Effect of capsazepine on [Ca²⁺]_i in MDCK renal tubular cells

The current study explored whether capsazepine changed basal cytosolic free Ca²⁺ concentrations ([Ca²⁺]_i) levels in suspended Madin Darby canine kidney (MDCK) cells cells by using fura-2 as a Ca²⁺-selective fluorescent dye. At concentrations of 10-200 μM, capsazepine increased [Ca²⁺]_i in a concentration-dependent manner. The Ca²⁺ signal was partially reduced by 40% by removing extracellular Ca²⁺. Capsazepine induced Mn²⁺ quench of fura-2 fluorescence, indirectly implicating Ca²⁺ entry. Capsazepine-induced Ca²⁺ influx was unchanged by L-type Ca ²⁺ entry inhibitors and protein kinase C modulators [phorbol 12-myristate 13-acetate (PMA) and GF109203X]. In Ca²⁺-free medium, 100 μM capsazepine-induced Ca²⁺ release was substantially suppressed by pretreatment with thapsigargin (an endoplasmic reticulum Ca ²⁺ pump inhibitor). Pretreatment with capsazepine nearly abolished thapsigargin-induced Ca²⁺ release. Inhibition of phospholipase C with U73122 did not change capsazepine-induced [Ca²⁺]_i rises. Collectively, in MDCK cells, capsazepine induced [Ca²⁺]_i rises by causing phospholipase C-independent Ca²⁺ release from the endoplasmic reticulum and Ca²⁺ influx via non-L-type Ca²⁺ channels.