PKA, novel PKC isoforms, and ERK is mediating PACAP auto-regulation via PAC1R in human neuroblastoma NB-1 cells

The neuropeptide PACAP is expressed throughout the central and peripheral nervous system where it modulates diverse physiological functions including neuropeptide gene expression. We here report that in human neuroblastoma NB-1 cells PACAP transiently induces its own expression. Maximal PACAP mRNA expression was found after stimulation with PACAP for 3 h. PACAP auto-regulation was found to be mediated by activation of PACAP specific PAC(1)R as PACAP had >100-fold higher efficacy than VIP, and the PAC(1)R selective agonist Maxadilan potently induced PACAP gene expression. Experiments with pharmacological kinase inhibitors revealed that both PICA and novel but not conventional PKC isozymes were involved in the PACAP auto regulation. Inhibition of MAPK/ERK kinase (MEK) also impeded the induction, and we found that PICA, novel PKC and ERK acted in parallel and were thus not part of the same pathways. The expression of the transcription factor EGRI previously ascribed as target of PACAP signalling was found to be transiently induced by PACAP and pharmacological inhibition of either PKC or MEKI/2 abolished PACAP mediated EGRI induction. In contrast, inhibition of PICA mediated increased PACAP mediated EGR1 induction. Experiments using siRNA against EGR1 to lower the expression did however not affect the PACAP auto-regulation indicating that this immediate early gene product is not part of PACAP auto-regulation in NB-I cells. We here reveal that in NB-1 neuroblastoma cells, PACAP induces its own expression by activation of PAC(1)R, and that the signalling is different from the PAC(1)R signalling mediating induction of VIP in the same cells. PACAP auto-regulation depends on parallel activation of PICA, novel PKC isoforms, and ERIC, while EGR1 does not seem to be part of the PACAP auto-regulation. (C) 2016 Elsevier Ltd. All rights reserved.