Submitted on February 28, 2007
Accepted on April 18, 2008
Inosine and equilibrative nucleoside transporter 2 (ENT2) contribute to hypoxic preconditioning in the murine cardiomyocyte cell line, HL-1
Zlatina Naydenova1, Jennifer B Rose1, and Imogen R Coe1*
1 Biology, York University, Toronto, Canada
* To whom correspondence should be addressed. E-mail: coe{at}yorku.ca.
The purine nucleoside adenosine (Ado) is a physiologically important molecule in the heart. Brief exposure of cardiomyocytes to hypoxic challenge results in the production of extracellular Ado, which then interacts with Ado receptors to activate compensatory signaling pathways that lead to cellular resistance to subsequence hypoxic challenge. This phenomenon is known as preconditioning (PC) and while Ado is clearly involved, other components of the response are less well understood. Flux of nucleosides such as Ado and inosine, across cardiomyocyte membranes is dependent on equilibrative nucleoside transporters, ENT1 and ENT2. We have previously shown in the murine cardiomyocyte cell line, HL-1, that hypoxic challenge leads to an increase in intracellular Ado, which exits the cell via ENT1 and preconditions via A1 and A3 Ado-receptor dependent mechanisms. However, the role and contribution of inosine and ENT2 is unclear. In this study, we confirmed that ENT1 and ENT2 are both capable of transporting inosine. Moreover, we found that hypoxic challenge leads to a significant increase in levels of intracellular inosine, which exits the cell via both ENT1 and ENT2. Exogenously added inosine (5µM) preconditions cardiomyocytes in an A1 Ado receptor dependent manner since preconditioning can be blocked by the A1 Ado receptor antagonist, DPCPX (1µM) but not the A3 Ado receptor antagonist MRS 1220 (200nM). These data suggest that cardiomyocyte responses to hypoxic PC are more complex than previously thought involving both Ado and inosine and differing, but overlapping, contributions of the two ENT isoforms.
