Cell cycle-dependent calcium oscillations in mouse embryonic stem cells

During cell cycle progression, somatic cells exhibit different patterns of intracellular Ca2+ signals during the G(0) phase, the transition from G(1) to S, and from G(2) to M. Because pluripotent embryonic stem ( ES) cells progress through cell cycle without the gap phases G1 and G2, we aimed to determine whether mouse ES ( mES) cells still exhibit characteristic changes of intracellular Ca2+ concentration during cell cycle progression. With confocal imaging of the Ca2+-sensitive dye fluo-4 AM, we identified that undifferentiated mES cells exhibit spontaneous Ca2+ oscillations. In control cultures where 50.4% of the cells reside in the S phase of the cell cycle, oscillations appeared in 36% of the cells within a colony. Oscillations were not initiated by Ca2+ influx but depended on inositol 1,4,5- trisphosphate ( IP3)- mediated Ca2+ release and the refilling of intracellular stores by a store- operated Ca2+ influx ( SOC) mechanism. Using cell cycle synchronization, we determined that Ca2+ oscillations were confined to the G(1)/S phase ( similar to 70% oscillating cells vs. G(2)/M with similar to 15% oscillating cells) of the cell cycle. ATP induced Ca2+ oscillations, and activation of SOC could be induced in G(1)/S and G(2)/M synchronized cells. Intracellular Ca2+ stores were not depleted, and all three IP3 receptor isoforms were present throughout the cell cycle. Cell cycle analysis after EGTA, BAPTA- AM, 2-aminoethoxydiphenyl borate, thapsigargin, or U-73122 treatment emphasized that IP3-mediated Ca2+ release is necessary for cell cycle progression through G(1)/S. Because the IP3 receptor sensitizer thimerosal induced Ca2+ oscillations only in G1/S, we propose that changes in IP3 receptor sensitivity or basal levels of IP3 could be the basis for the G(1)/S-confined Ca2+ oscillations.