Signalling pathways associated with the bicarbonate-dependent rat sperm capacitation process

DA ROS, VG; COHEN, DJ; PIGNATARO, O; VISCONTI, PE; CUASNICÚ, PS

Buenos Aires, Argentina

Congreso; VIII Jornadas Multidisciplinarias de la Sociedad Argentina de Biología (SAB); 2006

Sociedad Argentina de Biología (SAB)

Results from our group demonstrated that bicarbonate is required to support rat sperm capacitation by regulating protein tyrosine phosphorylation, migration of epididymal protein DE/CRISP-1 to the equatorial segment and expression of sperm fusion ability. However, while phosphorylation occurred even when bicarbonate was replaced by a cAMP analogue, the other two parameters seem to require additional bicarbonate properties. To further study the pathway involve in the bicarbonate regulation of rat sperm capacitation, intracellular cAMP levels were measured by RIA in sperm incubated in the presence or absence of the anion. Results showed a time-dependent increase in cAMP levels only in sperm incubated in the presence of bicarbonate. To investigate whether this bicarbonate-dependent cAMP accumulation was induced by phorbol esters, sperm were incubated in the presence or absence of bicarbonate, with or without 10 µM PMA. Results showed that PMA produces a significant increase in sperm cAMP levels only in the presence of bicarbonate. In order to test whether the cAMP produced was required for the three capacitation parameters previously analyzed, the involvement of PKA was studied using the PKA inhibitor H89. Tyrosine phosphorylation was significant reduced by addition of 30 µM H89. However, migration of DE/CRISP-1 was not affected, and the expression of the fusion ability could only be partially inhibited by H89, even in the presence of the inhibitor during gamete co-incubation. Taken together, our results provide further information on the regulation of rat sperm capacitation by bicarbonate, confirming that some capacitation-associated events, although bicarbonate-dependent, would be cAMP/PKA-independent.