Presence and function of dopamine transporter (DAT) in stallion sperm: dopamine modulates sperm motility and acrosomal integrity.

Javier A Urra,Joan Enric Rodríguez-Gil,Ilona I Concha,Alejandra A Covarrubias,Alfredo Ramírez-Reveco,Franz Villaroel-Espíndola,Chris D Wood

doi:10.1371/journal.pone.0112834

Javier A Urra, Joan Enric Rodríguez-Gil + Show 5 more

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https://doi.org/10.1371/journal.pone.0112834

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Abstract

Dopamine is a catecholamine with multiple physiological functions, playing a key role in nervous system; however its participation in reproductive processes and sperm physiology is controversial. High dopamine concentrations have been reported in different portions of the feminine and masculine reproductive tract, although the role fulfilled by this catecholamine in reproductive physiology is as yet unknown. We have previously shown that dopamine type 2 receptor is functional in boar sperm, suggesting that dopamine acts as a physiological modulator of sperm viability, capacitation and motility. In the present study, using immunodetection methods, we revealed the presence of several proteins important for the dopamine uptake and signalling in mammalian sperm, specifically monoamine transporters as dopamine (DAT), serotonin (SERT) and norepinephrine (NET) transporters in equine sperm. We also demonstrated for the first time in equine sperm a functional dopamine transporter using 4-[4-(Dimethylamino)styryl]-N-methylpyridinium iodide (ASP+), as substrate. In addition, we also showed that dopamine (1 mM) treatment in vitro, does not affect sperm viability but decreases total and progressive sperm motility. This effect is reversed by blocking the dopamine transporter with the selective inhibitor vanoxerine (GBR12909) and non-selective inhibitors of dopamine reuptake such as nomifensine and bupropion. The effect of dopamine in sperm physiology was evaluated and we demonstrated that acrosome integrity and thyrosine phosphorylation in equine sperm is significantly reduced at high concentrations of this catecholamine. In summary, our results revealed the presence of monoamine transporter DAT, NET and SERT in equine sperm, and that the dopamine uptake by DAT can regulate sperm function, specifically acrosomal integrity and sperm motility.

Highlights

Dopamine is a catecholamine that participates in many biological processes in mammals, acting on functions related to cognition, emotions and control of motor activity, among others [1]
Using immunofluorescence and Western blot assays, we have demonstrated the presence of the dopamine transporter in equine sperm (Figure 1) and other species (Figure S1)
Considering the effect of high doses of dopamine in in vitro sperm function, in the control of sperm motility, maintenance of acrosomal integrity and tyrosine phosphorylation in a wide range of target proteins, it might be suggested that these higher catecholamine concentrations could participate in the modulation of in vivo sperm function

Summary

Introduction

Dopamine is a catecholamine that participates in many biological processes in mammals, acting on functions related to cognition, emotions and control of motor activity, among others [1]. There is evidence of expression of tyrosine hydroxylase (TH), a ratelimiting enzyme in catecholamine synthesis, in the uterus and cervical cells of mare [8], in neuronal-type cells of non-human primates [9] and in human Leydig cells [10]. This raises the possibility that catecholamines are synthesized from a source other than that of the innervation present in testis and oviduct, implying that sperm would be in contact with catecholamines, or at least with L-DOPA, a precursor of dopamine, from a very early stage during their passage through the male and female reproductive tract. They have been shown to induce capacitation in mouse [11], hamster [12,13] and bull sperm [14], as well as stimulating motility in vitro, in hamster sperm [15] and promoting the acrosome reaction in hamster [16,13] and bovine sperm [14]

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