Cyclic AMP counteracts mitogen-induced inositol phosphate generation and increases in intracellular Ca2+ concentrations in human lymphocytes

1. The effects of increases in intracellular adenosine 3':5'-cyclic monophosphate (cyclic AMP) on mitogen-induced generation of inositol phosphates and increases in intracellular Ca2+ concentration were investigated in human peripheral blood mononuclear leukocytes (MNL). 2. The mitogens concanavalin A (Con A), pokeweed mitogen (PWM) and phytohaemagglutinin (PHA) concentration-dependently stimulated generation of inositol phosphates. Catecholamines inhibited this process with an order of potency: isoprenaline greater than adrenaline greater than noradrenaline indicating involvement of beta 2-adrenoceptors. This order of potency was also consistent with the catecholamine potencies for stimulating the generation of cyclic AMP. 3. In addition to catecholamines, the cyclic AMP formation-stimulating agents prostaglandin E1 (PGE1) and forskolin concentration-dependently inhibited mitogen-induced inositol phosphate generation, too. Moreover, the inhibitory effect of isoprenaline was potentiated by co-incubation with the phosphodiesterase inhibitor isobutylmethylxanthine demonstrating that these inhibitory effects were mediated by cyclic AMP. 4. Con A and PHA concentration-dependently increased the intracellular Ca2+ concentration in human MNL (assessed by the fluorescent indicator dye Fura-2). This increase was almost completely blocked by chelation of extracellular Ca2+, demonstrating influx rather than mobilization from intracellular stores. 5. The elevation of intracellular Ca2+ was not blocked by pretreatment with pertussis toxin, 100 ng ml-1, for 16 h. 6. Isoprenaline, PGE1, and forskolin, however, inhibited the mitogen-stimulated elevation of intracellular Ca2+. This inhibition was enhanced by the phosphodiesterase inhibitors isobutylmethylxanthine and Ro 20-1724, demonstrating mediation by cyclic AMP. 7. We conclude that catecholamines and other cyclic AMP increasing agents can inhibit mitogen-stimulated generation of inositol phosphates and elevation of intracellular Ca2+ in resting human MNL