This study evaluated the contribution of isozymes of cAMP phosphodiesterase (PDE) to total PDE activity in human and rat islets using type-selective inhibitors. The effects of selected PDE inhibitors on insulin secretion from human and rat islets were also measured in order to assess the contribution of the various PDE isozymes to the modulation of insulin secretion. The data suggest that PDE III is quantitatively the most important PDE isozyme present in islets, accounting for up to 70% of the total activity. Lower, but measurable, levels of PDE IV activity were present. Approximately 20% of islet PDE is not inhibitable by agents selective either for PDE III or IV. Selective inhibition of PDE III stimulated insulin secretion, but inhibition of PDE IV had no effect. The effects of type-selective inhibitors on PDE activity and insulin secretion were similar in human and rat islets.