Pharmacological Evidence for the Involvement of Calciuml Calmodulin in Serotonin 5-HT3 Receptor-Mediated Cation Permeability in NG 108-15 Cells

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Abstract
In NG 108–15 clonal cells, extracellular application of micromolar concentrations of serotonin [5-hydroxy-tryptamine (5-HT)] and substance P induces the opening of a cation permeability monitored by the influx of [14C]-guanidinium. The serotoninergic component of this cation permeability Is linked to 5-HT3 receptor activation, whereas the substance P component probably involves an “N-terminal-dependent substance P receptor.” In this study, [14C]guanidinium influx triggered by 1 μM 5-HT plus 10 μM substance P was shown to be insensitive to tetrodotoxin, verapamil, diltiazem, nimodipine, and ω-conotoxin, as expected from a process independent of voltage-sensitive sodium and calcium channels. In contrast, [14C]guanidinium influx was inhibited by millimolar concentrations of extracellular calcium and by the chelation of intracellular calcium by bis-O-aminophenoxyethanetetraacetic acid. The inhibition by extracellular calcium apparently involved a competition between the divalent cation and [14C]guanidinium for the same channel. When NG 108–15 cells were exposed to X537A, an ionophore that specifically induces release of calcium from intracellular stores, [14C]guanidinium uptake was markedly increased even in the absence of 5-HT and/or substance P. Conversely, [14C]guanidinium influx due to the latter substances could be reversibly and dose-dependently blocked by various drugs that possess calmodulin-antagonizing properties. These results strongly suggest that the cation permeability opened by 5-HT and substance P in NG 108–15 cells involves a calcium/calmodulin-dependent process. However, as the phosphodiesterase inhibitor isobutylmethylxanthine, the nitric oxide synthase inhibitor A/monomethylarginine, the protein kinase C inhibitor staurosporine, and the protein kinase C activator 12-O-tetradeca-noylphorbol 13-acetate did not alter [14C]guanidinium influx in NG 108–15 cells exposed to 5-HT and substance P., this process probably does not involve the calcium-dependent nitric oxide pathway and protein kinase C activation.