DIFFERENTIAL-EFFECTS OF PEPTIDE HISTIDINE ISOLEUCINE (PHI) AND RELATED PEPTIDES ON STIMULATION AND SUPPRESSION OF NEUROBLASTOMA CELL-PROLIFERATION - A NOVEL VIP-INDEPENDENT ACTION OF PHI VIA MAP KINASE

The growth rate of rodent embryonic neuroblasts and human neuroblastom a cell lines is regulated in part by autocrine or paracrine actions of neuropeptides of the family that includes vasoactive intestinal pepti de (VIP), peptide histidine isoleucine (PHI), and pituitary adenylate cyclase-activating peptide (PACAP). These peptides act via seven trans membrane G-protein-linked receptors coupled to cAMP elevation, phospho lipase C activation, intracellular Ca2+ release, and/or of mitogen-act ivated protein (MAP) kinase activation. Here we investigated the actio n of these peptides on the mouse neuroblastoma cell line Neuro2a PHI a nd VIP inhibited proliferation at concentrations as low as 10(-13) ns and 10(-10) M, respectively. In contrast, PACAP action was biphasic, w ith stimulation occurring at subnanomolar doses and inhibition at high er doses. Peptide actions were studied further by measuring cAMP and E RK1/2 MAP kinase activity and by assessing H-3-thymidine in corporatio n in conjunction with a panel of signal transduction pathways inhibito rs. The data obtained indicated that the PHI-inhibitory and PACAP-stim ulatory activities were mediated by corresponding changes in activity of the MAP kinase pathway and independent of protein kinase A (PRA) or protein kinase C (PKC). In contrast, the inhibitory actions of VIP an d PACAP were specifically blocked by antagonists of PKA Northern blot analysis revealed gene expression for only the PACAP-preferring (PAC(1 )) receptor. However, binding experiments using I-125-labeled PACAP27, PHI, and VIP, demonstrated the presence of PACAP-preferring sites, bi valent VIP/PACAP sites, and PHI-binding sites that did not interact wi th VIP. The studies demonstrate potent regulatory actions of PACAP, PH I, and VIP on neuroblastoma cell proliferation which al,pear to be med iated by multiple subsets of receptors which differentially couple to MAP kinase and PKA signaling pathways.