PACAP-38 Induces Transcriptomic Changes in Rat Trigeminal Ganglion Cells Related to Neuroinflammation and Altered Mitochondrial Function Presumably via PAC1/VPAC2 Receptor-Independent Mechanism - PubMed (original) (raw)

Krisztina Takács-Lovász et al. Int J Mol Sci. 2022.

Abstract

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a broadly expressed neuropeptide which has diverse effects in both the peripheral and central nervous systems. While its neuroprotective effects have been shown in a variety of disease models, both animal and human data support the role of PACAP in migraine generation. Both PACAP and its truncated derivative PACAP(6-38) increased calcium influx in rat trigeminal ganglia (TG) primary sensory neurons in most experimental settings. PACAP(6-38), however, has been described as an antagonist for PACAP type I (known as PAC1), and Vasoactive Intestinal Polypeptide Receptor 2 (also known as VPAC2) receptors. Here, we aimed to compare the signaling pathways induced by the two peptides using transcriptomic analysis. Rat trigeminal ganglion cell cultures were incubated with 1 µM PACAP-38 or PACAP(6-38). Six hours later RNA was isolated, next-generation RNA sequencing was performed and transcriptomic changes were analyzed to identify differentially expressed genes. Functional analysis was performed for gene annotation using the Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Reactome databases. We found 200 common differentially expressed (DE) genes for these two neuropeptides. Both PACAP-38 and PACAP(6-38) treatments caused significant downregulation of NADH: ubiquinone oxidoreductase subunit B6 and upregulation of transient receptor potential cation channel, subfamily M, member 8. The common signaling pathways induced by both peptides indicate that they act on the same target, suggesting that PACAP activates trigeminal primary sensory neurons via a mechanism independent of the identified and cloned PAC1/VPAC2 receptor, either via another target structure or a different splice variant of PAC1/VPAC2 receptors. Identification of the target could help to understand key mechanisms of migraine.

Keywords: intracellular calcium; mitochondrial electron transport chain; pituitary adenylated cyclase-activating polypeptide (PACAP); transcriptomics; trigeminal ganglion.

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Conflict of interest statement

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Figures

Figure 1

Figure 1

Transcripts Per Million (TPM) values for receptors with known or potential affinity for PACAP in untreated control trigeminal ganglion (TG) cell cultures. Data represent mean ± SD (n = 3).

Figure 2

Figure 2

Numerical representation of differentially expressed genes for PACAP-38 and PACAP6-38. The expression of genes was compared to the respective untreated control groups.

Figure 3

Figure 3

Heatmap of fold change (FC) values for differentially expressed (DE) genes shared in trigeminal ganglion (TG) cell cultures treated with PACAP-38 or PACAP(6-38) compared to the control group (1 µM, 6 h). p < 0.05 was considered as significantly different. (See the list of gene name abbreviations in Appendix A). DE genes associated with neurological disorders were selected from total hit according to databases (

https://rgd.mcw.edu/

;

https://www.genecards.org/

;

https://www.ncbi.nlm.nih.gov

last accessed date: 10 April 2021).

Figure 4

Figure 4

Highlighted GO analysis results which were significant in both PACAP-38- (red) and PACAP(6-38)- (blue) treated trigeminal ganglion cells (1 µM, 6 h). Panel A represents the number of differentially expressed (DE) genes related to each GO term, while panel B shows p values for the identical terms (p < 0.05).

Figure 5

Figure 5

Significantly affected Reactome pathways found for both PACAP-38 and PACAP(6-38)-treated cultures ranked according to the _p_-value. Blue bars indicate downregulated, while red bars show upregulated pathways.

Figure 6

Figure 6

The KEGG Ca2+ signaling pathway was significantly altered in both PACAP-38 and PACAP(6-38)-treated trigeminal ganglion cells. Rectangles in color indicate genes that were significantly changed after treatment (left sides of rectangles represent PACAP-38, while right sides stand for PACAP(6-38)). The red and blue color scale show normalized fold change values for each differentially expressed gene. The KEGG Ca2+ signaling pathway also significantly implicated by ranked list enrichment performed on all genes can be found in Supplementary Figure S1.

Figure 7

Figure 7

Schematic representation of the hypothetical mechanisms initiated by PACAP in trigeminal ganglion cells. PACAP-38/PACAP6-38 activate G- protein coupled receptors (GPCR). Activation of Guanine nucleotide-binding protein G(i) subunit alpha-1 (GnaI) leads to protein kinase A (PKA; Protein Kinase cAMP-Activated Catalytic Subunit Beta—Prkacb) activation via increased cAMP levels. Phosphorylation by PKA may induce TRPM8 channel inhibition which could cause overexpression of TRPM8 ion channel. Calcium overload could lead to suppressed Complex I biogenesis.

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References

    1. Somogyvári-Vigh A., Reglődi D. Pituitary Adenylate Cyclase Activating Polypeptide: A Potential Neuroprotective Peptide. Curr. Pharm. Des. 2004;10:2861–2889. doi: 10.2174/1381612043383548. - DOI - PubMed
    1. Reglődi D., Kiss P., Lubics A., Tamás A. Review on the Protective Effects of PACAP in Models of Neurodegenerative Diseases in Vitro and in Vivo. Curr. Pharm. Des. 2011;17:962–972. doi: 10.2174/138161211795589355. - DOI - PubMed
    1. Dickson L., Finlayson K. VPAC and PAC Receptors: From Ligands to Function. Pharmacol. Ther. 2009;121:294–316. doi: 10.1016/j.pharmthera.2008.11.006. - DOI - PubMed
    1. Vaudry D., Falluel-Morel A., Bourgault S., Basille M., Burel D., Wurtz O., Fournier A., Chow B.K.C., Hashimoto H., Galas L., et al. Pituitary Adenylate Cyclase-Activating Polypeptide and Its Receptors: 20 Years after the Discovery. Pharm. Rev. 2009;61:283–357. doi: 10.1124/pr.109.001370. - DOI - PubMed
    1. Harmar A.J., Fahrenkrug J., Gozes I., Laburthe M., May V., Pisegna J.R., Vaudry D., Vaudry H., Waschek J.A., Said S.I. Pharmacology and Functions of Receptors for Vasoactive Intestinal Peptide and Pituitary Adenylate Cyclase-Activating Polypeptide: IUPHAR Review 1. Br. J. Pharmacol. 2012;166:4–17. doi: 10.1111/j.1476-5381.2012.01871.x. - DOI - PMC - PubMed

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