CROSS-TALK BETWEEN CELLULAR SIGNALING PATHWAYS ACTIVATED BY SUBSTANCE-P AND VASOACTIVE-INTESTINAL-PEPTIDE IN RAT LACTOTROPH-ENRICHED PITUITARY CELL-CULTURES

We have investigated cross-talk between the cAMP/protein kinase A (PKA ) and protein kinase C (PKC)/inositol 1,4,5-trisphosphate (Ins(1,4,5)P -3) messenger systems probed by vasoactive intestinal peptide (VIP) an d substance P (SP), respectively, in rat pituitary cell cultures enric hed in lactotrophs. VIP and forskolin had no effect on the basal distr ibution pattern of the four PKC isozymes (alpha, beta, delta, and zeta ) detectable in lactotroph-enriched cell cultures derived from peripub ertal male rats. whereas both compounds significantly increased transl ocation of PKC alpha and beta from the cytosol to the plasma membrane induced by SP. The delta and zeta subspecies were not affected by VIP and forskolin. Moreover, VIP and forskolin also stimulated SP-induced formation of Ins(1,4,5)P-3 while having no effect on basal inositol ph osphate turnover. The effects of VIP and forskolin on PKC isozyme dist ribution could be blocked by pretreating cells with the PKA inhibitor rp-cAMP. On the other hand, SP potentiated the effect of VIP and forsk olin on cAMP formation while having no effect on the cAMP pathway when it was not triggered by an appropriate agonist. Down-regulation of PK C activity by long term 12-O-tetradecanoylphorbol 13-acetate (TPA) tre atment (24 h) diminished, but did not abolish, the effect of SP on VIP -stimulated cAMP production. Staurosporine and dopamine inhibited the potentiating effect of SP on cAMP accumulation. TPA, which translocate s PKC alpha, beta, and delta in lactotrophs. had a synergistic effect on cAMP formation induced by VIP, but dib also, unlike SP, display cAM P rising abilities when cells were not exposed to VIP and forskolin. D ischarging intracellular Ca2+ by thapsigargin pretreatment had no effe ct on the basal cAMP concentration or the VIP-induced cAMP response, w hereas exposure of cells to SP, thapsigargin, and VIP resulted in a de crease of the cAMP response compared with SP + VIP. The potentiating e ffect uf SP on the VIP response could also be inhibited, but not block ed, by staurosporine. On the basis of these results, it is concluded t hat there exists substantial cross-talk between the cAMP/PKA and PKC/I ns(1,4,5)P-3 messenger systems in lactotroph-enriched cell cultures. K ey effecters seem to be PKA, one or more of PKC alpha, beta, delta and Ins(1,4,5)P-3-sensitive Ca2+ stores.