Cellular energy metabolism correlates with cell fate, but the metabolic properties of chicken embryonic stem (chES) cells are poorly understood. Using a previously established chES cell model and electron microscopy (EM), we found that undifferentiated chES cells stored glycogen. Additionally, undifferentiated chES cells expressed lower levels of glucose transporter 1 (GLUT1) and phosphofructokinase (PFK) mRNAs but higher levels of hexokinase 1 (HK1) and glycogen synthase (GYS) mRNAs compared with control primary chicken embryonic fibroblast (CEF) cells, suggesting that chES cells direct glucose flux towards the glycogenic pathway. Moreover, we demonstrated that undifferentiated chES cells block gluconeogenic outflow and impede the accumulation of glucose-6-phosphate (G6P) from this pathway, as evidenced by the barely detectable levels of pyruvate carboxylase (PCX) and mitochondrial phosphoenolpyruvate carboxykinase (PCK2) mRNAs. Additionally, cell death occurred in undifferentiated chES cells as shown by Hoechst 33342 and propidium iodide (PI) double staining, but it could be rescued by exogenous G6P. However, we found that differentiated chES cells decreased the glycogen reserve through the use of PAS staining. Moreover, differentiated chES cells expressed higher levels of GLUT1, HK1 and PFK mRNAs, while the level of GYS mRNA remained similar in control CEF cells. These data indicate that undifferentiated chES cells continue to synthesize glycogen from glucose at the expense of G6P, while differentiated chES cells have a decreased glycogen reserve, which suggests that the amount of glycogen is indicative of the chES cell state.