Temporal and spatial resolution of Ca2+ release and influx in human neutrophils using a novel confocal laser scanning mode.

Confocal laser scanning demonstrated that stimulation of neutrophils with the surface receptor agonist, f-met-leu-phe, resulted in the release of stored Ca2+ from a single site. From reconstructions of neutrophils stained with acridine orange, it was shown that the central Ca2+ release site was always close to the nucleus, and correlated with a site which stained with DiOC(6)3. Elevated Ca2+ was locally restricted to within 1 micron of this site. Release of Ca2+ by this pathway was accompanied by the influx of Ca2+, less than 1 second after store release. In each neutrophil, the Ca2+ store release component preceded the Ca2+ influx, and their spatial separation suggested communication between the Ca2+ store and the plasma membrane by a messenger. The (distance)2 of the release site from the plasma membrane was correlated to the time between Ca2+ release and influx, consistent with the diffusion of a messenger from the storage site signalling Ca2+ influx.