Acute insulin treatment in rats has recently been shown to cause a rapid increase in liver low-Km cAMP phosphodiesterase (PDE) activity, which selectively affects Golgi fractions. To assess the physiological significance of this observation, the cAMP PDE activity associated with liver Golgi fractions has been measured in genetically obese Zucker rats, which spontaneously develop hyperinsulinemia, in rats receiving a continuous infusion of insulin, and in rats treated with anti-insulin serum. In genetically obese Zucker rats, a significant increase in Golgi-associated cAMP PDE relative to age-matched lean animals occurred after 3 wk, coinciding with the development of hyperinsulinemia. This change was maximal at 5-8 wk and affected the light (Gl) and intermediate (Gi) Golgi fractions (100-110% increase) to a greater extent than the heavy (Gh) fraction (30% increase). After 7 wk, despite the further increase in insulinemia, the increase in Golgi-associated cAMP PDE became progressively less marked, and at 18 wk it was no longer detectable except in Gh, suggesting the development of a hepatic insulin resistance. Infusion of insulin through chronically implanted intracardiac catheters led to a 30-50% increase in Golgi-associated cAMP PDE, which occurred earlier in Gi (3 h) than in Gh (7 h) and persisted for greater than 96 h. Injection of anti-insulin serum led to a 30-50% decrease in Golgi-associated cAMP PDE, which occurred sequentially in Gl (5 min), Gi (15 min), and Gh (30 min) and affected predominantly Gl and Gh. These results suggest that the cAMP PDE associated with Golgi fractions is a physiological effector of plasma insulin in vivo.