Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, PC311

Poster Communications

P2X subunits expression in mouse Leydig cells at different ages

L. S. Antonio1, R. R. Costa1, J. F. Aguiar1, M. D. Gomes2, W. A. Varanda1

1. Department of Physiology, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil. 2. Department of Biochemistry and Immunology, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil.

ATP acts on the plasma membrane by interacting with purinergic receptors P2X (ligand-gated ion channels) and P2Y (G protein-coupled receptors). Until now, seven subunits of P2X receptors were identified and cloned (P2X1 - P2X7). They form functional receptors with distinct biophysical and pharmacological properties depending of their structure. Calcium influx through these channels is in part responsible for the increase in intracellular [Ca2+] observed in a number of physiological situations. ATP treatment of Leydig cells from mice and rats, leads to an increase in [Ca2+]i and testosterone secretion. As testosterone production is an age dependent process, our hypothesis here is that the pattern of specific P2X subunits expression in Leydig cells changes along the development. Based on the developmental stages defined by Wu et al. (2010) the purpose of this work was to investigate the expression of P2X receptor subunits in mouse Leydig cells aged 07, 14, 21, 24, 28, 35, 45, 60, 75 and 90 days old. Western blot experiments were performed in freshly isolated Leydig cells purified in a percoll discontinuous gradient essentially as described in Salva et al., (2001). Polyclonal antibodies (Alomone Labs Ltd., Jerusalem, Israel) (1:200) against P2X2, P2X4 and P2X7 subunits were used in order to detect a particular type of receptor. Our results showed P2X2 expression from day 28 till day 100, P2X4 expression from day 21 till day 100 and P2X7 expression from day 35 till day 75. The expression pattern of P2X4 and P2X7 subunits matches the temporal expression of the steroidogenic enzymes P450scc and P450c17alpha (day 24 till day 90) and the expression pattern of P2X2 match the plasma testosterone levels in mice (day 24 till day 180) as describe by Wu et al., 2010. We suggest that purinergic P2X receptors in mouse Leydig cells could modulate the steroidogenic process and influence testoterone production.

Where applicable, experiments conform with Society ethical requirements