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Alcohol-induced decrease in muscle protein synthesis associated with increased binding of mTOR and raptor: Comparable effects in young and mature rats.

Lang CH, Pruznak AM, Nystrom GJ, Vary TC - Nutr Metab (Lond) (2009)

Bottom Line: Acute alcohol (EtOH) intoxication decreases muscle protein synthesis via inhibition of mTOR-dependent translation initiation.Furthermore, some in vivo-produced effects of EtOH vary in an age-dependent manner.The EtOH-induced changes in both groups were associated with a concomitant reduction in 4E-BP1 phosphorylation, and redistribution of eIF4E between the active eIF4E.eIF4G and inactive eIF4E.4EBP1 complex.

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Affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033, USA. clang@psu.edu.

ABSTRACT

Background: Acute alcohol (EtOH) intoxication decreases muscle protein synthesis via inhibition of mTOR-dependent translation initiation. However, these studies have been performed in relatively young rapidly growing rats in which muscle protein accretion is more sensitive to growth factor and nutrient stimulation. Furthermore, some in vivo-produced effects of EtOH vary in an age-dependent manner. The hypothesis tested in the present study was that young rats will show a more pronounced decrement in muscle protein synthesis than older mature rats in response to acute EtOH intoxication.

Methods: Male F344 rats were studied at approximately 3 (young) or 12 (mature) months of age. Young rats were injected intraperitoneally with 75 mmol/kg of EtOH, and mature rats injected with either 75 or 90 mmol/kg EtOH. Time-matched saline-injected control rats were included for both age groups. Gastrocnemius protein synthesis and the activity of the mTOR pathway were assessed 2.5 h after EtOH using [³H]-labeled phenylalanine and the phosphorylation of various protein factors known to regulate peptide-chain initiation.

Results: Blood alcohol levels (BALs) were lower in mature rats compared to young rats after administration of 75 mmol/kg EtOH (154 ± 23 vs 265 ± 24 mg/dL). However, injection of 90 mmol/kg EtOH in mature rats produced BALs comparable to that of young rats (281 ± 33 mg/dL). EtOH decreased muscle protein synthesis similarly in both young and high-dose EtOH-treated mature rats. The EtOH-induced changes in both groups were associated with a concomitant reduction in 4E-BP1 phosphorylation, and redistribution of eIF4E between the active eIF4E.eIF4G and inactive eIF4E.4EBP1 complex. Moreover, EtOH increased the binding of mTOR with raptor in a manner which appeared to be AMPK- and TSC-independent. In contrast, although muscle protein synthesis was unchanged in mature rats given low-dose EtOH, compared to control values, the phosphorylation of rpS6 and eIF4G was decreased.

Conclusion: These data indicate that muscle protein synthesis is equally sensitive to the inhibitory effects of EtOH in young rapidly growing rats and older mature rats which are growing more slowly, but that mature rats must be given a relatively larger dose of EtOH to achieve the same BAL. Based on the differential response in mature rats to low- and high-dose EtOH, the decreased protein synthesis was associated with a reduction in mTOR activity which was selectively mediated via a reduction in 4E-BP1 phosphorylation and an increase in mTOR.raptor formation.

No MeSH data available.


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Effect of acute alcohol intoxication on the binding of mTOR with raptor in skeletal muscle from young and mature rats. Groups are the same as described in Figure 1. Gastrocnemius was collected 2.5 h after administration of alcohol or saline (control). Inset at top: raptor was immunoprecipitated (IP) and the amount of mTOR and raptor determined by immunoblotting (IB). Bottom graph: densitometric analysis of immunoblots of mTOR bound to immunoprecipitated raptor in muscle. There was no difference among the various groups for the amount of immunoprecipitated raptor (mean data not shown). Values (means ± SEM) are expressed relative to the young saline-treated control group. Sample size was 10, 10, 6, 7, 9, and 9 for the six groups, respectively. Values with different letters are significantly different from each other, P < 0.05. Values which share a common letter are not statistically different.
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Figure 5: Effect of acute alcohol intoxication on the binding of mTOR with raptor in skeletal muscle from young and mature rats. Groups are the same as described in Figure 1. Gastrocnemius was collected 2.5 h after administration of alcohol or saline (control). Inset at top: raptor was immunoprecipitated (IP) and the amount of mTOR and raptor determined by immunoblotting (IB). Bottom graph: densitometric analysis of immunoblots of mTOR bound to immunoprecipitated raptor in muscle. There was no difference among the various groups for the amount of immunoprecipitated raptor (mean data not shown). Values (means ± SEM) are expressed relative to the young saline-treated control group. Sample size was 10, 10, 6, 7, 9, and 9 for the six groups, respectively. Values with different letters are significantly different from each other, P < 0.05. Values which share a common letter are not statistically different.

Mentions: As a result of its predominant role in regulating mTOR activity, the amount of the mTOR complex 1 (mTORC1) was determined. The mTORC1 is composed of at least four proteins, including mTOR, raptor, GβL, and PRAS40 [32]. Western blot analysis of whole muscle homogenate did not demonstrate a significant age- or alcohol-induced change for total mTOR, raptor, GβL or PRAS40 (data not shown). However, in muscle from either alcohol-treated young rats or mature rats given the high-dose of alcohol, the association of mTOR with immunoprecipitated raptor was increased approximately 35% (Figure 5). Such a change in mTOR·raptor binding was not seen in muscle of mature rats given the lower dose of alcohol. In contrast, there was no age-or alcohol-induced change in the binding of GβL or PRAS40 to raptor (data not shown).


Alcohol-induced decrease in muscle protein synthesis associated with increased binding of mTOR and raptor: Comparable effects in young and mature rats.

Lang CH, Pruznak AM, Nystrom GJ, Vary TC - Nutr Metab (Lond) (2009)

Effect of acute alcohol intoxication on the binding of mTOR with raptor in skeletal muscle from young and mature rats. Groups are the same as described in Figure 1. Gastrocnemius was collected 2.5 h after administration of alcohol or saline (control). Inset at top: raptor was immunoprecipitated (IP) and the amount of mTOR and raptor determined by immunoblotting (IB). Bottom graph: densitometric analysis of immunoblots of mTOR bound to immunoprecipitated raptor in muscle. There was no difference among the various groups for the amount of immunoprecipitated raptor (mean data not shown). Values (means ± SEM) are expressed relative to the young saline-treated control group. Sample size was 10, 10, 6, 7, 9, and 9 for the six groups, respectively. Values with different letters are significantly different from each other, P < 0.05. Values which share a common letter are not statistically different.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2651172&req=5

Figure 5: Effect of acute alcohol intoxication on the binding of mTOR with raptor in skeletal muscle from young and mature rats. Groups are the same as described in Figure 1. Gastrocnemius was collected 2.5 h after administration of alcohol or saline (control). Inset at top: raptor was immunoprecipitated (IP) and the amount of mTOR and raptor determined by immunoblotting (IB). Bottom graph: densitometric analysis of immunoblots of mTOR bound to immunoprecipitated raptor in muscle. There was no difference among the various groups for the amount of immunoprecipitated raptor (mean data not shown). Values (means ± SEM) are expressed relative to the young saline-treated control group. Sample size was 10, 10, 6, 7, 9, and 9 for the six groups, respectively. Values with different letters are significantly different from each other, P < 0.05. Values which share a common letter are not statistically different.
Mentions: As a result of its predominant role in regulating mTOR activity, the amount of the mTOR complex 1 (mTORC1) was determined. The mTORC1 is composed of at least four proteins, including mTOR, raptor, GβL, and PRAS40 [32]. Western blot analysis of whole muscle homogenate did not demonstrate a significant age- or alcohol-induced change for total mTOR, raptor, GβL or PRAS40 (data not shown). However, in muscle from either alcohol-treated young rats or mature rats given the high-dose of alcohol, the association of mTOR with immunoprecipitated raptor was increased approximately 35% (Figure 5). Such a change in mTOR·raptor binding was not seen in muscle of mature rats given the lower dose of alcohol. In contrast, there was no age-or alcohol-induced change in the binding of GβL or PRAS40 to raptor (data not shown).

Bottom Line: Acute alcohol (EtOH) intoxication decreases muscle protein synthesis via inhibition of mTOR-dependent translation initiation.Furthermore, some in vivo-produced effects of EtOH vary in an age-dependent manner.The EtOH-induced changes in both groups were associated with a concomitant reduction in 4E-BP1 phosphorylation, and redistribution of eIF4E between the active eIF4E.eIF4G and inactive eIF4E.4EBP1 complex.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033, USA. clang@psu.edu.

ABSTRACT

Background: Acute alcohol (EtOH) intoxication decreases muscle protein synthesis via inhibition of mTOR-dependent translation initiation. However, these studies have been performed in relatively young rapidly growing rats in which muscle protein accretion is more sensitive to growth factor and nutrient stimulation. Furthermore, some in vivo-produced effects of EtOH vary in an age-dependent manner. The hypothesis tested in the present study was that young rats will show a more pronounced decrement in muscle protein synthesis than older mature rats in response to acute EtOH intoxication.

Methods: Male F344 rats were studied at approximately 3 (young) or 12 (mature) months of age. Young rats were injected intraperitoneally with 75 mmol/kg of EtOH, and mature rats injected with either 75 or 90 mmol/kg EtOH. Time-matched saline-injected control rats were included for both age groups. Gastrocnemius protein synthesis and the activity of the mTOR pathway were assessed 2.5 h after EtOH using [³H]-labeled phenylalanine and the phosphorylation of various protein factors known to regulate peptide-chain initiation.

Results: Blood alcohol levels (BALs) were lower in mature rats compared to young rats after administration of 75 mmol/kg EtOH (154 ± 23 vs 265 ± 24 mg/dL). However, injection of 90 mmol/kg EtOH in mature rats produced BALs comparable to that of young rats (281 ± 33 mg/dL). EtOH decreased muscle protein synthesis similarly in both young and high-dose EtOH-treated mature rats. The EtOH-induced changes in both groups were associated with a concomitant reduction in 4E-BP1 phosphorylation, and redistribution of eIF4E between the active eIF4E.eIF4G and inactive eIF4E.4EBP1 complex. Moreover, EtOH increased the binding of mTOR with raptor in a manner which appeared to be AMPK- and TSC-independent. In contrast, although muscle protein synthesis was unchanged in mature rats given low-dose EtOH, compared to control values, the phosphorylation of rpS6 and eIF4G was decreased.

Conclusion: These data indicate that muscle protein synthesis is equally sensitive to the inhibitory effects of EtOH in young rapidly growing rats and older mature rats which are growing more slowly, but that mature rats must be given a relatively larger dose of EtOH to achieve the same BAL. Based on the differential response in mature rats to low- and high-dose EtOH, the decreased protein synthesis was associated with a reduction in mTOR activity which was selectively mediated via a reduction in 4E-BP1 phosphorylation and an increase in mTOR.raptor formation.

No MeSH data available.


Related in: MedlinePlus