Effect of Riluzole on Ca2+ Movement and Cytotoxicity in Madin-Darby Canine Kidney Cells

Abstract

Riluzole is a drug used in the treatment of amyotrophic lateral sclerosis; however, its in vitro action is unclear. In this study, the effect of riluzole on intracellular Ca^{2 -} concentration ([Ca^{2 -}]_i) in Madin-Darby canine kidney (MDCK) cells was investigated using the Ca^{2 -}-sensitive fluorescent dye, fura-2. Riluzole (100 -500 mM) caused a rapid and sustained increase of [Ca^{2 -}]_i in a concentration-dependent manner (EC₅₀ = 150 mM). Some 40 and 50% of this [Ca^{2 -}]_i increase was prevented by the removal of extracellular Ca^{2 -} and the addition of La^{3 -}, respectively, but was unchanged by dihydropyridines, verapamil and diltiazem. In Ca^{2 -}-free medium, thapsigargin -an inhibitor of the endoplasmic reticulum (ER) Ca^{2 -}-ATPase -caused a monophasic [Ca2-]_i increase, after which the increasing effect of riluzole on [Ca^{2 -}]_iwas attenuated by 70%; in addition, pre-treatment with riluzole abolished thapsigargin-induced [Ca^{2 -}]_i increases. U73122, an inhibitor of phospholipase C (PLC), abolished ATP (but not riluzole)-induced [Ca^{2 -}]_i increases. At concentrations of 250 and 500 mM, riluzole killed 40 and 95% cells, respectively. The cytotoxic effect of riluzole (250 mM) was unaltered by pre-chelating cytosolic Ca^{2 -} with BAPTA. Collectively, in MDCK cells, riluzole rapidly increased [Ca^{2 -}]_i by stimulating extra-cellular Ca^{2 -} influx via an La^{3 -}-sensitive pathway and intracellular Ca^{2 -} release from the ER via, as yet, unidentified mechanisms. Furthermore, riluzole caused Ca^{2 -}-unrelated cytotoxicity in a concentration-depen-dent manner.