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Thrombin‐dependent calcium signalling in single human erythroleukaemia cells
Author(s) -
Somasundaram Baggi,
Mason Michael J.,
MahautSmith Martyn P.
Publication year - 1997
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.1997.485bm.x
Subject(s) - thapsigargin , thrombin , biophysics , chemistry , intracellular , divalent , extracellular , patch clamp , calcium , fura 2 , microbiology and biotechnology , biochemistry , platelet , biology , cytosol , enzyme , receptor , immunology , organic chemistry
1 A combination of single cell fluorescence and patch clamp techniques were used to study the mechanisms underlying thrombin‐evoked Ca 2+ signals in human erythroleukaemia (HEL) cells, a leukaemic cell line of platelet–megakaryocyte lineage. 2 Thrombin caused a transient increase in intracellular Ca 2+ ([Ca 2+ ] j ), consisting of both release of Ca 2+ from intracellular stores and influx of extracellular Ca 2+ . Mn 2+ quench studies indicated that the thrombin‐evoked divalent cation‐permeable pathway was activated during, but not prior to, release from internal stores. 3 Thapsigargin (1 μ m ) irreversibly released internal Ca 2+ from the same store as that released by thrombin and continuously activated a Ca 2+ ‐influx mechanism. The amplitude of the thrombin‐ and thapsigargin‐induced Ca 2+ influx displayed a marked single cell heterogeneity which showed no correlation with the size of the store Ca 2+ transient. 4 In whole‐cell patch clamp recordings, both thrombin and thapsigargin evoked an inwardly rectifying Ca 2+ current which developed with little or no increase in current noise, showed no reversal in the voltage range −110 to +60 mV and was blocked by 1 m m Zn 2+ . The apparent divalent cation permeability sequence of this pathway was Ca 2+ ≫ Ba 2+ > Mn 2+ , Mg 2+ . The thapsigargin‐evoked current density at −100 mV varied between 0.42 and 2.1 pA pF −1 in different cells. Thrombin failed to activate additional Ca 2+ current if it was added after the thapsigargin‐induced inward current had fully developed. 5 These studies indicate that thrombin activates Ca 2+ influx in HEL cells entirely via a Ca 2+ ‐store‐release‐activated Ca 2+ current ( i erac ) rather than via receptor‐operated or second messenger‐dependent Ca 2+ channels. The level of expression of I erac appears to be a major factor in determining the duration of the thrombin‐evoked [Ca 2+ ] i response and therefore represents a means by which cells can exert control over [Ca 2+ ] i ‐dependent events.