Effect of p-chloroamphetamine on calcium movement and viability in renal tubular cells

In Madin-Darby canine kidney (MDCK) cells, the effect of p-chloroamphetamine, a neurotoxin that depletes intracellular serotonin, on intracellular Ca²⁺ concentration ([Ca²⁺]_i) and viability was measured by using the Ca²⁺-sensitive fluorescent dye fura-2 and the viability detecting fluorescent dye tetrazolium. p-Chloroamphetamine (<10 μM) caused a rapid rise of [Ca²⁺] _i in a concentration-dependent manner. p-Chloroamphetamine-induced [Ca²⁺]_i rise was partly reduced by removal of extracellular Ca²⁺. p-Chloroamphetamine-induced extracellular Ca ²⁺ influx was also suggested by Mn²⁺ influx-induced fura-2 fluorescence quench. In Ca²⁺-free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca²⁺-ATPase, caused a monophasic [Ca²⁺]_i rise, after which p-chloroamphetamine failed to increase [Ca²⁺]_i; also, pretreatment with p-chloroamphetamine reduced 50% of thapsigargin-sensitive Ca²⁺ stores. U73122, an inhibitor of phospholipase C, abolished ATP (but not p-chloroamphetamine)-induced [Ca²⁺]_i rise. Overnight incubation with 1-500 μM p-chloroamphetamine decreased cell viability. These findings suggest that p-chloroamphetamine evokes a rapid increase in [Ca ²⁺]_i in renal tubular cells by stimulating both extracellular Ca²⁺ influx and intracellular Ca²⁺ release, and is cytotoxic.