Nuclear factor-κB activated by capacitative Ca2+ entry enhances muscarinic receptor-mediated soluble amyloid precursor protein (sAPPα) release in SH-SY5Y cells

Gq/11 protein-coupled muscarinic receptors are known to regulate the release of soluble amyloid precursor protein (sAPP alpha) produced by alpha-secretase processing; however, their signaling mechanisms remain to be elucidated. It has been reported that a muscarinic agonist activates nuclear factor (NF)-kappa B, a transcription factor that has been shown to play an important role in the Alzheimer disease brain, and that NF-kappa B activation is regulated by intracellular Ca2+ level. In the present study, we investigated whether NF-kappa B activation plays a role in muscarinic receptor-mediated sAPP alpha release enhancement and contributes to a changed capacitative Ca2+ entry (CCE), which was suggested to be involved in the muscarinic receptor-mediated stimulation of sAPP alpha release. Muscarinic receptor-mediated NF-kappa B activation was confirmed by observing the translocation of the active subunit (p65) of NF-kappa B to the nucleus by the muscarinic agonist, oxotremorine M (oxoM), in SH-SY5Y neuroblastoma cells expressing muscarinic receptors that are predominantly of the M3 subtype. NF-kappa B activation and sAPP alpha release enhancement induced by oxoM were inhibited by NF-kappa B inhibitors, such as an NF-kappa B peptide inhibitor (SN50), an I kappa B alpha kinase inhibitor (BAY11-7085), a proteasome inhibitor (MG132), the inhibitor of proteasome activity and I kappa B phosphorylation, pyrrolidine dithiocarbamate, the novel NF-kappa B activation inhibitor (6-amino-4-(4-phenoxyphenylethylamino) quinazoline), and by an intracellular Ca2+ chelator (TMB-8). Furthermore, both oxoM-induced NF-kappa B activation and sAPP alpha release were antagonized by CCE inhibitors (gadolinium or SKF96365) but not by voltage-gated Ca2+-channel blockers. On the other hand, treatment of cells with NF-kappa B inhibitors (SN50, BAY11-7085, MG132, or pyrrolidine dithiocarbamate) did not inhibit muscarinic receptor-mediated CCE. These findings provide evidence for the involvement of NF-kappa B regulated by CCE in muscarinic receptor-mediated sAPP alpha release enhancement.