Osmolytes are rapidly lost from the ischemic heart, an effect thought to benefit the heart by reducing the osmotic load. However, the observation that chronic lowering of one of the prominent osmolytes,
taurine, is more beneficial to the ischemic heart than acute
taurine loss suggests that osmotic stress may benefit the ischemic heart through multiple mechanisms. The present study examines the possibility that chronic osmotic stress preconditions the heart in part by stimulating a cardioprotective, osmotic-linked signaling pathway. Hyperosmotic stress was produced by treating rat neonatal cardiomyocytes during the pre-hypoxic period with either the
taurine depleting agent,
beta-alanine (5 mM), or with medium containing 25 mM
mannitol. The cells were then subjected to chemical
hypoxia in medium containing 3 mM
Amytal and 10 mM
deoxyglucose but lacking
beta-alanine and
mannitol. Cells that had been pretreated with either 5 mM
beta-alanine or 25 mM
mannitol exhibited resistance against
hypoxia-induced apoptosis and
necrosis. Associated with the osmotically preconditioned state was the activation of Akt and the inactivation of the pro-apoptotic factor, Bad, both events blocked by the inhibition of
PI 3-kinase. However, preconditioning the cardiomyocyte with
mannitol had no effect on the generation of
free radicals during the hypoxic period. Osmotic stress also promoted the upregulation of the anti-apoptotic factor, Bcl-2. Since inhibition of
PI 3-kinase with
Wortmannin also prevents osmotic-mediated cardioprotection, we conclude that hyperosmotic-mediated activation of the
PI 3-kinase/Akt pathway contributes to osmotic preconditioning.