Gluconeogenesis (GNG), a key metabolic process, involves the formation of glucose and glycogen from non-glucose precursors via pyruvate. In the strict sense, it also includes the contribution of glycerol as well as recycled glucose carbon (Cori's cycle). The developmental expression of GNG in the fetus and newborn and the quantitative contribution of GNG to glucose has been extensively investigated in humans and other mammalian species. Data from studies in rodents, rabbits, and sheep fetuses show that the development of GNG is a well-orchestrated process that is regulated by the expression of specific factors involved in the transcription of the genes for specific regulating enzymes, which catalyze GNG. These transcription factors and the genes for gluconeogenic enzymes are expressed at specific time periods during development. Although the fetus has the potential for GNG, the actual formation of glucose from pyruvate is not apparent until after birth because the rate limiting enzyme phosphoenolpyruvate carboxykinase appears only after birth in the immediate newborn period. Several tracer isotope methods have been employed to quantify the contribution of GNG to glucose. Of these, the recently developed stable isotope techniques with deuterium labeled water and the mass isotopomer distribution analysis appear to be the most precise and easily applicable in human studies. The available data show that in the human newborn, GNG appears soon after birth and contributes 30% to 70% to glucose produced. Application of new molecular biology techniques, in combination with sensitive tracer isotopic methods, will allow us to identify and examine metabolic disorders that impact GNG and help develop intervention strategies.