High micromolar Ca2+ beneath the plasma membrane in stimulated neutrophils

Ca2+ near the inner face of the plasma membrane, as reported by the membrane associated fluorescent Ca2+ probe FFP-18, was higher than the bulk cytosolic free Ca2+ concentration both in resting neutrophils and in response to f-met-leu-phe. Influx caused Ca2+ close to the plasma membrane to rise more rapidly than the bulk cytosolic free Ca2+ and to reach a peak concentration of at least 30 microM. This zone of high Ca2+ was localised to just beneath the plasma membrane and did not extend more than 0.1 micron into the cell, as it was undetected by the bulk cytosolic free Ca2+ probes magfura2 and fura2. From these data, reconstruction of the distribution of Ca2+ within the neutrophil showed that the high Ca2+ signal at the cell cortex rapidly subsided to give a uniform free Ca2+ across the cell.