A novel action of the antianginal drug bepredil: induction of internal Ca(2+) release and external Ca(2+) influx in Madin-Darby canine kidney (MDCK) epithelial cells

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