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Title:Use of a dual isotope method to measure the interaction of branched chain amino acids and insulin in the stimulation of skeletal muscle protein synthesis
Author(s):Wibert, Gregory James
Doctoral Committee Chair(s):Layman, Donald K.
Department / Program:Nutritional Sciences
Discipline:Nutritional Sciences
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Biology, Animal Physiology
Health Sciences, Nutrition
Abstract:The extent to which branched-chain amino acids (bcaa) and insulin affect the regulation of in vivo skeletal muscle protein synthesis is controversial. To determine their roles in protein synthesis, a dual isotope method using $\sp{14}$C dansyl chloride was adapted for use with radioactive tyrosine. The primary difficulty in adapting the method was defining the stability of the $\sp3$H-label on the tyrosine molecule and characterizing the dansyl chloride reaction with tyrosine. To mitigate these problems: (1) L- (2,3,5,6-$\sp3$H) tyrosine was used, reducing $\sp3$H loss, (2) an acetonitrile/lithium carbonate solvent system was used to increase the efficiency and reproducibility of dansylated tyrosine recovery. These modifications yielded reproducible measurements of protein synthesis and fractional synthesis rates (%/day) that were comparable to literature values.
The effect of bcaa on protein synthesis was examined in fasted rats using a steady-state, constant infusion of a $\sp3$H tyrosine tracer and a primed constant leucine infusion with or without added isoleucine and valine. Protein synthesis in red gastrocnemius muscles increased at all levels of leucine regardless of isoleucine and valine concentrations. Leucine stimulated insulin secretion in a non-dose dependent manner. Infusion of isoleucine and valine in the absence of leucine increased insulin concentration without a comcomitant increase in protein synthesis.
Similar infusion experiments were conducted to examine the effects of insulin on protein synthesis. Elevations of insulin within physiological ranges failed to stimulate protein synthesis in the absence of elevated leucine. However, supraphysiological insulin concentrations stimulated protein synthesis without added leucine.
In summary: (1) a dual isotope method, using a $\sp3$H tyrosine tracer, provided a sensitive, precise measurement of protein synthesis, (2) leucine stimulated skeletal muscle protein synthesis in fasted rats in the absence of added isoleucine and valine, (3) physiological elevations of insulin stimulated protein synthesis in the presence, but not in the absence, of increased leucine, (4) valine and isoleucine did not interact with insulin to stimulate skeletal muscle protein synthesis. These studies imply that leucine is an important regulator of in vivo skeletal muscle protein synthesis.
Issue Date:1992
Rights Information:Copyright 1992 Wibert, Gregory James
Date Available in IDEALS:2011-05-07
Identifier in Online Catalog:AAI9236622
OCLC Identifier:(UMI)AAI9236622

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