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In neutrophils, intracellular Ca 2+ levels are regulated by several transporters and pathways, namely SERCA [sarco(endo)plasmic reticulum Ca 2+ -ATPase], SOCE (store-operated calcium entry), and ROCE (receptor-operated calcium entry). However, the exact mechanisms involved in the communication among these transporters are still unclear. In the present study, thapsigargin, an irreversible inhibitor of SERCA, and ML-9, a broadly used SOCE inhibitor, were applied in human neutrophils to better understand their effects on Ca 2+ pathways in these important cells of the immune system. The thapsigargin and ML-9 effects in the intracellular free Ca 2+ flux were evaluated in freshly isolated human neutrophils, using a microplate reader for monitoring fluorimetric kinetic readings. The obtained results corroborate the general thapsigargin-induced intracellular pattern of Ca 2+ fluctuation, but it was also observed a much more extended effect in time and a clear sustained increase of Ca 2+ levels due to its influx by SOCE. Moreover, it was obvious that ML-9 enhanced the thapsigargin-induced emptying of the internal stores. Indeed, ML-9 does not have this effect by itself, which indicates that, in neutrophils, thapsigargin does not act only on the influx by SOCE, but also by other Ca 2+ pathways, that, in the future, should be further explored.

Calcium Pathways in Human Neutrophils—The Extended Effects of Thapsigargin and ML-9