Neuregulin-1 (NRG-1) is a stress-mediated growth factor secreted by cardiovascular endothelial cells and provides the protection to myocardial cells, but the underlying mechanisms are not fully understood. This study aimed to demonstrate that NRG-1 protects myocardial cells exposed to oxidative damage by regulating endoplasmic reticulum (ER) stress. Neonatal rat cardiac myocytes (NRCMs) were isolated and treated with H2O2 as a cellular model of ER stress. NRCMs were pretreated with different concentrations of NRG-1. We found that NRG-1 increased the viability and reduced the apoptosis of NRCMs treated by H2O2. Moreover, NRG-1 reduced lactate dehydrogenase level, increased superoxide dismutase activity and decreased malondialdehyde content in NRCMs treated by H2O2. Finally, we demonstrated that NRG-1 alleviated ER stress and decreased CHOP and GRP78 protein levels in NRCMs treated by H2O2. Taken together, these data indicate that NRG-1 relieves oxidative and ER stress in NRCMs and suggest that NRG-1 is a promising agent for cardioprotection. Copyright (C) 2014 John Wiley & Sons, Ltd.