A major issue in stem cell biology is a lack of understanding of the basic biology of the pluripotent state. In vitro, human pluripotent stem cells (PSCs) are difficult to maintain as they are subject to spontaneous differentiation and cell death. We hypothesize that a better understanding of how these cells consume and process metabolites will shed light on the pluripotent state and lead to improved methods for the culture and differentiation of PSCs. We and others have shown that human embryonic stem cells (hESCs) exhibit increased glycolytic metabolism and reduced respiration relative to differentiated counterparts, and that altering glucose or oxygen levels can impact hESC differentiation efficiency and pluripotency. However, the mechanisms by which changes in PSC metabolism result in cell fate changes remain unclear. We are currently investigating the role of glycolytic metabolism in stem cell biology, specifically in hESC self-renewal and differentiation capacity.