1: J Cell Biol  1995 Nov;131(4):963-73 

Regulatory mechanisms of the acrosome reaction revealed by multiview microscopy
of single starfish sperm.

Sase I, Okinaga T, Hoshi M, Feigenson GW, Kinosita K Jr.

Department of Physics, Faculty of Science and Technology, Keio University,
Yokohama, Japan.

The acrosome reaction in many animals is a coupled reaction involving an
exocytotic step and a dramatic change in cell shape. It has been proposed that
these morphological changes are regulated by intracellular ions such as Ca2+ and
H+. We report here simultaneous visualization, under a multiview microscope, of
intracellular free Ca2+ concentration ([Ca2+]i), intracellular pH (pHi), and
morphological changes in a single starfish sperm (Asterina pectinifera). [Ca2+]i
and pHi were monitored with the fluorescent probes indo-1 and SNARF-1,
respectively. The acrosome reaction was induced with ionomycin. After the
introduction of ionomycin in the medium, [Ca2+]i increased gradually and reached
a plateau in approximately 30 s. The fusion of the acrosomal vacuole took place
abruptly before the plateau, during the rising phase. Although the speed of the
[Ca2+]i increase varied among the many sperm tested, exocytosis in all cases
occurred at the same [Ca2+]i of approximately 2 microM (estimated using the
dissociation constant of indo-1 for Ca2+ of 1.1 microM). This result suggests
that the exocytotic mechanism in starfish sperm responds to [Ca2+]i rapidly,
with a reaction time of the order of one second or less. Unlike the change in
[Ca2+]i, an abrupt increase in pHi was observed immediately after exocytosis,
suggesting the presence of a proton mobilizing system that is triggered by
exocytosis. The rapid increase in pHi coincided with the formation of the
acrosomal rod and the beginning of vigorous movement of the flagellum, both of
which have been proposed to be pHi dependent. The exocytotic event itself was
visualized with the fluorescent membrane probe RH292. The membrane of the
acrosomal vacuole, concealed from the external medium in an unreacted sperm, was
seen to fuse with the plasma membrane.

PMID: 7490297 [PubMed - indexed for MEDLINE]