Effects of diadenosine polyphosphates and seminal fluid vesicles on rabbit sperm cells

Membrane vesicles were isolated from rabbit seminal plasma. Electron microscopy analyses showed the presence of numerous small, round vesicles with a diameter of about 70 nm. Determination of enzyme activities was carried out by high performance liquid chromatography and showed that the vesicles can degrade the diadenosine polyphosphates (ApnA), Ap3A and Ap4A and ATP and ADP, but not AMP. Studies of the degradation of diadenosine compounds by the vesicles present in seminal fluid showed an increasing production of AMP as the by-product and a time-dependent generation of dephosphorylated products consistent with the presence of ecto-ATP diphosphophosphatase (ecto-apyrase). In the presence of rabbit spermatozoa, AMP did not accumulate because 5ʹnucleotidase and adenosine deaminase, present at the surface of sperm cells, transformed AMP into adenosine and inosine. The effects of seminal fluid vesicles and diadenosine compounds on the acquisition of fertilizing capacity by rabbit spermatozoa were evaluated by Pisum sativum agglutinin fluorescein isothiocyanate conjugated staining. The results obtained with uncapacitated spermatozoa showed that the capacitating effector BSA could be substituted efficiently by the addition of diadenosine compounds and vesicles previously incubated for 2 h to the capacitative medium. Under these experimental conditions, the spontaneous acrosome reaction rate was not increased. Capacitated rabbit spermatozoa did not undergo acrosome reaction when l-alpha-lysophosphatidylcholine was substituted by diadenosine compounds previously incubated with vesicles. In conclusion, this study has shown that rabbit seminal fluid vesicles can degrade diadenosine compounds to AMP and that the addition of the vesicles and diadenosine compounds to uncapacitated rabbit spermatozoa favours the acquisition of the fertilizing capacity.