Multiple effects of 1-[β-[3-(4-methoxyphenyl)propoxy]-4-methoxy- phenethyl]-1H-imidazole hydrochloride (SKF 96365) on Ca2+ signaling in MDCK cells

C. R. Jan, C. M. Ho, S. N. Wu, C. J. Tseng

研究成果: Article同行評審

50 引文 斯高帕斯(Scopus)


The effect of 1-[β-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]- 1H-imidazole hydrochloride (SKF 96365) on Ca2+ signaling in Madin Darby canine kidney (MDCK) cells was examined. SKF 96365 at 25-100 μM evoked a robust [Ca2+](i) transient in a dose-dependent manner, measured by fura-2 fluorimetry. A concentration of 10 μM SKF 96365 did not have an effect. The transient consisted of a slow rise, a gradual decay, and a sustained plateau in physiological Ca2+ medium. Removal of extracellular Ca2+ reduced the Ca2+ signals evoked by 50-100 μM SKF 96365 by nearly half in the area under the curve, suggesting that SKF 96365 induced intracellular Ca2+ release and also extracellular Ca2+ influx. A concentration of 100 μM SKF 96365 caused significant Mn2+ quench of fura-2 fluorescence, which was partly inhibited by La3+ (1 mM) or Gd3+ (0.1 mM), indicating that the SKF 96365-induced Ca2+ influx had two components: one is sensitive to La3+ (1 mM) or Gd3+ (0.1 mM), the other is not. The internal Ca2+ source for the SKF 96365-induced [Ca2+](i) transient was the endoplasmic reticulum Ca2+ store because, pretreatment with thapsigargin and cyclopiazonic acid, two inhibitors of the endoplasmic reticulum Ca2+ pump nearly abolished the SKF 96365-induced [Ca2+](i) increase in Ca2+-free medium. In contrast, pretreatment with 100 μM SKF 96365 only partly depleted the thapsigargin- sensitive Ca2+ store. Addition of 10 mM Ca2+ induced a significant [Ca2+](i) increase after prior incubation with 100 μM SKF 96365 in Ca2+- free medium, demonstrating that SKF 96365 induced capacitative Ca2+ entry. This capacitative Ca2+ entry was about 40% of that induced by 1 μM thapsigargin. Additional to inducing its own capacitative Ca2+ entry, 100 μM SKF 96365 partly inhibited thapsigargin- or uridine trisphosphate (UTP)- induced capacitative Ca2+ entry. We also investigated the mechanisms underlying the decay of the SKF 96365-induced [Ca2+](i) transient. Inhibition of the plasma membrane Ca2+ pump with La3+ or Gd3+, or lowering extracellular Na+ level to 0.35 mM, significantly increased the SKF 96365-induced [Ca2+](i) transient. In contrast, the mitochondrial uncoupler carbonylcyanide m-chlorophenylhydrazone had little effect. In Ca2+-free medium, the thapsigargin-induced [Ca2+](i) increase was greatly reduced by pretreatment with SKF 96365. Collectively, we have found that besides its well-known inhibitory action on capacitative Ca2+ entry in many cell types, in MDCK cells SKF 96365 exerted multiple and complex effects on Ca2+ signaling. It induced a considerable increase in [Ca2+](i) by releasing Ca2+ from the endoplasmic reticulum store followed by capacitative Ca2+ entry. It also caused a direct Ca2+ entry. The decay of the SKF 96365 response was significantly governed by efflux via the plasma membrane Ca2+ pump or Na+/Ca2+ exchange. Sequestration by mitochondria or the endoplasmic reticulum played a minor role. We caution use of SKF 96365 as an inhibitor of capacitative Ca2+ entry.

頁(從 - 到)92-101
期刊Naunyn-Schmiedeberg's Archives of Pharmacology
出版狀態Published - 1999

All Science Journal Classification (ASJC) codes

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