Differential catabolism of muscle protein in Garden Warblers (Sylvia borin): flight and leg muscle act as a protein source during long-distance migration

Samples of flight and leg muscle tissue were taken from migratory garden warblers at three different stages of migration: (1) pre-flight: when birds face an extended flight phase within the next few days, (2) post-flight: when they have just completed an extended flight phase, and (3) recovery: when they are at the end of a stop-over period following an extended flight phase. The changes in body mass are closely related to the changes in flight (P<0.001) and leg muscle mass (P<0.001), suggesting that the skeletal muscles are involved in the protein metabolism associated with migratory flight. From pre- to post-flight, the flight and the leg muscle masses decrease by about 22%, but are restored to about 12% above the pre-flight masses during the recovery period. Biochemical analyses show that following flight a selective reduction occurred in the myofibrillar (contractile) component of the flight muscle (P<0.01). As this selective reduction accounts only for a minor part of the muscle mass changes, sarcoplasmic (non-contractile) and myofibrillar proteins of both the flight and leg muscle act as a protein source during long-distance migration. As a loss of leg muscle mass is additionally observed besides the loss in flight muscle mass, mass change seems not to be strictly associated with the mechanical power output requirements during flight. Whereas the specific content of sarcoplasmic proteins in the flight muscle is nearly twice as high as that in the leg muscle (P<0.001), the specific content of myofibrillar proteins differs only slightly (P < 0.05), being comparably low in both muscles. The ratio of non-contractile to contractile proteins in the flight muscle is one of the highest observed in muscles of a vertebrate.     

Effects of insulin-like growth factor-1/binding protein-3 complex on muscle atrophy in rats

Muscle atrophy and wasting is a serious problem that occurs in patients with prolonged debilitating illness, burn injury, spinal injury, as well as with space flight. Current treatment for such atrophy, which often relies on nutritional supplementation and physical therapy, is of limited value in preventing the muscle wasting that occurs. Considerable recent attention has focused on the use of anabolic growth factors such as insulin-like growth factor (IGF-1) in preventing muscle atrophy during limb disuse or with various catabolic conditions. However, potential side effects such as hypoglycemia appear to be limiting factors in the usefulness of IGF-1 for clinical treatment of muscle wasting conditions. The formulation of IGF-1 used in this study (IGF-1/BP3) is already bound to its endogenous-binding protein (BP3) and, as a result, has a greater specificity of action and significantly less hypoglycemic effect. Using a rat model of hind limb suspension (HLS) for 10 days, we induced marked muscle atrophy that was accompanied by enhanced muscle proteolysis and reduced muscle protein content. When HLS rats were treated with IGF-1/BP3 (50 mg/kg, b.i.d.), they retained greater body and muscle mass. Muscle protein degradation was significantly reduced and muscle protein content was preserved. The rate of protein synthesis, although somewhat reduced in HLS muscle, was not significantly elevated by IGF-1/BP3 treatment. Volume density of HLS-treated muscles were increased compared to untreated HLS rats and the actual number of fibers per area of muscle was likewise increased. The results of the current study suggest that IGF-1/BP3 might be useful for inhibiting muscle proteolysis in catabolic conditions and thus preserving muscle protein content and mass.     

Atrophy responses to muscle inactivity. I. Cellular markers of protein deficits.

The goal of this study was to use the model of spinal cord isolation (SI), which blocks nearly all neuromuscular activity while leaving the motoneuron muscle-fiber connections intact, to characterize the cellular processes linked to marked muscle atrophy. Rats randomly assigned to normal control and SI groups were studied at 0, 2, 4, 8, and 15 days after SI surgery. The slow soleus muscle atrophied by approximately 50%, with the greatest degree of loss occurring during the first 8 days. Throughout the SI duration, muscle protein concentration was maintained at the control level, whereas myofibrillar protein concentration steadily decreased between 4 and 15 days of SI, and this was associated with a 50% decrease in myosin heavy chain (MHC) normalized to total protein. Actin relative to the total protein was maintained at the control level. Marked reductions occurred in total RNA and DNA content and in total MHC and actin mRNA expressed relative to 18S ribosomal RNA. These findings suggest that two key factors contributing to the muscle atrophy in the SI model are 1). a reduction in ribosomal RNA that is consistent with a reduction in protein translational capacity, and 2). insufficient mRNA substrate for translating key sarcomeric proteins comprising the myofibril fraction, such as MHC and actin. In addition, the marked selective depletion of MHC protein in the muscles of SI rats suggests that this protein is more vulnerable to inactivity than actin protein. This selective MHC loss could be a major contributor for the previously reported loss in the functional integrity of SI muscles. Collectively, these data are consistent with the involvement of pretranslational and translational processes in muscle atrophy due to SI.     

Biochemical effects of mild iron deficiency and cold acclimatization on rat skeletal muscle

Most of the previous studies on the effects of iron deficiency on skeletal muscle respiratory capacity and work performance have been investigated in severe or moderate iron-deficiency anemia. We report here that even in mild iron deficiency where the hemoglobin concentration was 10 g/dl and the iron stores in livers and spleen were not completely depleted, a marked reduction in succinate dehydrogenase was observed in skeletal muscles but not in heart. Similarly, cytochrome oxidase activities were reduced. Although no significant change in glycerophosphate dehydrogenase was detected in the iron-deficient rats, exposure to cold in this group greatly reduced this enzyme activity. As cold acclimatization accelerates marrow erythropoiesis (20) which in turn, demands more iron, it seems that in the iron-insufficient state, this iron demand for marrow activity may persist at the expense of the tissue iron pool, resulting in a marked reduction in glycerophosphate dehydrogenase activities. Since succinate dehydrogenase plays a significant role in the impairment of mitochondrial function and early fatigue of iron-deficient muscle (11), the present study shows that even in mild iron deficiency, some loss of muscle functions could result as succinate dehydrogenase activities were greatly reduced.     

Oogenesis-flight syndrome in crickets: age-dependent egg production, flight performance, and biochemical composition of the flight muscles in adult female Gryllus bimaculatus

Age-dependent changes in flight performance, biochemical composition of flight muscles, and fresh mass of the flight muscles and ovaries were analysed in adult female two-spotted crickets, Gryllus bimaculatus. After the final moult the flight muscle mass increased significantly to a maximum at days 2 and 3. On day 2 the highest flight activity was also observed. Between days 2 and 3 the ovary weight started to rapidly increase due to vitellogenic egg growth, which continued at a high rate until day 10. With the onset of ovarial growth, flight performance decreased and the flight muscles started to histolyse. A high correlation between flight muscle mass and the content of protein, lipid, glycogen, and free carbohydrate in the flight muscle indicated that energy-rich substrates from the degrading flight muscles were used to fuel oogenesis, although flight muscle histolysis can provide only a small fraction of the substrates needed for egg production. In general, there was a clear trade-off between egg production and flight ability. Surprisingly, however, some females possessed well-developed ovaries but displayed no signs of flight muscle histolysis. This observation was corroborated by flight experiments which revealed that, although most flying females had small ovaries, some of them carried an appreciable amount of mature eggs, and thus, somehow managed to evade the oogenesis-flight syndrome.     

Effects of flight training on flight speed and substrate utilization in locusts

ABSTRACT. Regular flight exercise of adult male Locusta migratoria migratorioides (R and F) accelerated the development of maximum flight speed and disrupted the development of the typical pattern of change of flight speed exhibited when normal (untrained) adult male laboratory locusts are flown on roundabouts. Thus, while untrained mature locusts fly fast initially and then slow to a steady cruising speed after 20 min, trained locusts flew at a relatively constant speed throughout a 60-min test period. Flight training also led to a marked reduction in the size of the fat body and the flight muscles, but flight muscle ultrastructural development was not affected. Regular flight exercise had no long-term effect on haemolymph carbohydrate concentration but lipid levels were significantly depressed.     

The myosin converter domain modulates muscle performance

Myosin is the molecular motor that powers muscle contraction as a result of conformational changes during its mechanochemical cycle. We demonstrate that the converter, a compact structural domain that differs in sequence between Drosophila melanogaster myosin isoforms, dramatically influences the kinetic properties of myosin and muscle fibres. Transgenic replacement of the converter in the fast indirect flight muscle with the converter from an embryonic muscle slowed muscle kinetics, forcing a compensatory reduction in wing beat frequency to sustain flight. Conversely, replacing the embryonic converter with the flight muscle converter sped up muscle kinetics and increased maximum power twofold, compared to flight muscles expressing the embryonic myosin isoform. The substitutions also dramatically influenced in vitro actin sliding velocity, suggesting that the converter modulates a rate-limiting step preceding cross-bridge detachment. Our integrative analysis demonstrates that isoform-specific differences in the myosin converter allow different muscle types to meet their specific locomotion demands.     

Effect of sciatic nerve crush on enzyme activities of skeletal muscles of the rat.

Weight loss and reduction in specific activity of cytochrome oxidase of both red (soleus) and white muscles (gastrocnemius and plantaris) of the rat was greatest 5 days after sciatic nerve crush (p less than 0.001) and then became minimal. In neither was there a significant, concomitant loss of protein. By 19 days, the specific activity of cytochrome oxidase was the same in both muscle types. The specific activity of lactate dehydrogenase was reduced significantly (p less than 0.001) in the white muscles, to a value approaching that of the red soleus by 19 days postoperatively, but remained unaltered in the soleus. Nerve crush is proposed as a model experimental system for studying neural regulation of skeletal muscle metabolism.     

Responses of muscle sympathetic nerve activity to static handgrip exercise after 14 days of exposure to simulated microgravity

Decrease in muscle perfusion affects on cardiovascular response to exercise. Muscle hypoperfusion enhances the increase in blood pressure responses to exercise. Muscle perfusion depends not only on central blood pressure but also how fit the active muscle is above or below the heart level; muscle perfusion decreases as arm is elevated. Static exercise increases muscle sympathetic nerve activity (MSNA) innervating vessels in non-active muscles. The exercise-induced increase in MSNA is mainly mediated by stimulating chemosensitive muscle afferents in active muscles. However, the effect of arm elevation on MSNA during forearm exercise is not examined. On the other hand, space flight and simulated microgravity exposure causes reduction in muscle blood flow, suggesting chronic muscle hypoperfused condition during simulated microgravity. Therefore, there is a possibility that arm elevation after microgravity exposure alters MSNA responsiveness during exercise. However, arm elevation effect after exposure to simulated microgravity is not examined.     

The physiological costs of flight capability in wing-dimorphic crickets

Nutritional indices, triglyceride levels and flight muscle developmental profiles were compared between long-winged (LW) and short-winged (SW; flightless) morphs of the cricketsGryllus rubens Scudder andG. firmus Scudder. This was done to identify potential physiological costs of flight capability in adults. The LW morph of each species converted a lower proportion of assimilated nutrients into biomass (reduced ECD) than did the SW morph. This documents increased respiratory metabolism in the LW morph. Triglyceride concentration was higher in LW vs. SW adults. This suggests that the elevated respiration in the LW morph may be at least partially due to the increased biosynthesis of this high energy substance. Preliminary data indicate higher respiration rates of LW functional vs. SW vestigial flight muscles. Collectively, these data suggest that the energetic cost of flight capability in adults results from biosynthesis of triglyceride flight fuel and flight muscle maintenance but not flight muscle growth. No flight muscle growth was observed in adults.     

Myosin heavy chain isoforms regulate muscle function but not myofibril assembly.

Myosin heavy chain (MHC) is the motor protein of muscle thick filaments. Most organisms produce many muscle MHC isoforms with temporally and spatially regulated expression patterns. This suggests that isoforms of MHC have different characteristics necessary for defining specific muscle properties. The single Drosophila muscle Mhc gene yields various isoforms as a result of alternative RNA splicing. To determine whether this multiplicity of MHC isoforms is critical to myofibril assembly and function, we introduced a gene encoding only an embryonic MHC into Drosophila melanogaster. The embryonic transgene acts in a dominant antimorphic manner to disrupt flight muscle function. The transgene was genetically crossed into an MHC null background. Unexpectedly, transformed flies expressing only the embryonic isoform are viable. Adult muscles containing embryonic MHC assemble normally, indicating that the isoform of MHC does not determine the dramatic ultrastructural variation among different muscle types. However, transformed flies are flightless and show reduced jumping and mating ability. Their indirect flight muscle myofibrils progressively deteriorate. Our data show that the proper MHC isoform is critical for specialized muscle function and myofibril stability.     

Effect of glucocorticoid on protein and creatine content of inactivated muscle of rats

In denervation, there was loss of protein in gastrocnemius muscles and this loss of was more in prednisolone treated animals. There was significant change of protein loss between tenotomy and tenotomy with prednisolone treatment. The reduction of protein in denervation and denervation with prednisolone treatment were also highly significant. Significant loss of muscle creatine was observed in denervation with prednisolone treatment. It was about 50% of the normal control group and about 40% when compared to other limb. In denervation alone, the creatine loss was about 24%. In tenotomy and in tenotomy with prednisolone treatment, the loss of creatine was also significantly high. All these figures regarding the reduction of muscle creatine in different experiments were highly significant. The reduction of muscle weight, protein and creatine content of muscle in denervation were due to inactivation of the muscle and due to trophic changes caused by loss of motor supply to the muscle. But in tenotomy, the reductions were only due to inactivation.     

Nutrition‐ and sex‐dependent utilization of body resources in relation to reproduction in a scorpionfly

Reproduction often comes at a cost of a reduction in body functions. In order to enhance their reproductive output, some insect species degenerate their thoracic muscles, typically resulting in reduced flight ability. From a life‐history trade‐off perspective, we expect the importance of body resource utilization to be amplified both with increased reproductive expenditure and with increased resource limitation. In this study, we measured age‐related changes in thorax weight, as a measure of flight muscle size, during a major part of the adult lifespan in males and females of the scorpionfly Panorpa vulgaris. The aim of the study was twofold: first to investigate whether scorpionflies have the potential to degenerate their flight muscles; second, and more importantly, to determine whether the magnitude of flight muscle degeneration is a plastic response in relation to resource availability, and if it differs between the sexes. The results clearly demonstrate that food availability does influence investment in flight muscle development. The build‐up of the thoracic muscles was strongly influenced by nutrient availability. Furthermore, the age‐related decrease in thorax weight was significantly different for males and females. Only females showed a strong age‐dependent decrease in thorax weight, indicative of muscle degeneration, yet no difference between food treatments was detected. For males, there was no significant directional change in thorax weight. Nevertheless, with increasing age, the difference in thorax weight between food treatments increased significantly. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2011, 102 , 199–207.     

Transformation of Drosophila melanogaster with the wild-type myosin heavy-chain gene: rescue of mutant phenotypes and analysis of defects caused by overexpression

We have transformed Drosophila melanogaster with a genomic construct containing the entire wild-type myosin heavy-chain gene, Mhc, together with approximately 9 kb of flanking DNA on each side. Three independent lines stably express myosin heavy-chain protein (MHC) at approximately wild-type levels. The MHC produced is functional since it rescues the mutant phenotypes of a number of different Mhc alleles: the amorphic allele Mhc1, the indirect flight muscle and jump muscle-specific amorphic allele Mhc10, and the hypomorphic allele Mhc2. We show that the Mhc2 mutation is due to the insertion of a transposable element in an intron of Mhc. Since a reduction in MHC in the indirect flight muscles alters the myosin/actin protein ratio and results in myofibrillar defects, we determined the effects of an increase in the effective copy number of Mhc. The presence of four copies of Mhc results in overabundance of the protein and a flightless phenotype. Electron microscopy reveals concomitant defects in the indirect flight muscles, with excess thick filaments at the periphery of the myofibrils. Further increases in copy number are lethal. These results demonstrate the usefulness and potential of the transgenic system to study myosin function in Drosophila. They also show that overexpression of wild-type protein in muscle may disrupt the function of not only the indirect flight but also other muscles of the organism.     

Flight muscles polymorphism in a flightless bug, Pyrrhocoris apterus (L.): developmental pattern, biochemical profile and endocrine control

The flightless bug Pyrrhocoris apterus (L.) is polymorphic for both wing length and flight muscle development. The developed flight muscles of macropterous adults of both sexes first enlarge their volume during the first 5 days after adult emergence, but are then histolyzed in all males and females older than 10 and 14 days, respectively. The flight muscles of brachypterous adult males and females are underdeveloped due to their arrested growth. The total protein content of histolyzed dorsolongitudinal flight muscles from 21-day-old macropterous adults of both sexes is lower than that of developed dorsolongitudinal flight muscles in 5-10-days-old macropterous bugs, but substantially higher than the protein content of underdeveloped dorsolongitudinal flight muscles from adult brachypters. Histolyzed dorsolongitudinal flight muscles differ from the developed ones by decreased quantities of 18 electrophoretically separated proteins. Histolysis of developed dorsolongitudinal flight muscles is accompanied by significant decreases in citrate synthase, glyceraldehyde-3-phosphate dehydrogenase and β-hydroxyacyl-CoA dehydrogenase enzyme activities and an increase in alanine aminotransferase activity, and can be precociously induced by application of a juvenile hormone analogue. This is the first report of flight muscle polymorphism, histolysis of developed flight muscles and its endocrine control in insects displaying non-functional wing polymorphism.     

Effect of ecdysterone and juvenoid on the developmental involution of flight muscles in Acheta domestica

Morphogenesis and degeneration of the flight muscles in Acheta domestica was studied. The dorso-longitudinal flight muscles (DLMs) degenerate during the fourth day after adult ecdysis and the dorso-ventral flight muscles (DVMs) on the fifteenth day. In the presence of an intact innervation the degeneration of the DLMs can be retarded for 2 days by the injection of ecdysterone into very young adults. This retardation may also result in hypertrophy of the muscle fibres. The injection of ecdysterone, even in high doses, did not affect the flight muscle remnants. No notable changes have been found in the degeneration of DLMs by ovarectomy. Thus, the degeneration of flight muscles and the development of ovaries appear to be independent processes.The DLMs are homogeneous in fibre pattern in respect to succinic dehydrogenase, an important oxidative enzyme, and to ATPase activity, but the muscle fibres do not show any phosphorylase activity.     

The effects of space flight during gestation on rat uterine smooth muscle

Pregnant rats were flown on a NASA space shuttle during days 11-20 of gestation and the effects of pregnancy maintenance and uterine myometrial smooth muscle were evaluated. There was no effect of space flight on fetal mass in rats examined at recovery on day 20 of gestation (G-20). Rats allowed to go to term delivered vaginally at days 22-23, but space flight significantly decreased pup mass at birth. Space flight did not alter myometrial smooth muscle volume at G-20 and postpartum when compared to flight delayed synchronous controls, but flight animals showed a 37% decrease in myometrial smooth muscle volume between G-20 and postpartum. In contrast, smooth muscle volume of the myometrium of controls, postpartum, was similar to G-20. We conclude that myometrial smooth muscle, which hypertrophies during space flight similar to controls, reacclimates acutely to earth's gravity between G-20 and parturition with a dramatic reduction in volume.     

The use of muscle protein for egg production in the Zebra Finch Taeniopygia guttata

Pectoral muscle can be an important source of protein for birds. During egg formation Zebra Finches Taeniopygia guttata are able to compensate for nutritional inadequacies in their diet by utilization of the protein in their flight muscles. This analysis of flight muscle sarcoplasm supported earlier observations of protein depletion during egg production. However, SDS gel electrophoresis of the sarcoplasm produced no evidence to support a previous suggestion of the existence of a high molecular weight storage protein, and it is thought that the original observation may have arisen as an artefact of experimental methodology. During laying, protein removal from the sarcoplasm occurred over a range of different proteins and was not confined to any one specific protein band. Additionally, the protein band most reduced over the course of laying did not contain elevated levels of the amino acids most limiting to egg production. These results indicate that during laying, flight muscle sarcoplasm contributes towards the nutrient requirements of egg production from general protein reserves, rather than from a specific storage protein containing elevated levels of limiting amino acids.     

Increase in rat soleus myotendinous interface after a 14-d spaceflight

Myotenclinous junctions (MTJs) transmit contractile force from skeletal muscles to tendons. The effects of a 14-d spaceflight on MTJ were studied in the soleus muscle of male adult Sprague Dawley rats by transmission electron microscopy and histomorphometric techniques. We showed that the length of the junctional membrane relative to the muscle fiber diameter increased by 58% after 14 d of spaceflight. This increase accompanies morphological changes at MTJs. The flight MTJs appeared more shredded. The ends of the muscle fibers exhibited T tubule dilatation, swollen mitochondria, Z-disk streaming, loss of myofilaments, a thinning down of subplasmalemmal densities, multivesicular bodies and signs of junctional membrane and basal lamina remodelling. The ultrastructural observations suggest that the increase in myotendinous interface could result from the extracellular matrix spreading into remodelling muscle fiber, whereas the constraints related to unloading were reduced by spaceflight conditions.     

An improved technique for estimating pectoral muscle protein condition from body measurements of live gulls

Body measurements, which could be taken from live birds, were used to estimate total pectoral muscle protein in Lesser Black-backed Gulls Larus fuscus. The maximum cross-sectional area of the flight muscles was measured from the profile of the muscle surface over the keel, and this was used in conjunction with the length of the flight muscle to estimate muscle volume. The estimate of muscle volume was then used with fresh body weight to estimate total flight muscle protein. A highly significant correlation was found between the estimated values and actual pectoral muscle protein mass determined by carcass analysis. The model developed from the source group was then validated using a second independent sample, in which flight muscle protein was estimated from the model. Carcass analysis again demonstrated a good correlation between estimated and actual total protein. Different methods of controlling for body-size to calculate protein condition from measures of total protein were considered. The technique described here provides a simple and reliable method of estimating pectoral muscle protein condition in live gulls which could be applied to studies of body condition in other species.