Role of calcium in the shape control of human granulocytes.

: The possible role of cytoplasmic calcium in the shape control of human blood granulocytes was explored with two complementary sets of experiments. First, cells were stimulated with N-formyl methionyl leucyl phenylalanine (FMLP) with or without depletion of free intracellular calcium (by incubation with BAPTA/AM in calcium-deprived medium). Control cells displayed a rapid and transient rise of free intracellular calcium (peaked after a few seconds, returned to the basal level after 2-3 minutes), extensive morphological polarization (more than 90% polarized cells after 15 minutes), and reorganization of actin filaments (as assessed by image analysis on cells labeled with a fluorescent phallacidin derivative). Calcium-depleted cells displayed no calcium rise, but both morphological polarization and actin reorganization were indistinguishable from those of controls. In a second set of experiments, individual granulocytes were labeled with fluorescent calcium probes and aspirated in a micropipette on the stage of a confocal laser microscope. About half of the cells displayed a transient calcium rise, which was concomitant with the formation of a protrusion in most cases. However, extensive protrusions were shown by the cells that did not exhibit calcium flashes. Furthermore, if cells were treated with ionomycin, in calcium-containing or calcium-deprived medium, then aspirated, they showed protrusions comparable to those of controls. Finally, when cells were treated with ionomycin during micropipette aspiration, a calcium rise was followed by a moderate increase of the rate of protrusion growth. It is concluded that free calcium alone cannot play a major role in the control of cell shape.