Hwm. Niessen et al., IMMUNOELECTRON MICROSCOPY REVEALS SIGNIFICANT GRANULE FUSION UPON STIMULATION OF ELECTROPERMEABILIZED HUMAN NEUTROPHILS, Cellular signalling, 6(1), 1994, pp. 47-58
Although electropermeabilization has become an important tool for stud
ying the signal requirements of exocytosis, relatively little is known
about the morphological changes accompanying this response in electro
permeabilized cells. In this study, we determined that electropermeabi
lization of human neutrophils by itself caused only minor changes in t
he morphology as determined by transmission electron microscopy. The s
tructure of the plasma membrane did not show detectable changes, where
as the cytoplasm was more electron lucent as compared to intact cells.
Activation of intact neutrophils with formyl-methionyl-leucyl-phenyla
lanine (FMLP), in the presence of cytochalasin B, caused the developme
nt of invaginations of the plasma membrane. In contrast, activation of
electropermeabilized cells with 1 mu M Ca2+ and/or 50 mu M GTP-gamma-
S caused the development of vacuoles that did not seem to be in contac
t (or had previously been in contact) with the extracellular environme
nt. However, fusion of azurophilic and specific granules with these va
cuoles clearly had taken place. The response characteristics of this f
usion induced by Ca2+ and GTP-gamma-S were quite similar to those of t
he direct fusion of granules with the plasma membrane. We conclude tha
t in electropermeabilized human neutrophils, two processes involving g
ranule fusion can be distinguished. First, a direct fusion of granules
with the plasma membrane. Secondly, the fusion of granules leading to
the formation of vacuoles, not in contact with the extracellular spac
e.