Glucocorticoids activate a suicide process in thymocytes through an elevation of cytosolic Ca²⁺ concentration

Glucocorticoid hormones kill immature thymocytes through the induction of a suicide process commonly referred to as "apoptosis." A characteristic marker for this process is the stimulation of endogenous endonuclease activity which results in the extensive cleavage of cell chromatin. In an attempt to characterize the biochemical events involved in this process, we studied the role of Ca²⁺ in glucocorticoid-induced DNA fragmentation and cell killing in thymocytes. Treatment of thymocytes from immature rats with the synthetic glucocorticoid methylprednisolone resulted in extensive DNA fragmentation which was preceded by an early, sustained increase in cytosolic Ca²⁺ concentration. This increase in Ca²⁺ level was blocked by cycloheximide and actinomycin D, inhibitors of de novo protein and mRNA synthesis, respectively. Prevention of the Ca²⁺ increase by buffering cytosolic Ca²⁺ with quin-2, or through incubation of the thymocytes in a "Ca²⁺-free" medium, prevented endonuclease activation and cell killing. Inhibitors of calmodulin also prevented DNA fragmentation without inhibiting the glucocorticoid-stimulated elevation of cytosolic Ca²⁺ concentration. The Ca²⁺ increase appeared to be due to the action of a heat-labile cytosolic factor, synthesized in response to glucocorticoids, which facilitated the influx of extracellular Ca²⁺. Our findings suggest that glucocorticoids induce thymocyte suicide through an elevation of cytosolic Ca²⁺ concentration resulting in endonuclease activation, DNA fragmentation, and cell death.