Investigating ethanol toxicity at the cellular level has been the overarching theme of my research career. Much of my focus has been trying to understand how alcohol kills Chinese hamster ovary (CHO) cells, an easy-to-culture mammalian cell line that offers expedience and convenience. My work in CHO cells has focused on whole-cell approaches to determine how certain factors influence cell survival and viability following ethanol exposure. There are two limitations to this approach. First, the hallmark of human alcoholism is neuronal cell death, and CHO cells, while practical, are not complex neurons. Second, a whole-cell focus does not take into account subcellular processes, and will be insufficient for uncovering the precise mechanism of ethanol neurotoxicity. The proposed research seeks to broaden my intellectual perspectives by moving my ethanol toxicity research into human neurons (SH-SY5Y cells) in order to investigate the subcellular mechanism of ethanol neurotoxicity. This new endeavor will transition my current work at the cellular level to the subcellular realm where neuronal calcium dynamics in response to ethanol will be explored. This is a dramatic shift for my research, requiring the introduction of new techniques and model systems into my laboratory. While the aims outlined in this proposal are new endeavors for me, requiring new technical approaches and equipment, they build upon existing expertise and laboratory infrastructure and are highly feasible and achievable. The potential impact of the proposed work is significant, as I will be able to expand my current whole-cell thinking to the subcellular level. Simply put, this renewed, deeper focus will help reinvigorate and stimulate my mid-career research.