Involvement of a P2X7 Receptor in the Acrosome Reaction Induced by ATP in Rat Spermatozoa

Abstract

The acrosome reaction (AR) is the exocytosis of the acrosomal vesicle in response to different physiological and non-physiological stimuli. Particularly in mammals, the AR is needed for sperm to fuse with the oocyte plasma membrane, and it occurs only in capacitated sperm. Previous evidence in the literature indicates that extracellular ATP induces the AR in capacitated human and bovine spermatozoa, but its receptor has not yet been identified. The aim of this work was to define a putative ATP receptor in rat spermatozoa using pharmacological and biochemical approaches. We found that ATP induced the AR only in capacitated rat spermatozoa, which was inhibited in the presence of two general inhibitors of ATP receptors (P2 receptors), Suramin, and oxidized ATP (oATP), and one inhibitor of P2X receptor (pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid [PPADS]). In addition, the AR induced by ATP in capacitated rat spermatozoa was inhibited by brilliant blue-G (BB-G) and 17-β-oestradiol, two blockers of P2X7 receptors. Moreover, the ATP analog 2'(3')-O-(4-benzoylbenzoyl) ATP (BzATP) was almost 500 times more potent than ATP to induce the AR, which agrees with the pharmacology of a P2X7 receptor. Here, we show the presence of P2X7 receptor by Western blot and its localization in the tail and acrosome by indirect immunofluorescence. Finally, we quantify the presence of ATP in the rat oviduct during the estrous cycle. We found that the ATP concentration within the lumen of the oviduct is similar to those required to induce acrosome reaction, which agree with its role during in vivo fertilization. Therefore, our results strongly suggest that ATP induces the AR in capacitated rat spermatozoa through a P2X7 receptor, which may be functional during in vivo fertilization.

Acrosomal alkalization triggers Ca2+ release and acrosome reaction in mammalian spermatozoa

“…At the present time, the IP 3 /Ryanodine receptor families have been detected in this organelle (Bahat & Eisenbach, ; Kuroda, Kaneko, Yoshimura, Nozawa, & Mikoshiba, ; Nguyen, Duittoz, Praud, Combarnous, & Blesbois, ; Treviño et al, ; Walensky & Snyder, ; Zapata et al, ). In addition to these Ca 2+ channels, TRPs (particularly TRPML), TPCs (Arndt et al, ) and P2Xs (Banks, Calvert, & Burnstock, ; Navarro, Miki, & Clapham, ; Torres‐Fuentes, Rios, & Moreno, ) are channels found in acidic Ca 2+ stores that could reside in the acrosome. Several of these channels are pH dependent (Christensen, Myers, & Swanson, ; Peiter, ; Pitt et al, ; Venkatachalam, Wong, & Zhu, ; Xu & Ren, ) and some like TRPs may respond to mechanical stress (Basso & Altier, ; Echeverry, Rodriguez, & Torres, ; Xiao, Chen, & Zhang, ).…”

Mouse sperm begin to undergo acrosomal exocytosis in the upper isthmus of the oviduct

“…It is also possible that exocytosis can be triggered by more than one compound in the middle-to-upper isthmus and lower ampulla. Several molecules were reported to induce acrosome reaction by other groups such as GABA (Shi et al, 1997), Glycine (Bray et al, 2002), atrial natriuretic peptide (Zamir et al, 1995), ATP (Torres-Fuentes et al, 2015), among others. Redundant mechanisms for inducing AE could insure that fertilization is successful.…”

Purinergic Signalling: Therapeutic Developments

“…In rat testes, during spermatogenesis there is differential, stage-dependent immunostaining for P2XR subtypes ( Glass et al, 2001 ), suggesting purinergic targets for both fertility and contraception. ATP triggers the acrosome reaction via P2X7R in human spermatozoa ( Torres-Fuentes et al, 2015 ). Purinergic signalling plays a role in the maturation of sperm cells in the testes.…”