A novel action of the antianginal drug bepredil: Induction of internal Ca²⁺ release and external Ca²⁺ influx in Madin-Darby canine kidney (MDCK) epithelial cells

The effect of the antianginal drug bepridil on Ca²⁺ signaling in Madin-Darby canine kidney (MDCK) cells was investigated by using fura-2 as a Ca²⁺ probe. Bepridil at 10-50 μM evoked a significant rise in cytosolic free Ca²⁺ concentration ([Ca²⁺](i)) in a dose-dependent manner. The [Ca²⁺](i) rise consisted of an immediate initial rise and a slow decay. Removal of external Ca²⁺ partly inhibited the Ca²⁺ signals by reducing both the initial rise and the decay phase, suggesting that bepridil activated both external Ca²⁺ influx and internal Ca²⁺ release. In Ca²⁺-free medium, pretreatment with 50 μM bepridil nearly abolished the Ca²⁺ release induced by thapsigargin (1 μM), an endoplasmic reticulum Ca²⁺ pump inhibitor, and vice versa, pretreatment with thapsigargin inhibited most of the bepridil-induced Ca²⁺ release, suggesting that the thapsigargin-sensitive Ca²⁺ store was the main source of bepridil-induced Ca²⁺ release. Bepridil (50 μM) induced considerable Mn²⁺ quench of fura-2 fluorescence at an excitation wavelength of 360 nm, which was partly inhibited by La³⁺ (0.1 mM). Consistently, La³⁺ (0.1 mM) pretreatment significantly inhibited the bepridil-induced [Ca²⁺](i) rise. Addition of 3 mM Ca²⁺ induced a significant [Ca²⁺](i) rise after prior incubation with 10-50 μM bepridil in Ca²⁺-free medium, suggesting that bepridil induced dose-dependent capacitative Ca²⁺ entry. However, 50 μM bepridil inhibited 1 μM thapsigargin-induced capacitative Ca²⁺ entry by 38%. Pretreatment with aristolochic acid (40 μM) so as to inhibit phospholipase A₂ inhibited 50 μM bepridil-induced internal Ca²⁺ release by 42%, but inhibition of phospholipase C with U73122 (2 μM) or inhibition of phospholipase D with propranolol (0.1 mM) had little effect, suggesting that bepridil induced internal Ca²⁺ release in an inositol 1,4,5-trisphosphate-independent manner that could be modulated by phospholipase A₂-coupled events. This is the first report providing evidence that bepridil, currently used as an antianginal drug, induced a rise in [Ca²⁺](i) in a non-excitable cell line. Copyright (C) 2000 Elsevier Science Inc.