ABSTRACT

No abstract

Neurogenesis in the peripheral and central nervous systems proceeds in region-specific fashion, although underlying mechanisms remain undefined. Emerging evidence indicates that the neuropeptide PACAP and its G-protein-coupled receptor are expressed widely in the embryonic brain, suggesting that the ligand/receptor system plays a role in development. We found previously that PAC1-R activation elicited opposing mitogenic effects in neurogenetic cultures, stimulating peripheral sympathetic neuroblasts while inhibiting cerebral cortical precursors. We have now defined the expression of PAC1-R mRNA isoforms and activation of second-messenger pathways in these model populations. Sympathetic neuroblasts express the "hop" receptor isoform, through which PACAP elicits increased levels of cAMP and activation of the PI signaling pathway. In contrast, cerebral cortical precursors express primarily the "short" (non-insert) receptor isoform and exhibit increased cAMP levels alone following PACAP treatment. Thus, opposing mitogenic regulation in sympathetic and cortical precursors correlates with differential receptor isoform expression and distinct second-messenger signaling. In addition to receptor, PACAP ligand mRNA was expressed by both populations, suggesting that the peptide is produced and acts locally to regulate precursor proliferation. These observations indicate that the PACAP ligand/receptor system is expressed in both the peripheral and central nervous system during development. More generally, these studies suggest that widely expressed extracellular factors mediate region-specific neurogenesis by activating lineage-restricted receptor isoforms and intracellular pathways.

The PACAP ligand/type I receptor system is expressed throughout the embryonic nervous system, suggesting roles in regulating neural patterning and neurogenesis. In the forebrain, precursors of the six-layered cerebral cortex cease dividing in a highly reproducible spatiotemporal sequence. The time of cell cycle exit in fact determines neuron laminar fate. Our studies indicate that PACAP signaling may elicit cortical precursor withdrawal from the cell cycle, antagonizing mitogenic stimulators. PACAP inhibited embryonic day 13.5 rat cortical precursor [3H]thymidine incorporation, decreasing the proportion of mitotic cells. PACAP promoted morphological and biochemical differentiation, indicating that PACAP-induced cell cycle withdrawal was accompanied by neuronal differentiation. In vivo, embryonic cortex contains PACAP. In culture, 85% of cells expressed PACAP while 64% exhibited receptor. Co-localization studies indicated that PACAP ligand and receptor were expressed by the mitotic precursors that divided in response to bFGF, suggesting that precursors integrate mitogenic and anti-mitogenic signals to determine the timing of cell cycle exit. The expression of PACAP ligand and receptor in precursors raised the possibility of autocrine function. Indeed, peptide antagonists increased proliferation, suggesting that the PACAP system is expressed to elicit cell cycle exit. During ontogeny, an inhibitory signal, such as PACAP, may be required to counter the stimulatory activity of mitogenic bFGF and IGFI whose expression during cortical neurogenesis is sustained. The dynamic interplay of positive and negative regulators would regulate the timing of cell cycle withdrawal, and thus neuronal phenotype and laminar position.

No abstract