Cardioprotection in an in vitro model of hypoxic preconditioning

Short periods of myocardial ischemia appear to provide protection against subsequent prolonged ischemic episodes in experimental animals and in man. This phenomenon, known as ischemic preconditioning, has not yet been characterized at the cellular or molecular levels; however, tissue hypoxia appears to be required. In this study, we used a previously developed method for hypoxic cardiac myocyte culture in order to establish a model for ischemic (or hypoxic) preconditioning in cell culture. We demonstrate that cultured neonatal rat cardiac myocytes preconditioned by 25 min of exposure to hypoxia followed by reoxygenation were protected against membrane damage for up to 6 h of prolonged severe hypoxia, as determined by arachidonic acid release and contractile recovery. In contrast, non-preconditioned myocytes exhibited significant hypoxic damage after 2-4 h. Pretreatment of cells with PMA, a tumor-promoting phorbol ester, mimicked the protective effects of hypoxic preconditioning; pretreatment with the muscarinic cholinergic agonist carbachol had no effect. Our data suggests that isolated myocytes in culture remain competent to be preconditioned by hypoxia, through a pathway that may involve the activation of protein kinase C.