Extracellular adenosine triphosphate stimulates acrosomal exocytosis in bovine spermatozoa via P2 purinoceptor.
The presence of ATP in the genital tract fluid of mammals provokes questions regarding its function in the fertilization process. We investigated the effect of extracellular ATP (ATPe) on the activation of bovine spermatozoa. A signal transduction mechanism for ATP involving the receptor-mediated release of second messengers is described. Treatment of spermatozoa with ATP, uridine triphosphate (UTP), or 2-methylthio-ATP resulted in a concentration-dependent increase of acrosomal exocytosis, whereas treatment with either AMP or adenosine induced little exocytosis. This suggested that the receptor involved is of the P2 and not the P1 type. Several lines of evidence also suggest that the ATP purinoceptor is of the P2y and not the P2x type. First, the acrosome reaction was induced by the P2y-agonists ATP, UTP, or 2-methylthio-ATP, but no effects were shown by the P2x-agonists alpha,beta-methylene-ATP or beta,gamma-methylene-ATP. Second, ATP-induced acrosomal exocytosis was inhibited by the P2y antagonists, but not by the P2x antagonists. Third, enhanced Ca2+ uptake into the cells was observed with ATP and 2-methylthio-ATP, but not with beta,gamma-methylene-ATP. Additionally, ATP induced elevation of intracellular Ca2+ and cAMP, and the effect on cAMP was predominantly enhanced by including Ca2+ and the Ca2+-ionophore A23187 in the incubation medium. Extracellular ATP also activates protein kinase Calpha (PKCalpha), and the acrosome reaction, stimulated by ATPe, is inhibited by a PKC-specific inhibitor. In summary, we suggest that ATPe activates the P2 purinoceptor that elevates [Ca2+]i, which leads to PKCalpha activation and culminates in acrosomal exocytosis.
Luria
Ayala
A
Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan 52900, Israel.
Rubinstein
Sara
S
Lax
Yehudit
Y
Breitbart
Haim
H
D000177:Acrosome / Q000187:drug effects*; D020101:Acrosome Reaction / Q000187:drug effects*; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D000242:Cyclic AMP / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005089:Exocytosis / Q000187:drug effects*; D015151:Immunoblotting; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D011493:Protein Kinase C / Q000378:metabolism; D051571:Protein Kinase C-alpha; D018048:Receptors, Purinergic P2 / Q000187:drug effects*; D013075:Sperm Capacitation / Q000187:drug effects; D013094:Spermatozoa / Q000187:drug effects*; D013268:Stimulation, Chemical; D013347:Subcellular Fractions / Q000187:drug effects
D007527:Isoenzymes; D018048:Receptors, Purinergic P2; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D002118:Calcium