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Nutrient Physiology, Metabolism, and Nutrient-Nutrient Interactions

Glutamate Is the Major Anaplerotic Substrate in the Tricarboxylic Acid Cycle of Isolated Rumen Epithelial and Duodenal Mucosal Cells from Beef Cattle^1,2

³ Department of Animal and Avian Sciences, University of Maryland, College Park, MD 20742; ⁴ Bovine Genomic Laboratory, Animal and Natural Resources Institute, USDA-Agricultural Research Service, Beltsville, MD 20705; and ⁵ University of Kentucky, Lexington, KY 40546

In this study, we aimed to determine the contribution of substrates to tricarboxylic acid (TCA) cycle fluxes in rumen epithelial cells (REC) and duodenal mucosal cells (DMC) isolated from Angus bulls (n = 6) fed either a 75% forage (HF) or 75% concentrate (HC) diet. In separate incubations, [¹³C₆]glucose, [¹³C₅]glutamate, [¹³C₅]glutamine, [¹³C₆]leucine, or [¹³C₅]valine were added in increasing concentrations to basal media containing SCFA and a complete mixture of amino acids. Lactate, pyruvate, and TCA cycle intermediates were analyzed by GC-MS followed by ¹³C-mass isotopomer distribution analysis. Glucose metabolism accounted for 10–19% of lactate flux in REC from HF-fed bulls compared with 27–39% in REC from HC and in DMC from bulls fed both diets (P < 0.05). For both cell types, as concentration increased, an increasing proportion (3–63%) of -ketoglutarate flux derived from glutamate, whereas glutamine contributed <3% (P < 0.05). Although leucine and valine were catabolized to their respective keto-acids, these were not further metabolized to TCA cycle intermediates. Glucose, glutamine, leucine, and valine catabolism by ruminant gastrointestinal tract cells has been previously demonstrated, but in this study, their catabolism via the TCA cycle was limited. Further, although glutamate's contribution to TCA cycle fluxes was considerable, it was apparent that other substrates available in the media also contributed to the maintenance of TCA fluxes. Lastly, the results suggest that diet composition alters glucose, glutamate, and leucine catabolism by the TCA cycle of REC and DMC.

^* To whom correspondence should be addressed. E-mail: bbequett{at}umd.edu.

Manuscript received 5 December 2008. Initial review completed 5 January 2009. Revision accepted 6 February 2009.

Published online 12 March 2009.