Molecular Human Reproduction Vol. 2, NUMBER 4 pp. 225-232, 1996
© European Society of Human Reproduction and Embryology 1996
research-article |
Regulators of sperm function
Single cell analysis of tyrosine kinase dependent and independent Ca2+ fluxes in progesterone induced acrosome reaction
Department of Biochemistry and Molecular Biology, University of Granada Faculty of Sciences, Campus Universitario ‘Fuentenueva’ 18071 Granada, Spain
To whom correspondence should be addressed at: 1To whom correspondence should be addressed
In this study we developed a single cell analysis protocol with which protein tyrosine kinase (PTK)-dependent and independent Ca2+ fluxes occurring in human spermatozoa in response to progesterone were evaluated. By recording the fluorescence emitted by fluo-3-loaded spermatozoa using a confocal laser scanning microscopy system it was possible not only to monitor relative changes in the intracellular free Ca2+ concentration ([Ca2+]i) but also to determine the time at which the acrosomal exocytosis began. The addition of progesterone produced a rapid transient [Ca2+]i increase in 35% of spermatozoa. In 
10% of spermatozoa, this initial [Ca2+]i increase was followed by a secondary [Ca2+]i increase beginning 2–10 min after the progesterone addition and leading to the acrosomal exocytosis in most of these spermatozoa. On the other hand, a rapid triggering of exocytosis during the initial [Ca2+]i increase was a relatively infrequent observation. The inhibition of PTK with genistein or herbimycin A did not influence the initial progesterone-induced [Ca2+]i increase but inhibited the secondary [Ca2+]i increase and the ensuing acrosomal exocytosis. The initial PTK-independent Ca2+ response could be induced by progesterone in both non-capacitated and capacitated spermatozoa, whereas the ability to generate the secondary, PTK-dependent response developed during in-vitro capacitation.
acrosome reaction/Ca2+ fluxes/non-genomic steroid effects/progesterone/protein tyrosine kinase
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