Recently, we have shown that T cells exposed to concentrations of prostaglandin E2 (PGE2) or the beta-adrenergic receptor agonist isoproterenol (ISO) that elicit equimolar levels of cAMP accumulation do not inhibit anti-CD3 monoclonal antibody-induced T cell proliferation to the same extent. This report extends these studies by investigating the induction of cAMP-dependent protein kinase (PKA) in T cells stimulated with PGE2 or ISO. The kinetics of PKA activity induced by PGE2 or ISO in T cells are similar but PGE2 induces more PKA activity. When T cells were treated with concentrations of PGE2 or ISO that elicited similar PKA activities, PGE2 was found to be more immunosuppressive than ISO. T cells stimulated with PGE2 or ISO showed similar levels of increased PKA activity in both the cytosolic and the particulate fractions. Quantitation of the activity of PKA I and PKA II isozymes in T cells stimulated with PGE2 or ISO revealed that both types were activated; however, while PGE2 induced the utilization of an equal amount of both isozymes in T cells, ISO-treated cells utilized twice as much PKA I compared to PKA II. Overall, these results suggest that qualitative differences in the concentration of cAMP and PKA activity are important elements in modulatory T cell proliferative responses.