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Previous reports indicated that glucose uptake in denervated muscle is resistant to insulin, while in non-weight-bearing (unweighted) muscle this effect of insulin is enhanced. To extend the comparison of these differences, insulin effects on amino acid uptake and protein metabolism were studied in soleus muscles subjected to denervation or unweighting. Denervated muscle showed insulin resistance of both 2-deoxy[1,2-3H]glucose and alpha-[methyl-3H]aminoisobutyric acid uptake whereas unweighted muscle showed an increased or normal response, respectively. Atrophy was greater in denervated than in unweighted muscle, apparently due to faster protein degradation. The stimulation of protein synthesis and the inhibition of protein degradation by insulin was generally less in denervated than in unweighted muscle. Since metabolic measurements in denervated-unweighted muscles did not differ from those in denervated-weight-bearing muscles, effects of denervation must be independent of leg posture.

Mechanisms of accelerated proteolysis were compared in denervated and unweighted (by tail-cast suspension) soleus muscles. In vitro and in vivo proteolysis were more rapid and lysosomal latency was lower in denervated than in unweighted muscle. In vitro, lysosomotropic agents (eg, chloroquine, methylamine) did not lessen the increase in proteolysis caused by unweighting, but abolished the difference in proteolysis between denervated and unweighted muscle. Leucine methylester, an indicator of lysosome fragility, lowered latency more in denervated than in unweighted muscle. 3-Methyladenine, which inhibits phagosome formation, increased latency similarly in all muscles tested. Mersalyl, a thiol protease inhibitor, and 8-(diethylamino)octyl-3,4,5-trimethoxybenzoate hydrochloride (TMB-8), which antagonizes sarcoplasmic reticulum release of Ca2+, reduced accelerated proteolysis caused by unweighting without diminishing the faster proteolysis due to denervation. Calcium ionophore (A23187) increased proteolysis more so in unweighted than control muscles whether or not Ca2+ was present. Different mechanisms of accelerated proteolysis were studied further by treating muscles in vivo for 24 hours with chloroquine or mersalyl. Chloroquine diminished atrophy of the denervated but not the unweighted muscle, whereas mersalyl prevented atrophy of the unweighted but not of the denervated muscle, both by inhibiting in vivo proteolysis. These results suggest that (1) atrophy of denervated, but not of unweighted, soleus muscle involves increased lysosomal proteolysis, possibly caused by greater permeability of the lysosome, and (2) cytosolic proteolysis is important in unweighting atrophy, involving some role of Ca2(+)-dependent proteolysis and/or thiol proteases.

Effects of stretching on muscle amino acids were tested in unloaded soleus by casting the foot in dorsiflexion on one limb of tail-casted, hindquarter-suspended rats. For comparison with unloading, amino acids also were measured in shortened extensor digitorum longus (EDL) in the same casted limb and in denervated leg muscles. Concentrations of tyrosine and glutamate were lower, while aspartate, ammonia, and the ratio of glutamine to glutamate were greater in the stretched than in the freely moving, unloaded soleus, but stretched did not differ from weight-bearing, control muscle. Therefore, stretching the soleus muscle prevented changes in certain amino acids due to unloading. Aspartate, ammonia, glutamine, and the ratio of glutamine to glutamate were lower in the shortened EDL than in the freely moving muscle of the contralateral limb, or in the control muscle. When denervated, these leg muscles also showed lower aspartate, ammonia, and ratio of glutamine to glutamate relative to innervated muscles. Since muscle shortening or denervation produced amino acid changes that mimicked the effects of unloading on the soleus, these responses must reflect the effect of muscle disuse. These data suggested that lower ammonia might cause the lower ratio of glutamine to glutamate with disuse. Because the fresh muscle energy charge, one factor which controls AMP deaminase, generally was not affected by disuse, altered deamination of glutamate via glutamate dehydrogenase may explain the variations in muscle ammonia.(ABSTRACT TRUNCATED AT 250 WORDS)

The time course of the response of carbohydrate metabolism to unloading was studied in the soleus muscle of rats subjected to tail-cast suspension. In the fresh soleus, just 12 hours of unloading led to higher concentrations of glycogen and lower activity ratios of both glycogen synthase and glycogen phosphorylase. These changes were still evident on day 3. This initial accumulation of glycogen was likely due to its decreased degradation in response to muscle disuse. Thereafter, the increased glycogen concentration apparently diminished the activity ratio of glycogen synthase, leading to a subsequent fall in the total glycogen content after day 1. After 24 hours of unloading, when no significant atrophy was detectable, there was no differential response to insulin for in vitro glucose metabolism. As reported for day 6 (reference 6), on day 3 the soleus atrophied significantly and displayed a greater sensitivity to insulin for most of these parameters compared to the weight-bearing control muscle. However, insulin sensitivity for glycogen synthesis was unchanged. These results showed that the increased sensitivity to insulin of the unloaded soleus is associated with the degree of muscle atrophy, likely due to an increased insulin binding capacity relative to muscle mass. This study also showed that insulin regulation of glucose uptake and of glycogen synthesis is affected differentially in the unloaded soleus muscle.

The influence of glucocorticoids on muscle glutamine production in starvation was studied by using cortisol-treated or non-cortisol-treated, starved, adrenalectomized rats. Administration of cortisol at physiological doses in vivo (1 mg/100 g body weight) to fasted, adrenalectomized rats increased the muscle ratio of glutamine/glutamate and the activity of glutamine synthetase after only 6 hours. Prior treatment of fasted, adrenalectomized animals with actinomycin D or proflavine abolished these increases by cortisol. Therefore, cortisol induces muscle glutamine synthetase, and this induction can be detected by changes in the fresh-muscle ratio of glutamine/glutamate. Using this ratio as a qualitative indicator of muscle glutamine synthesis, the role of glucocorticoids in modifying muscle glutamine production in starvation was studied. In fresh-frozen soleus, extensor digitorum longus, and diaphragm muscle, starvation led to greater ratios of glutamine/glutamate and higher levels of tyrosine, which are indicative of enhanced muscle protein turnover. These effects were not apparent in starved, adrenalectomized animals but were restored, at least partially, by administering a physiological dose of cortisol. Therefore, glucocorticoids seem essential for promoting muscle glutamine production in starvation probably by inducing the activity of glutamine synthetase.