Amitriptyline modulated Ca²⁺ signaling and induced Ca²⁺-independent cell viability in human osteosarcoma cells

Abstract

Amitriptyline is a widely used tricyclic antidepressant, which acts primarily as a serotonin-norepinephrine reuptake inhibitor. This study examined the effect of amitriptyline on Ca homeostasis and its related mechanism in MG63 human osteosarcoma cells. Amitriptyline evoked cytosolic-free Ca concentrations ([Ca]) rises concentration dependently. Amitriptyline-evoked Ca entry was confirmed by Mn-induced quench of fura-2 fluorescence. This entry was inhibited by Ca entry modulators nifedipine, econazole, SKF96365, the protein kinase C (PKC) activator phorbol 12-myristate 13 acetate but was not affected by the PKC inhibitor GF109203X. In Ca-free medium, treatment with the endoplasmic reticulum Ca pump inhibitor thapsigargin (TG) inhibited amitriptyline-evoked [Ca] rises by 95%. Conversely, treatment with amitriptyline abolished TG-evoked [Ca] rises. Inhibition of phospholipase C (PLC) with U73122 inhibited amitriptyline-evoked [Ca] rises by 70%. Amitriptyline killed cells at 200-500 μM in a concentration-dependent fashion. Chelating cytosolic Ca with 1,2-bis(2-aminophenoxy)ethane- N, N, N', N'-tetraacetic acid/AM did not reverse amitriptyline-induced cytotoxicity. Collectively, our data suggest that in MG63 cells, amitriptyline induced [Ca] rises by evoking PLC-dependent Ca release from the endoplasmic reticulum and Ca entry via PKC-regulated store-operated Ca entry. Amitriptyline also induced Ca-disassociated cell death.

Action of the insecticide cyfluthrin on Ca2+ signal transduction and cytotoxicity in human osteosarcoma cells

“…In MG63 cells, store-operated Ca 2þ channels have been shown to play a crucial role in [Ca 2þ ] i rises triggered by different compounds such as niflumic acid, amitriptyline, and methoxsalen. [17][18][19] Although there are so far no selective inhibitors for this Ca 2þ entry, nifedipine, econazole, and SKF96365 were commonly applied for this purpose. [25][26][27] Our results suggest that all of these three chemicals inhibited cyfluthrin-induced [Ca 2þ ] i rises.…”

“…In MG63 cells, the endoplasmic reticulum was thought to play a key role in releasing store-operated Ca 2þ . [17][18][19] To exclude the contribution of Ca 2þ entry, the experiments were performed in Ca 2þ -free medium. Figure 4(a) shows that addition of 1 mM thapsigargin (TG), an endoplasmic reticulum Ca 2þ pump inhibitor, 22…”

“…Since previous researches have shown that in MG63 cells, 16 [Ca 2þ ] i rises can be evoked in response to the stimulation of external compounds such as niflumic acid, 17 amitriptyline, 18 and methoxsalen 19 . The goal of this study was to investigate the underlying pathways of this Ca 2þ movement.…”