Kinetics of empty store-activated Ca2+ influx in HeLa cells

L. Missiaen, H. De Smedt, J. B. Pary, M. Oike, R. Casteels

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Abstract

The intracellular Ca2+ indicator Indo-1 was used to monitor changes in cytosolic [Ca2+] ([Ca2+](i)) in single HeLa cells upon readmission of external Ca2+ after a short incubation in Ca2+-free solution. HeLa cells were responsive to histamine but not to caffeine, and their histamine- sensitive store was totally depleted by a 60-min exposure to 2 μM thapsigargin. The resting [Ca2+](i) in thapsigargin-treated cells was higher than in control cells and low amplitude [Ca2+](i) oscillations were observed in about 20% of the cells. Readmission of external Ca2+ after a brief withdrawal of extracellular Ca2+ resulted in a transient [Ca2+](i) rise, which then decayed to the same elevated [Ca2+](i) measured before the Ca2+ withdrawal period. The [Ca2+](i) rise was associated with an increased rate of Mn2+ entry, measured as the rate of quenching of intracellular Fura-2. The same procedure did not affect the [Ca2+](i) in control cells not pretreated with thapsigargin. The amplitude of this [Ca2+](i) transient in thapsigargin pretreated cells depended on the duration of prior incubation in Ca2+-free medium. The [Ca2+](i) rise induced by elevating the extracellular [Ca2+] from 1.5 to 10 mM was more pronounced if the [Ca2+](i) during the initial incubation in 1.5 mM Ca2+ was first lowered by depolarizing the cells. We conclude that an empty store stimulates a Ca2+ entry pathway consisting of two components: a continuously elevated basal leak and a second component that is transient due to the high [Ca2+](i)-induced inhibition of the Ca2+ entry pathway. This inhibition and the subsequent recovery from it as the [Ca2+](i) is brought to resting levels could cause the oscillatory Ca2+ entry that we recorded in a fraction of the thapsigargin-treated cells.

Original languageEnglish
Pages (from-to)5817-5823
Number of pages7
JournalJournal of Biological Chemistry
Volume269
Issue number8
Publication statusPublished - Jan 1 1994
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Missiaen, L., De Smedt, H., Pary, J. B., Oike, M., & Casteels, R. (1994). Kinetics of empty store-activated Ca2+ influx in HeLa cells. Journal of Biological Chemistry, 269(8), 5817-5823.