Pituitary adenylate cyclase-activating polypeptide promotes cell survival and neurite outgrowth in rat cerebellar neuroblasts
Section snippets
Peptides
PACAP38 and VIP were synthesized by the solid phase methodology described previously.[7]PACAP(6–38) was from the American Peptide Company (Sunnyvale, CA, U.S.A.).
Cell culture
Granule cell suspensions were prepared from cerebelli of eight-day-old Wistar rats, as described previously.[19]Dispersed cells were seeded in multiwell plates (Costar, Cambridge, MA, U.S.A.) coated with poly-l-lysine (5×10−3 M) at a density of 3.5×105 cells per well. The cells were cultured in a chemically defined culture medium
Morphological examination of cultured granule cells
Five hours after plating, cultured granule cells were homogeneously distributed and exhibited a round shape with very few processes (Fig. 1A). Addition of 10−8 M PACAP38 produced no apparent effect on the overall distribution of cells and the number of processes (Fig. 1B). After 24 h, granule cells were still evenly distributed, but their aspect was markedly different (Fig. 1C) as compared to 2-h cultured cells (Fig. 1A). Most round-shaped cells were dying, while surviving cells generally
Discussion
Previous studies have shown the occurrence of substantial amounts of PACAP in the rat cerebellum during development.34, 45It has also been found that PACAP binding sites are expressed by immature granule cells in the rat cerebellar cortex3, 4and that these recognition sites correspond to authentic type I PACAP receptors positively coupled to the adenylyl cyclase and phospholipase C pathways.5, 14The present study has demonstrated that incubation of cultured immature granule cells with PACAP38
Conclusions
The present report indicates that PACAP promotes cell survival and neurite outgrowth in cultured immature granule cells. The neurotrophic activity of PACAP is mediated through high-affinity type I PACAP receptors. These data, together with previous findings showing the presence of PACAP and PACAP receptors in the rat cerebellar cortex, provide strong evidence for the involvement of PACAP in the regulation of apoptosis and differentiation of germinative cells in the developing cerebellum.
Note added in proof
An effect of PACAP on apoptosis of granule cells has been recently reportted by Cavallaro et al., (1996) Am. Soc. Pharm. exp. Ther. 50, 60-66.
Acknowledgements
This work was supported by the Institut National de la Santé et de la Recherche Médicale (INSERM U413), a France–Québec exchange programme (Coopération Scientifique et Technologique Franco-Québécoise) and the Conseil Régional de Haute-Normandie. H.V. is an Affiliated Professor at the Institut National de la Recherche Scientifique-Santé, Montréal.
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Light-Triggerable Liposomes for Enhanced Endolysosomal Escape and Gene Silencing in PC12 Cells
2017, Molecular Therapy Nucleic AcidsCitation Excerpt :Such enhancement of the gene knockdown was further confirmed by the neurite-inducing activity of PACAP in treated PC12 cells, where a limited neurite outgrowth was an indicator of the response to PAC1R gene knockdown achieved by using lipVP-asODN. PACAP-induced differentiation of PC12 cells is generally investigated by either measuring the levels of signaling molecules involved in this processor or inhibiting the action of PACAP and its receptor with synthetic antagonists, including PACAP6-27 and PACAP6-38.74,75 However our light-triggered liposomal delivery platform incorporating VP provided another useful strategy to interfere with PACAP-induced cell differentiation.