was measured with either 20 mM glucose or 0.5 mM (Bu)2cAMP. Calcium was an absolute prerequisite for reaggregation of B-cells. Glucose-induced insulin release from reaggregated B-cells was 4-fold higher than from single B-cells; this difference was not caused by some metabolic priming effect of glucose or (Bu)2cAMP, but appeared primarily mediated by the aggregated state of the cells. It is concluded that the secretory response of pancreatic B-cells is highly dependent on their aggregation with other B-cells. Both glucose and cAMP promote the adhesion of B-cells, and this may contribute to their well known insulinotropic effects.