Metabolic fates of ammonia-N in ruminal epithelial and duodenal mucosal cells isolated from growing sheep.
The objective of this experiment was to determine the capability of ruminant gut tissues to detoxify ammonia-N using short-term incubations of isolated cells in vitro. Ruminal epithelial cells (REC) and duodenal mucosal cells (DMC) were isolated from growing Texel-Polypay ram lambs (n = 4) fed a pelleted forage:concentrate-based diet. Immediately after isolation, primary cells were incubated for 60 min with glucose (1mM), glutamate (1mM), [15N]ammonium chloride (5, 10, 20, or 40 mM), and 1 of 4 combinations of substrates (1 mM each) that could support urea synthesis [control, N-carbamoylglutamate (NCG); NCG + ornithine (ONCG); and ONCG + aspartate (AONCG)]. Treatments were arranged in a 4 x 4 factorial design. Incorporation of ammonia-15N into alanine, citrulline, arginine, and urea was determined by gas chromatography-mass spectrometry. For both cell types, ammonia-N transfer to alanine was lower when incubation medium contained NCG compared with control, whereas use of ammonia-N for net alanine synthesis increased quadratically with ammonia concentration regardless of substrate treatment. For REC, ammonia-N was not incorporated into citrulline, arginine, or urea, nor into arginine or urea by DMC. Ammonia-N use for net citrulline synthesis exhibited an inverse relationship with ammonia concentration, decreasing linearly as media ammonia concentration increased. Thus, ala-nine synthesis may be a significant metabolic pathway for ruminant gut tissues to detoxify ammonia-N when it is presented luminally at high concentrations as compared with detoxification by the ornithine-urea cycle. Furthermore, DMC do exhibit a metabolic capability to incorporate ammonia-N into citrulline, but low or absent activity of downstream enzymes of the ornithine-urea cycle appears to limit ammonia-N transfers to urea.
Oba
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Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Canada T6G 2P5. masahito.oba@ualberta.ca
Baldwin
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Owens
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Bequette
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BJ
D000409:Alanine / Q000378:metabolism; D000641:Ammonia / Q000378:metabolism*; D000818:Animals; D001120:Arginine / Q000378:metabolism; D001224:Aspartic Acid / Q000378:metabolism; D002956:Citrulline / Q000378:metabolism; D004386:Duodenum / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D018698:Glutamic Acid / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism*; D009584:Nitrogen / Q000378:metabolism*; D009587:Nitrogen Isotopes; D012417:Rumen / Q000378:metabolism*; D012756:Sheep / Q000378:metabolism*; D014508:Urea / Q000378:metabolism
D009587:Nitrogen Isotopes; D002956:Citrulline; D001224:Aspartic Acid; D018698:Glutamic Acid; D000641:Ammonia; D014508:Urea; D001120:Arginine; D009584:Nitrogen; D000409:Alanine