Integrating Macronutrient Metabolism In Developing Chicken Embryos
Sunny, Nishanth Edakulathur
Bequette, Brian J
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The objective of this thesis research was to determine the pathways of glucose metabolism and utilization in small and large egg embryos during the latter half of development, and whether <em>in ovo</em> nutrient supplementation alters glucose use. A further objective was to determine the contribution of glutamate, glutamine and glycerol to glucose, glycogen and non essential amino acid (NEAA) synthesis during embryo development. In ovo stable isotope ([U-<sup>13</sup>C]glucose, [U-<sup>13</sup>C] glutamate, [U-<sup>13</sup>C]glutamine and [U-<sup>13</sup>C]glycerol) injection approaches were developed along with mass isotopomer distribution analysis of metabolic intermediates and end-products to acquire a metabolic phenotype of the fluxes and partition of these substrates through central pathways. Embryos developing in small and large eggs maintained similar rates of glucose metabolism. Thus, glucose entry and utilization gradually increased from day 12 to 18 embryonic. By embryonic day 20, gluconeogenesis accounted for >80% of glucose entry, a part (65%) of which was represented by glucose carbon recycling. Glutamate and glutamine were not found to be significant gluconeogenic precursors in day 19 embryos. However, catabolism of these amino acids contributed to ~25% of proline flux in the liver. By contrast, there was significant [M+3] <sup>13</sup>Cisotopomer abundance in blood glucose and in liver and muscle glycogen when [U- <sup>13</sup>C]glycerol was injected <em>in ovo</em>. These observations clearly confirmed that glycerol derived from triacylglycerides is a significant precursor for glucose and glycogen synthesis. In ovo supplementation on day 9 embryonic of glucose and/or amino acids (5 non-essential amino acids) did not alter gluconoegenesis. However, these supplemental treatments significantly reduced catabolism of glucose via glycolysis. <sup>13</sup>C-Mass isotopomer abundances of most substrates differed when each was individually compared in blood and in the various tissues, indicating differences in substrate utilization between tissues. In summary, this thesis research has provided new information on the degree and pathways of nutrient (glucose, glycerol, amino acids) use by the developing embryo and the rapid adjustments in the activity of networks of enzymes involved in non-essential amino acid, glucose and glycogen metabolism to support embryo survival. Most importantly, this work has systematically evaluated the potential substrates that the embryo utilizes for glucose synthesis, in particular, the significant role of glycerol.