Modulation of prolactin-stimulated Nb2 lymphoma cell mitogenesis by cholera toxin and pertussis toxin

We have investigated whether cholera toxin (CT)- or pertussis toxin (IAP)-sensitive G proteins are involved in ovine (o) PRL-stimulated mitogenesis in the lactogen-dependent rat Nb2 node lymphoma cell line. Addition of IAP to medium caused a biphasic effect on oPRL-stimulated cell number. Low doses (10(-3) ng/ml) enhanced (mean +/- SEM, 15 +/- 3%) whereas higher doses (greater than or equal to 10 ng/ml) inhibited (24 +/- 3%) mitogenesis stimulated by a submaximal dose of oPRL (0.1 ng/ml) compared to control values. The cAMP analog 8-bromo-cAMP also had a biphasic effect on cell division stimulated by submaximal doses of PRL. Low doses (10(-5) M) enhanced whereas higher doses (10(-3) M) inhibited Nb2 cell growth in response to PRL. Incubation with CT only inhibited oPRL-stimulated mitogenesis in a dose-dependent manner. Maximal inhibition (63 +/- 7%) occurred at a concentration of 10 ng/ml or more. Phorbol myristate acetate (PMA) enhanced mitogenesis stimulated by PRL alone and in the presence of either stimulatory or inhibitory doses of IAP, but PMA did not block IAP inhibition. In contrast, PMA had no effect on cells incubated with CT; the inhibition of PRL-stimulated cell division by CT remained unchanged. Lactogenic receptor-binding sites per cell and affinity were not significantly affected by PMA, IAP, or CT, suggesting a postreceptor mechanism of action. In summary, these data demonstrate that cAMP modifies PRL-stimulated Nb2 cell mitogenesis. The differences between IAP and CT (i.e. biphasic effect, degree of inhibition, and differential effect of PMA) suggest that these agents could also modulate PRL actions in the Nb2 cell through different mechanisms, including a cAMP-independent pathway.