Int. J. Dev. Biol. 63: 589 - 595 (2019)

https://doi.org/10.1387/ijdb.190092aw

Vol 63, Issue 11-12

Acrosome reaction-inducing substance triggers two different pathways of sperm intracellular signaling in newt fertilization

Original Article | Published: 27 February 2020

Shinnosuke Kon¹, Akio Takaku¹, Fubito Toyama², Eriko Takayama-Watanabe*^,3 and Akihiko Watanabe*^,1

¹Faculty of Science, Yamagata University, 1-4-12 Kojirakawa, Yamagata, ²Graduate School of Engineering, Utsunomiya University, Utsunomiya, Tochigi, ³Institutes of Arts and Sciences, Yamagata University, 1-4-12 Kojirakawa, Yamagata, Japan

Abstract

The acrosome reaction is induced in the sperm of Cynops pyrrhogaster immediately in response to a ligand protein called acrosome reaction-inducing substance (ARIS) in the egg jelly at fertilization, whereas a spontaneous acrosome reaction occurs time-dependently in correlation with the decline of sperm quality for fertilization. The ARIS-induced acrosome reaction was recently found to be mediated by TRPV4 in association with the NMDA type glutamate receptor, although the intracellular mediators for the acrosome reaction are largely unclear. In the present study, spontaneous acrosome reaction was significantly inhibited by Ni²⁺, RN1734, and diltiazem, which blocks Cav3.2, TRPV4 or TRPM8, and the cyclic nucleotide-gated channel, respectively. In contrast, expression of Ca²⁺-activated transmembrane and soluble adenylyl cyclases was detected in the sperm of C. pyrrhogaster by reverse transcription-polymerase chain reaction. Activator of transmembrane or soluble adenylyl cyclases (forskolin or HCO₃^-) independently promoted spontaneous acrosome reaction, while an inhibitor of each enzyme (MD12330A or KH7) inhibited it only in the sperm with high potential for spontaneous acrosome reaction. An inhibitor of protein kinase A (H89) inhibited spontaneous acrosome reaction in a manner independent of sperm potential for spontaneous acrosome reaction. Surprisingly, KH7 significantly inhibited ARIS-induced acrosome reaction, but its effect was seen in a small percentage of sperm. H89 had no effect on ARIS-induced acrosome reaction. These results suggest that C. pyrrhogaster sperm possess multiple intracellular pathways for acrosome reaction, involving Ca²⁺ permeable channels, adenylyl cyclases and PKA, and that two pathways having distinct dependencies on adenylyl cyclases may contribute to ARIS-induced acrosome reaction at fertilization.

Keywords

acrosome reaction, Ca²⁺ channel, adenylyl cyclase, PKA, sperm, urodele