Maintenance of intracellular free Ca2+ homeostasis following lethal heat shock

We have manipulated the extracellular Ca2+ concentration (1.8 mM in normal Dulbecco's modified Eagle's medium) to test whether the resulting effect on intracellular free Ca2+ homeostasis was similar in heat-sterilized and nonheated mouse NIH-3T3 cells. The responsiveness of the intracellular free Ca2+ concentration to changes in the extracellular Ca2+ concentration was not affected by prior treatment of the cells with trypsin, or by the extracellular Ca2+ concentration during dye loading (indo-1, AM). Rather, the intracellular free Ca2+ concentration was dependent upon the ambient Ca2+ concentration during analysis by flow cytometry. When the extracellular Ca2+ concentration was decreased to 0.017 mM, either before or after a lethal heat shock, the intracellular free Ca2+ concentration (approximately 300 nM) decreased to a similar extent in both heated and control cells (to approximately 30-100 nM). Similarly, when the extracellular Ca2+ concentration was increased to 15.0 mM, either before or after a lethal heat shock, the intracellular free Ca2+ concentration exhibited a quantitatively similar increase in both heated and nonheated cells (to approximately 400-1000 nM). These data indicate that a lethal heat dose does not inhibit the intact cell's ability to maintain intracellular free Ca2+ homeostasis.