INVESTIGADORES
CESARI Andreina
congresos y reuniones científicas
Título:
Extracellular cAMP activates molecular signalling pathways associated to sperm capacitation in bovines
Autor/es:
ALONSO, CAI; OSYCKA SALUT, C; CASTELLANO, L ; CESARI, A; MORRELL, J; JOHANNISSON, A; DAVIO, C; PEREZ MARTINEZ, S
Lugar:
San Diego
Reunión:
Congreso; 49th Anual Meeting of Society for the Study of Reproduction; 2016
Institución organizadora:
SSR
Resumen:
In order to fertilise the oocyte, ejaculated spermatozoa must undergo a series of physiological changes in the female reproductive tract formerly known as capacitation. Sperm capacitation correlates with a number of membrane and metabolic modifications that include an increased bicarbonate and Ca2+ influx, activation of a soluble adenylyl cyclase (sAC) to produce cAMP, protein kinase A (PKA) activation, protein tyrosine phosphorylation, and the development of hyperactivated motility.Although cAMP concentration is mainly regulated by the modulation of its synthesis by sAC and its degradation by phosphodiesterases, increasing evidence shows that the extrusion of cAMP through multidrug resistance associated protein 4 (MRP4) is also involved in the regulation of this nucleotide levels, mantaining cell homeostasis in physiological and pathophysiological processes.In our laboratory we found that the cAMP efflux by MRP4 occurs during sperm capacitation in bovines and the blockade of the transporter inhibits sperm capacitation. Moreover, the supplementation of sperm incubation media with external non-permeable cAMP abolishes the inhibition and leads to capacitation, suggesting that extracellular cAMP regulates signalling cascades involved to this process.In the present work we aim to elucidate the molecular events that are involved in the sperm capacitation induced by extracellular cAMP in bovines.Cryopreserved spermatozoa were incubated in sp-TALP in the presence of cAMP (10 nM) with or without inhibitors of key molecules involved in signalling cascades related to sperm capacitation (PLC, PKC, AC, PKA). Different events associated to sperm capacitation were studied: CTC patterns and LPC-induced acrosome reaction, Ca2+levels, Ser/Thr and tyrosine phosphorylation, mithocondria membrane potential changes and hyperactivation of the flagellum.Results showed that external cAMP induces sperm capacitation, through the A1 receptor, depending upon the action of its associated PLC. Downstream this enzyme, cAMP increased pERK1-2 levels through PKC activation and elicited a rise of intracellular Ca2+. On the other hand, cAMP-induced capacitation also depends upon the activity of sAC and PKA and increased the tyrosine phosphorylation. In addition, cAMP induced hyperactivation of the motility, and concomitantly increased the number of sperm with high mithocondrial activity.Finally, in vitro fertilisation assays were carried out in the presence of cAMP. Results indicated that the nucleotide increased the number of 2-cell embryos respect to control conditions.All of these findings suggest an important role of external cAMP in the regulation of the signalling pathways involved in sperm capacitation in bovines.