Co-existence of nonmuscle-specific and cardiac muscle-specific myosin in myofibrils of cultured adult cardiac muscle cells.

Immunofluorescence microscopy revealed that nonmuscle myosin and cardiac muscle-specific myosin were present in the myofibrils of cultured adult cardiac muscle cells at different stages of redifferentiation including fully redifferentiated cells. Nonmuscle myosin and cardiac muscle-specific myosin were observed in amorphous or fibroamorphous form predominantly in unspread and partially spread cardiac myocytes. Nonmuscle myosin and cardiac muscle-specific myosin were present in the same striated myofibrils of the same fully redifferentiated cells. Nonmuscle myosin was localized in the Z-lines of sarcomeres of myofibrils. In double stained cells, alpha-actinin was colocalized with nonmuscle myosin in the Z-lines. The in vivo adult cardiac muscle cells contained nonmuscle myosin and cardiac muscle-specific myosin in the same myofibrils of the same cells with the localization of nonmuscle myosin in Z-lines and intercalated discs. It is evident that the nonmuscle myosin is an integral part of the sarcomeric structure of myofibrils in cardiac muscle cells.

Capillary network patterning during angiogenesis.

1. Although much is known of signalling events associated with angiogenesis, starting with a sprout, less is known of how an intact microvascular network is correctly formed following the initiation of sprouting. The aim of the present report is to evaluate some of the factors that could be involved in directing the patterning of the resulting microvascular bed. 2. Hypothetical 'patterning rules' are discussed and considered in the context of data obtained from studies of angiogenesis. The guidelines suggest that both tissue- and stimulus-specific regulators are involved in directing patterning. 3. Examples of 'patterned' and 'non-patterned' angiogenesis in rat mesentery are given, with evidence to support stimulus-related differences in angiogenesis patterns. 4. Skeletal muscle angiogenesis follows metabolic patterns within the muscle, as discerned through traditional histochemical evaluation. However, further examination, using confocal microscopy and new patterning assays, indicates that branching patterns differ according to stimulus. Patterning guidelines within the tissue may be important in preventing the growth of sinusoidal vessels. 5. The concept of 'angiotypes' is proposed to direct future studies towards understanding how therapeutic angiogenesis or angiostasis applications can be used to obtain correctly patterned microvascular networks. Angiogenesis via 'branching' compared with 'non-branching' angiotypes will involve different signalling pathways. Both sprouting and dividing of capillaries are found in the 'branching' angiotype, with each pattern having several subtypes that would be expected to be mediated via different signalling mechanisms.

Localization of the ANG II type 2 receptor in the microcirculation of skeletal muscle.

Only functional studies have suggested the presence of the ANG II type 2 (AT2) receptor in the microcirculation. To determine the distribution of this receptor in the rat skeletal muscle microcirculation, a polyclonal rabbit anti-rat antiserum was developed and used for immunohistochemistry and Western blot analysis. The antiserum was prepared against a highly specific and antigenic AT2-receptor synthetic peptide and was validated by competition and sensitivity assays. Western blot analysis demonstrated a prominent, single band at approximately 40 kDa in cremaster and soleus muscle. Immunohistochemical analysis revealed a wide distribution of AT2 receptors throughout the skeletal muscle microcirculation in large and small microvessels. Microanatomic studies displayed an endothelial localization of the AT2 receptor, whereas dual labeling with smooth muscle alpha-actin also showed colocalization of the AT2 receptor with vascular smooth muscle cells. Other cells associated with the microvessels also stained positive for AT2 receptors. Briefly, this study confirms previous functional data and localizes the AT2 receptor to the microcirculation. These studies demonstrate that the AT2 receptor is present on a variety of vascular cell types and that it is situated in a fashion that would allow it to directly oppose ANG II type 1 receptor actions.

Lack of involvement of basic fibroblast growth factor (FGF-2) in capillary growth in skeletal muscles exposed to long-term contractile activity.

Fibroblast growth factors (FGFs) are involved in stimulation of angiogenesis in tumors and other pathological circumstances. Increased activity of normal skeletal muscles resulting from chronic electrical stimulation is a very potent stimulus for capillary growth but a relationship between the initiation of this angiogenesis and the involvement of autocrine growth factors has yet to be established. Although FGF expression has been reported in muscles stimulated for 3 weeks, capillary growth is underway significantly earlier, beginning around 3 days. The present experiments have therefore studied the possible involvement of basic fibroblast growth factor (FGF-2) in stimulated rat fast skeletal muscles prior to, and coincident with, capillary growth. Muscle contractions were induced via electrodes implanted in the vicinity of the peroneal nerve and maintained for 8h/day for 2, 4 or 7 days. Capillary/fiber ratio (C/F), based on staining of capillary endothelium for alkaline phosphatase, was not changed in either extensor digitorum longus (EDL) or tibialis anterior (TA) after 2 days stimulation, but increased in TA stimulated for 4 days and in both muscles after 7 days. The expression of mRNA for FGF-2, detected by ribonuclease protection assay, was decreased in all stimulated muscles compared with control or contralateral muscles; immunohistochemistry showed FGF-2 gene product in nerves and larger blood vessels but not in capillaries. There was no evidence from immunohistochemistry for up-regulation of receptors flg and bek for FGF-2. The presence of FGF-2, flg and bek in arterioles may indicate a possible role for FGF-2 in the regulation of blood flow since we have previously shown it to be a dilator of small arterioles. However, based on the lack of correlation between changes in capillary density and the expression of mRNA and protein for FGF-2 and its receptors, it is unlikely that it is directly linked with the initiation of angiogenesis resulting from chronic activity in skeletal muscles.

In vivo angiogenesis in adult rat skeletal muscle: early changes in capillary network architecture and ultrastructure.

The individual structural stages in capillary growth have been identified during development and under pathological circumstances in adults (wound healing, tumors), but there are no data to indicate whether these steps are similar when angiogenesis is induced in a fully differentiated microvascular bed in normal, uninjured adult skeletal muscle. In this study changes in capillary ultrastructure were correlated with capillary density and network morphology to elucidate the sequelae of angiogenesis in adult rat extensor digitorum longus (EDL) muscle whose activity was increased by stimulation at 10 Hz (8 h/day). This resulted in an increased capillary/fiber (C/F) ratio (based on staining for alkaline phosphatase) after 4 days; by 7 days C/F ratio was increased further, by approximately 50%. The ultrastructure of capillary endothelium in both the EDL and extensor hallucis proprius (EHP) was similar to control muscles after 2 days of stimulation, whereas endothelial cells in some capillaries in muscle stimulated for 4 days revealed signs of metabolic activation such as proliferation of organelles (Golgi apparatus, endoplasmic reticulum, ribosomes and mitochondria) and fewer pinocytic vesicles. Luminal surfaces were often irregular with numerous pseudopodial processes. Basement membranes were always present but amorphous regions were observed, particularly near pericyte processes. Unusually small capillary profiles, with either a slit-like lumen or with cisternae but no lumen, probably represented capillary sprouts. The interstitium contained increased collagenous and granular extracellular matrix surrounding capillaries, and numerous activated fibroblasts which were closely apposed to many capillaries. Capillary growth in EHP was also evaluated by confocal microscopy using whole mounts. The complex pattern of vessels underwent remodelling between 2 and 7 days of stimulation, resulting in more tortuous capillaries with numerous sprouts and loops. These combined observations suggest that angiogenesis may occur by a combination of sprouting, intussusceptive growth and elongation; also, that activation of endothelial cells occurs at the same time as disturbance of basement membranes during the earliest phase of growth and remodelling of the capillary bed. These changes are postulated to occur in connection with increased shear stress and/or capillary wall tension, which have been demonstrated previously.

Rapid microvessel rarefaction with elevated salt intake and reduced renal mass hypertension in rats.

To identify the sequence of events associated with the development of reduced vessel density (rarefaction) in hypertension, microvessel density and ultrastructure were assessed in the cremaster muscle of rats subjected to a 75% surgical reduction of renal mass and normotensive sham-operated control rats. Rats with reduced renal mass (RRM rats) and sham-operated rats were then maintained on either a high salt (4.0% NaCl) or a low salt (0.4% NaCl) diet for 3 days. Acute exposure to the high salt diet significantly increased mean arterial pressure in RRM rats but did not affect sham-operated control rats. Quantitative fluorescence microscopy of cremaster muscle whole mounts using rhodamine-labeled Griffonia simplicifolia I lectin revealed substantial rarefaction of microvessels in both RRM hypertensive rats and normotensive sham-operated rats on a high salt diet relative to corresponding control rats on a low salt diet. Confocal microscopy revealed a heterogeneous distribution of microvessels in RRM rats on a high salt diet, with some areas largely devoid of vessels. RRM and sham-operated rats on a high salt diet both exhibited changes in arteriolar ultrastructure, which included a loss of basement membranes and a dissociation of the endothelial and smooth muscle components of the vascular wall, resulting in a loss of vessel integrity. These observations demonstrate that a rapid loss of microvessels can occur not only in rats with RRM hypertension but also in normotensive rats on a high salt diet. This loss of microvessels results from structural alterations, which differ from the degenerative processes associated with microvascular rarefaction in rats with chronic RRM hypertension.

Regional differences in spontaneously occurring angiogenesis in the adult rat mesentery.

Despite intensive study in the area of angiogenesis, relatively little is known about normal angiogenesis in adult animals. Preliminary studies using the Griffonia simplicifolia I (GSI) lectin as a microvascular marker indicated that capillary sprouting occurs in the clear "windows" of normal intact adult rat mesentery. The purpose of this study was to determine whether angiogenesis occurs uniformly within the mesenteric windows and whether maturational age affects the extent of angiogenesis in the absence of any experimental or pathological perturbation. Four groups of female Sprague-Dawley rats were used, "weanling" (4-5 weeks), "juvenile" (6-8 weeks), "young adult" (10-13 weeks), and "adult" (16-20 weeks). Microvessels sprouting into proximal and distal windows were delineated in whole mounts by use of fluorescent derivative of the GSI lectin. Microvascular sprouting, indicating angiogenesis, was found in all age groups, but was most frequent in the windows sampled from the distal region of the small intestine when compared with those from the proximal region. The mean number of microvessels per sample site was significantly higher in the distal windows of adults than in the weanling or juvenile rats. Angiogenesis was found to occur asymmetrically within the individual windows in the two adult groups, with significantly more angiogenesis on the intestinal side compared to that along the portal vessels. We conclude that there is an age-related increase in angiogenesis into the mesenteric windows, and that the intestinal side is more prone to spontaneous angiogenesis than is the portal side.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression of muscle capillary alkaline phosphatase is affected by hypoxia.

Hypoxia stimulates angiogenesis in some microvascular beds, but no clear angiogenic effect of hypoxia has yet been demonstrated in adult skeletal muscle. In this study the distribution of alkaline phosphatase (APase) was compared with a novel microvascular marker, Griffonia simplicifolia I (GSI), to determine whether the respective markers were expressed by muscle capillaries during hypoxic conditions and to probe for the presence or absence of angiogenesis in response to short-term hypoxia. Mice were exposed to normobaric 8% oxygen for 7 or 21 days. Capillary density in the red and white areas of the gastrocnemius muscle was determined with the use of a double-labeling procedure for both APase and fluorescently tagged GSI. Little change in capillary density was found. Focal reductions in APase activity were observed within 1 wk of hypoxia, but no changes were observed in GSI binding. In controls, 74 and 92% of red and white muscle capillaries, respectively, were APase positive. This percentage declined to 60% in red and 43% in white muscle after 21 days of hypoxia. The results indicate that APase expression is labile under certain conditions and warrant a cautious approach to using the enzyme as a marker. Binding of the GSI lectin to muscle capillaries appeared to be unchanged by the exposure to hypoxia, indicating stability of this marker system. No significant change in the number of capillaries around individual muscle fibers was evident at 21 days when GSI was used to detect capillaries. These results confirm the absence of hypoxia-induced angiogenesis in muscle capillaries during the time period studied.

Microvessel changes in hypertension measured by Griffonia simplicifolia I lectin.

Commonly used methods for assessing reductions in microvascular density (rarefaction) in hypertension detect only perfused microvessels. In the present study, samples of cremaster and spinotrapezius muscles were taken from rats with chronic (4-week) reduced renal mass hypertension and normotensive sham-operated control rats, as well as from 12-week-old spontaneously hypertensive rats and their normotensive Wistar-Kyoto control strain. Mean arterial pressure was 149 +/- 8 mm Hg in the rats with reduced renal mass hypertension, 114 +/- 7 mm Hg in sham-operated rats, 177 +/- 9 mm Hg in spontaneously hypertensive rats, and 95 +/- 4 mm Hg in Wistar-Kyoto rats. Muscle samples were incubated with rhodamine-labeled Griffonia simplicifolia I lectin, which identifies both perfused and nonperfused microvessels. Microvascular density was assessed by counting intersections with a 20-microns grid. Microvessel density was significantly reduced in cremaster muscles of both spontaneously hypertensive and reduced renal mass hypertensive rats, and in the spinotrapezius muscle of spontaneously hypertensive rats, compared with their respective normotensive controls. Further studies in the reduced renal mass rats on low salt diets indicated that lectin binding was also decreased as salt intake was increased, independent of blood pressure. This change was not due to an alteration in lectin-binding affinity. These studies indicate that lectin binding can be a useful tool for assessing microvessel density that does not depend on the perfusion state of the vessels and that rarefaction due to hypertension is not evenly distributed in all vascular beds. These results also provide evidence that dietary salt intake alone can influence microvessel density, as measured by the lectin technique.

Postnatal changes in capillary density of rat sternomastoid muscle.

The changes in muscle capillarity during postnatal growth have been difficult to study using standard histochemical methods. This laboratory has proposed the use of a lectin, Griffonia simplicifolia I (GSI), as a histochemical marker that may be appropriate for developing muscle. The purpose of the study was to compare the capillary densities (capillaries per muscle fiber) determined by the GSI lectin method, alkaline phosphatase-periodic acid-Schiff (APase-PAS) staining method, and by direct counting from 1-micron plastic sections. Sternomastoid muscles from 10-day or 6-wk-old rats were used. Results from the 10-day rats showed that the GSI method and the 1-micron method gave comparable results (1.44, 1.58 capillaries per fiber), which were significantly higher than that with APase-PAS (0.56). Capillary densities in the white region of the sternomastoid from 6-wk-old rats were identical (2.80) using both the GSI and APase-PAS methods. In contrast, the GSI method yielded significantly higher capillary densities in the red region than did the APase-PAS method (4.80 vs. 3.83). These results indicate that the GSI method for visualizing capillaries is a sensitive method for visualizing capillaries in muscle during the postnatal and juvenile growth period.

Griffonia simplicifolia I: fluorescent tracer for microcirculatory vessels in nonperfused thin muscles and sectioned muscle.

Previous studies on mice have revealed that the Griffonia simplicifolia I (GSI) lectin selectively binds to capillaries in a number of microvascular beds. These observations suggest that the lectin might be a suitable microvascular marker for physiological studies of skeletal muscle, particularly when fluorescent visualization of vessels is desired independently of their perfusion status. Since species and strain heterogeneity has been demonstrated for certain lectins associated with the microcirculatory vessels, lectin binding was studied in a number of muscles taken from the major species of mammals used for experimental purposes. Staining of cryostat sections confirmed the utility of GSI as a marker for capillaries from muscle of mice, rats, hamsters, rabbits, dogs, and monkeys. Differential staining of arterioles and veins was revealed by double labeling with GSI and antisera to Factor VIII-related antigen. Double labeling for GSI binding and alkaline phosphatase activity revealed that the GSI method detects many more capillaries and terminal arterioles than does the alkaline phosphatase method. GSI binding to unfixed whole mounts of thin skeletal muscles (hamster cheek pouch, mouse diaphragm, and rat cremaster) was studied to determine whether the GSI lectin would be a suitable marker for intravital studies. An extensive microvascular bed, including terminal arterioles, venules, and capillaries, was revealed which could be visualized in the complete absence of perfusion with fluorescent markers. These observations suggest that the GSI lectin may be extremely useful as a probe for the microcirculation of skeletal muscle in many types of physiological experiments.

Formation of acetylcholine receptor clusters in mammalian sternohyoid muscle regenerating in the absence of nerves.

When the sternohyoid muscle from the rat is grafted, the original muscle fibers, including the membranes at the neuromuscular junction, degenerate irreversibly. New muscle fibers regenerate inside of the basal laminae remaining from the original muscle fibers. In this study rhodamine-alpha-bungarotoxin and electron microscopy have been used to demonstrate that acetylcholine receptor (AchR) clusters and synaptic folds are restored to the regenerating myotubes even when innervation to the grafts is prevented. The AchR clusters and synaptic folds colocalized with acetylcholinesterase that persisted at the original synaptic basal lamina. The AchR clusters were not restored if the original innervation band was removed from the muscle at the time of grafting. Lengths of the AchR clusters were measured in animals ranging in weight from 50 to 700 g. The lengths of clusters in the grafts were proportional to the lengths of those in the preoperative controls, suggesting that quantitative morphogenetic information persists through the period of degeneration and regeneration. However, the distribution of the AchRs within the clusters differed slightly from controls. Extrajunctional AchR clusters were present initially, but later disappeared. The sizes of these clusters were unrelated to the sizes of the junctional AchR clusters. This study demonstrates that morphogenetic cues persist within the region of the original motor and plate, possibly associated with the synaptic basal lamina.

Growth of regenerating skeletal muscle fibers in cats.

Extensor digitorum longus (EDL) muscles of cats were grafted heterotopically or orthotopically, and either with or without prior denervation. The autografted muscles were studied at times from 4 to 518 days after grafting. Muscle weight, fiber cross-sectional area, and ultrastructural, histochemical, and biochemical characteristics of regenerating muscle fibers were determined. Prior denervation reduced the mass of muscle at the time of grafting, but had no significant effect on the characteristics of the regenerating muscles. Orthotopic and heterotopic autografts achieved similar recovery of structure. Mean fiber area reached control values. Differentiation into fiber types occurred, but compared to control muscles, autografts had fewer Type I and Type IIA fibers. Electron microscopic analysis of regenerating muscle fibers revealed centrally located nuclei, but otherwise normal ultrastructure. The bimodal distribution of Z-band width was consistent with differentiation into fiber types. Mean data of some morphological variables did not stabilize.

Development and innervation of soleplates in the freely grafted extensor digitorum longus (EDL) muscle in the rat.

The ultrastructural events in the establishment of the neuromuscular junction of the freely grafted extensor digitorum longus (EDL) muscle of the rat were studied 1-120 days after grafting. The original axons and muscle fibers, including soleplates, degenerated during the first few days, but Schwann cells and basal laminae persisted. Myofibers regenerated within the original basal laminae. Indentations of the sarcolemma, termed "presumptive synaptic clefts" (PSC), were found on myotubes from 7-day grafts. Schwann cells and residual acetylcholinesterase were invariably associated with the PSC, suggesting that the PSC developed at the site of the original soleplate. Nerves entered the grafts 10 days postoperatively and contacted the PSC of the regenerating muscle fibers on the 18-20th day. The secondary synaptic clefts of these "reconstructed" soleplates extended far beyond the subaxonal region. A second type of soleplate appeared on the 18-20th day. These soleplates were similar to those found in embryonic muscle and were considered to have been induced to form "de novo" by the presence of the nerves. When grafts were placed in permanently denervated limbs the "reconstructed" soleplates appeared, but the "de novo" type did not. These results show that information directing the morphogenesis and innervation of the soleplate persists after the original muscle fibers and axons of a graft degenerate and regenerate.

Revascularization of the freely grafted extensor digitorum longus muscle in the rat.

The process of revascularization of free grafts of the extensor digitorum longus muscle in rats has been studied by gross, histological and electron microscopic methods. During the first day after transplantation the muscle is entirely avascular, and it consists of a thin peripheral zone of surviving muscle fibers and a large central area of ischemic muscle. The original blood vessels of the graft undergo a sequence of intrinsic and cell-mediated destruction. Scattered sinusoidal vessels begin to grow into the graft starting on the second day, and ingrowing blood vessels progressively invade the deeper tissues of the graft. Most new vessels form in the connective tissues, but some vessels, especially larger ones, grow into persisting basal laminae from preexisting fibers and nerves. The differentiation of new arterioles and venules in free muscle grafts is described. By the end of the first week, the entire graft is revascularized, and ultimately a fairly normal relationship between new capillaries and regenerating muscle fibers is established. In mature grafts, however, irregularities are sometimes found in the organization of smooth muscle cells associated with larger vessels.

Cellular responses to free grafting of the extensor digitorum longus muscle of the rat.

The cellular and subcellular responses related to the survival or destruction and subsequent regeneration of muscle fibers within the freely grafted extensor digitorum longus muscle of the rat were examined by light and electron microscopy. A small number of fibers at the periphery of the grafts survived the initial ischemia but underwent denervation changes and accumulated lipid deposits. The majority of fibers in the grafts, however, became ischemic and underwent an intrinsic degeneration within 4 hours. Cell-mediated destruction of the degenerating fibers occurred as the grafts became revascularized. The basal laminae and some of the satellite cells were the only elements of the original fibers that persisted. Regeneration began at the periphery of the graft within three days after grafting and reached the center about three days later. After phagocytosis of the original fibers, presumptive myoblasts within the grafts differentiated into myoblasts and myotubes. The formation of myotubes followed a biphasic pattern of development comparable to that of normal fetal muscle. Although most of the myotubes were formed within the basal lamina remaining from the original fiber, there was also evidence for regeneration outside the basal lamina. Myotubes matured into muscle fibers which were essentially normal in apperance when examined up to 180 days after grafting. Some fibers, however, were atrophic, presumably due to a failure to become innervated, and some fibers were joined by myo-myous junctions. Pre-denervated grafts and Marcaine-treated grafts were also examined. There were more surviving fibers in pre-denervated grafts, and cell-mediated destruction of degenerating fibers proceeded more rapidly than in normal grafts. No surviving fibers were found in Marcaine-treated grafts. The changes in these grafts were otherwise similar to normal grafts. A schematic model of the spatial and temporal sequence of degeneration and regeneration within a free muscle graft is presented.

Muscle satellite cells in malnourished and nutritionally rehabilitated children.

Satellite cells were examined qualitatively and quantitatively in muscle biopsies from children when malnourished, during nutritional rehabilitation, and after clinical recovery. The proportion of satellite cell nuclei relative to myonuclei was significantly lower in malnourished subjects than in well-nourished age-matched controls (4.5 +/- 1% vs. 8.1 +/- 1.0%). The proportion of satellite cells remained low during the early period of "catch-up growth" but was significantly increased in the recovered subjects (10.5 +/- 1.0%). Satellite cells were small and their nuclei were heterochromatic in biopsies from the malnourished subjects. The cells were often partially segregated from the parent fiber by an external lamina. In recovering and recovered subjects many of the satellite cells enlarged, and the appearance of their nuclei and cytoplasmic organelles suggested a more active state. Intervention of external lamina between the satellite cell and the myofibre was uncommon in the recovered subjects.

Growth of muscle fibres during recovery from severe malnutrition in Jamaican infants.

1. The growth of muscle fibres was analysed by light microscopy in biopsies from subjects when malnourished, during nutritional rehabilitation, and after clinical recovery. 2. Muscle fibres from malnourished subjects were extremely atrophic (cross-sectional area, 110 micrometers2). The fibres doubled in size during the early period of rehabilitation. Growth of muscle fibres during later periods of rehabilitation occurred at a slower rate. 3. The absolute rates of change in fibre sizes differed considerably between subjects, but the rates of change relative to the rate of gain of total body-weight (expressed as % recovery or % expected weight-for height (Nelson, 1975)) were similar between subjects after the initial growth spurt. The pattern of recovery appeared to differ between older and younger subjects. 4. Fibre sizes correlated with body-weight but not with age in the malnourished subjects. A significant correlation between fibre areas and either weight or age was observed during rehabilitation and after clinical recovery. 5. Fibre sizes of clinically-recovered subjects (mean age, 13.8 months; weight, 8.7 kg) were only approximately 60% of that for a well-nourished 6-month-old control subject (6.4 kg). These results suggest that a longer period of time is required for fibres to reach their expected size. Therefore, when the child has regained body-weight to that of a normal child of the same height, his muscles have not yet recovered and his body composition is abnormal.

Comparison of the chemical and biochemical composition of thirteen muscles of the rat after dietary protein restriction.

1. The objective of this study was to determine whether the chemical and biochemical changes induced by muscle wasting caused by dietary protein restriction are different in various skeletal muscles. 2. Rats were fasted for 3 d and then fed on a 10 g protein/kg diet for 21 d. Thirteen muscles from the trunk, forelimb, and hind-limb regions were analysed for muscle weight, and the content of water, fat, cellular and extracellular protein, DNA and RNA. Results were compared to values for an 'initial' control group killed at the start of the experiment. 3. Weight loss was greatest in trunk muscles and least in the distal forelimb muscles. Water content decreased in most muscles, but increased in three forelimb muscles. A significant loss of lipid was found in the gastrocnemius, while the biceps brachii gained lipid. Changes in lipid content of the muscles did not form a distinctive pattern. 4. All muscles except the distal forelimb muscles lost a significant amount of cellular protein, while all muscles except the diaphragm gained extracellular protein. 5. DNA content was unchanged in all muscles. The value for cellular protein:DNA was significantly reduced in the rectus abdominis and the diaphragm. A significant loss of RNA was found in all muscles; the percentage change was greatest in trunk muscles and least in the distal forelimb muscles. The values for RNA:protein and RNA:DNA were significantly lower in all muscles except two distal forelimb muscles. 6. With the exception of the water and lipid content of the muscles, the directions of the changes in the experimental animals were the same for all muscles. The results suggested, however, that the magnitude of changes in certain chemical and biochemical indices of composition may depend to some extent on the anatomical location of the muscle: trunk muscles tended to show the greatest percentage change, while the distal forelimbs changed the least.

Comparison of the chemical and biochemical composition of thirteen muscles of the rat after dietary protein restriction

The objective of this study was to determine whether the chemical and biochemical changes induced by muscle wasting caused by dietary protein restriction are different in various skeletal muscles. Rats were fasted for 3 d and then fed on a 10 g protein/kg diet for 21d. Thirteen muscles from the trunk, forelimb, and hind-limb regions were analysed for muscle weight, and the content of water, fat, cellular and extracellular protein, DNA and RNA. Results were compared to values for an `initial' control group killed at the start of the experiment. Weight loss was greatest in trunk muscles and least in the distal forelimb muscles. Water content decreased in most muscles, but increased in three forelimb muscles. A significant loss of lipid was found in the gastroenemius, while the biceps brachii gained lipid. Changes in lipid content of the muscles did not form a distinctive pattern. All muscles except the distal forelimb muscles lost a significant amount of cellular protein, while all muscles except the diaphragm gained extracellular protein. DNA content was unchanged in all muscles. The value for cellular protein: DNA was significantly reduced in the rectus abdominis and the diaphragm. A significant loss of RNA was found in all muscles; the percentage change was greatest in trunk muscle and least in the distal forelimb muscles. The values for RNA: protein and RNA: DNA were significantly lower in all muscles except two distal forelimb muscles. With the exception of the water and lipid content of the muscles, the directions of the changes in the experimental animals were the same for all muscles. The results suggested, however, that the magnitude of changes in certain chemical and biochemical indices of composition may depend to some extent on the anatomical location of the muscle: trunk muscles tended to show the greatest percentage change, while the distal forelimbs changed the least.(AU)