Effect of nortriptyline on intracellular Ca²⁺ handling and proliferation in human osteosarcoma cells

The effect of the antidepressant nortriptyline, on bone cells is unknown. In human osteosarcoma MG63 cells, the effect of nortriptyline on intracellular Ca²⁺ concentration ([Ca²⁺]_i) and proliferation was measured by using fura-2 and tetrazolium, respectively. Nortriptyline (≥10 μM) caused a [Ca²⁺]_i rise in a concentration-dependent manner (EC₅₀ = 200 μM). Nortriptyline-induced [Ca²⁺]_i rise was prevented by 60% by removal of extracellular Ca²⁺ but was not altered by voltage-gated Ca²⁺ channel blockers. In Ca²⁺-free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca²⁺-ATPaSe, caused a monophasic [Ca²⁺]_i rise, after which the increasing effect of nortriptyline on [Ca²⁺]_i was abolished; also, pretreatment with nortriptyline abolished thapsigargin-induced [Ca ²⁺]_i increase. U73122, an inhibitor of phospholipase C, did not affect nortriptyline-induced [Ca²⁺]_i rise; however, activation of protein kinase C decrease nortriptyline-induced [Ca ²⁺]_i rise by 32%. Overnight incubation with 50 and 100 μM nortriptyline killed 78% and 97% of cells, respectively; while 10 μM nortriptyline had no effect. These data suggest that nortriptyline rapidly increases [Ca²⁺]_i in human osteosarcoma cells by stimulating both extracellular Ca²⁺ influx and intracellular Ca ²⁺ release, and is cytotoxic at high concentrations.