Cyclic nucleotide phosphodiesterase from isolated mouse islets of Langerhans has been investigated and characterized. The enzyme has a pH optimum of 8.5 and is magnesium dependent, 10 mM. magnesium causing a fivefold increase in V max. Calcium is without effect on the enzyme. It is inhibited by theophylline and parachloromercuribenzoate. Glucose, has no effect on the activity of the enzyme. Tolbutamide at concentrations equivalent to those found under clinical conditions is a noncompetitive inhibitor of the enzyme.