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Glutamine metabolism and function in skeletal muscle

University of British Columbia

Glutamine is an amino acid possessing a number of physiological functions in addition to its contribution as a substrate for protein synthesis. Skeletal muscle is a major source of glutamine synthesized in vivo. The main objectives of this thesis were to study regulation of the glutamine synthesis and to characterize glutamine's effect on protein turnover in skeletal muscle. Regulation of glutamate dehydrogenase (GDH) by branched-chain amino acids (BCAA) was first investigated. Leucine and isoleucine were found to increase the activity of GDH in rat skeletal muscle in a dose-dependent manner [45% (P<0.01) and 27% (P<0.05) by 1 mM leucine and 1 mM isoleucine, respectively]. The effects of leucine and isoleucine on GDH activity are additive, suggesting different regulatory sites for each amino acid. Leucine, but not isoleucine, was also found to increase GDH activity in chick skeletal muscle (36% at 1 mM leucine, P<0.05). Acidosis, a condition under which glutamine requirement by the kidney is substantially increased, upregulated the activities of both GDH and glutamine synthetase (GS) in rat skeletal muscle. Following 5-d feeding 1.5% NH4C1, the activity of GDH and GS increased 88% and 66%, respectively (P<0.01). Acidosis also increased the sensitivity of rat skeletal muscle GDH to regulation by all three BCAA. These data provide information to complete our understanding in regulation of glutamine synthesis in skeletal muscle. In the second part of this thesis, the effect of glutamine on protein turnover in rat skeletal muscle cells was characterized using myotubes differentiated from L8 rat skeletal myoblasts. Glutamine (0.65 - 15 mM) was found to increase the rate of protein synthesis in stressed myotubes (43°C for 45 min) (P<0.05) but not in normal-cultured myotubes. Glutamine (0.65 - 15 mM) decreased the rate of degradation of long-lived proteins in both heat-stressed and normal-cultured myotubes (P<0.05) but had no influence on the rate of degradation of short-lived proteins. In an attempt to study the mechanism underlying the effects of glutamine on protein turnover in myotubes, glutamine was found to increase the expression of heat-shock protein 70 (HSP70) at both the protein level (7.5-fold increase by 10 mM glutamine, P<0.001) and the mRNA level (7.8-fold increase by 10 mM glutamine, P< 0.001) in heat-stressed myotubes but not in normal-cultured myotubes. This effect of glutamine may be specific to HSP70 as the expression of HSP27 was not altered by glutamine. In spite of an increase in the abundance of hsp7O mRNA, the DNA binding activity of heat shock transcription factor was not influenced by glutamine. It is concluded that the effect of glutamine on protein turnover in skeletal muscle is condition-dependent, and that HSP70 may play a role in the mechanism underlying the anabolic effect of glutamine in skeletal muscle.

Thesis/Dissertation

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2009-03-17

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http://hdl.handle.net/2429/6190

Animal Science

University of British Columbia

UBCV

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