How does Adenosine Alter Sperm Motility?

Abstract

Adenosine is a candidate modulator of motility of spermatozoa as they progress through the female reproductive tract. Past work demonstrated that the adenosine analog 2-chloro-deoxyadenosine (Cl-dAdo) accelerates the flagellar beat rate of mouse sperm by a cAMP/PKA-mediated pathway with a pharmacological profile suggestive of a Slc29a (ENT) equilibrative nucleoside transporter. Using qRT-PCR we find in adult mouse testis 160- and 32-fold greater expression of Slc29a1 than of the other surface-membrane ENT transporters Slc29a4 and Slc29a2. However, Slc29a1 protein was not found on mature sperm using immunocytochemistry. Moreover, wildtype and Slc29a1-null mice accelerate at similar rates (1.5-2.0 Hz min−1) in response to Cl-dAdo indicating that Slc29a1 is not required for Cl-dAdo action. Consistent with this observation, the accelerating action of Cl-dAdo resists the Slc29a-selective inhibitor nitrobenzylthioinosine (NBTI; 10 μM). The accelerating action of Cl-dAdo additionally resists replacement of external Na+ with NMDG+ indicating that Slc28a concentrative nucleoside transporters (CNTs) also are not required. Interestingly, the Adenosine A3 receptor-selective agonist Cl-IB-MECA (25 μM) is nearly as effective as Cl-dAdo in accelerating sperm beat frequency, suggesting a possible role for cell surface A3 receptors in Cl-dAdo-mediated increases in sperm motility. Two A3 isoforms are expressed in the mouse Adora3i1 and Adora3i2; Adora3i2 expression is testis specific. Adora3i1 null sperm increase beat frequency in response to both Cl-IB-MECA (25 μM) and Cl-dAdo (25 μM), so this isoform is not needed for sperm response to adenosine. Sperm response to Cl-dAdo and Cl-IB-MECA is diminished after pertussis toxin treatment of cells suggesting the receptor is Gαi/o coupled. We are currently testing functionality of the testis-specific novel Adora3i2 isoform in a heterologous system. Support from U54-HD12629 of the SCCPRR program of NICHD. L.A.B. supported in part by 5-T32-HD007453.