The 45Ca2+ uptake in beta-cell-rich ob/ob-islets was measured using the La3+ wash technique. Tetracaine (1 mM) markedly enhanced the 45Ca2+ net uptake (120 min) in the presence of 3 mM glucose, and at 7 and 20 mM glucose there were clear tendencies to dose-dependent increases with 0.1 to 1 mM tetracaine. Glibenclamide 1 microM to 0.2 mM, stimulated the 45Ca2+ net uptake in the presence of 3 mM glucose and 0.1 mM to 0.2 mM glibenclamide potentiated the uptake in the presence of 7 mM glucose. When the drugs were added for only a 10 min incubation period, glibenclamide, 1 microM to 0.2 mM, but not tetracaine (10 microM to 1 mM) increased the short-term uptake of 45Ca2+. After preincubation with either of the drugs, neither tetracaine (10 microM to 1 mM) nor glibenclamide (10 nM to 0.2 mM) had any effect on the short-term 45Ca2+ uptake. In islets incubated with 45Ca2+ and tetracaine and washed without La3+ the apparent net uptake of 45Ca2+ was reduced by 0.5 to 1 mM tetracaine both at 3 and 20 mM glucose. Tetracaine (0.5 mM) stimulated the 45Ca2+ efflux in the presence of 3 mM glucose. The results show that both drugs affected the Ca2+ handling. It is suggested that glibenclamide mainly increases Ca2+ influx by voltage-dependent pathways, whereas tetracaine, at certain concentrations, mobilizes Ca2+ from intracellular stores in the islet cells.