title: Inhibitors of receptor tyrosine kinases do not suppress progesterone-induced [Ca2+]i signalling in human spermatozoa.
abstract: Previous studies have implicated receptor tyrosine kinases in progesterone-induced [Ca2+]i signalling, and consequent induction of the acrosome reaction, in human spermatozoa. We have investigated the effects of tyrosine kinase inhibition on [Ca2+]i responses in large numbers of individual human spermatozoa. Genistein (5, 50 and 250 micromol/l), an inhibitor of receptor-linked tyrosine kinases, significantly inhibited the progesterone-induced acrosome reaction (P < 0.05). However, we could detect no effect of genistein on progesterone-induced [Ca2+]i signalling. In control experiments, application of progesterone induced a significant transient [Ca2+]i response in approximately 77% of cells and a sustained [Ca2+]i ramp/plateau in approximately 48% of cells (n = 26; 5411 cells). In preparations pretreated with genistein (50 micromol/l), significant transient and sustained responses were detected in 69.5 and 39.1% of cells respectively (n = 5; 1109 cells). The amplitudes of both transient and sustained [Ca2+]i responses were similar in control and genistein-pretreated preparations. Tyrphostin A47 (100 micromol/l), another receptor tyrosine kinase inhibitor, also failed to inhibit either the transient or sustained [Ca2+]i response (n = 3; 468 cells). Assessment of tyrosine phosphorylation of two sperm proteins (p105/81) showed greatly increased levels of phosphotyrosine in response to capacitation but a negligible increase in response to progesterone stimulation. Pretreatment with genistein (50 and 250 micromol/l) decreased capacitation-induced tyrosine phosphorylation and resulted in a loss of phosphorylation in response to progesterone treatment. We conclude that neither the transient nor sustained phases of the progesterone-induced [Ca2+]i response require receptor tyrosine kinase signalling. Previous reports of modulation of the progesterone-induced [Ca2+]i signal by tyrosine kinase inhibition probably reflect inhibition of the acrosome reaction.
authors.affiliation: School of Biosciences, University of Birmingham, B15 2TT, UK. j.kirkmanbrown@bham.ac.uk
authors.forename: J C
authors.initials: JC
authors.lastname: Kirkman-Brown
authors.forename: L
authors.initials: L
authors.lastname: Lefièvre
authors.forename: C
authors.initials: C
authors.lastname: Bray
authors.forename: P M
authors.initials: PM
authors.lastname: Stewart
authors.forename: C L R
authors.initials: CL
authors.lastname: Barratt
authors.forename: S J
authors.initials: SJ
authors.lastname: Publicover
mesh_terms: D000177:Acrosome / Q000201:enzymology / Q000378:metabolism; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D008297:Male; D011374:Progesterone / Q000378:metabolism / Q000494:pharmacology*; D020794:Receptor Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D015398:Signal Transduction / Q000502:physiology; D013094:Spermatozoa / Q000201:enzymology / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism; D020032:Tyrphostins / Q000494:pharmacology
chemical_list: D004791:Enzyme Inhibitors; D020032:Tyrphostins; C060152:tyrphostin 47; D014443:Tyrosine; D011374:Progesterone; D019833:Genistein; D020794:Receptor Protein-Tyrosine Kinases; D002118:Calcium