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PubMed2002

Insulin growth factor-1 decreases muscle atrophy following denervation.

Despite modern microsurgical techniques for nerve repair, functional outcome following proximal injury is often unsatisfactory because irreversible muscle atrophy may develop before reinnervation occurs. Because insulin growth factor-1 (IGF-1) has been shown to improve muscle regeneration after injury, and may have a role in muscle preservation following denervation, the purpose of this investigation was to evaluate the histological, immunohistochemical, and electrophysiological differences between normal, denervated, and IGF-1-injected denervated muscle over an 8-week period. Denervated mice gastrocnemius muscles demonstrated a decrease in muscle weight, a decrease in myofiber diameter, an absence of muscle regeneration, an early increase in the number of neuromuscular junctions (NMJs), and a decrease in fast-twitch and maximum tetanic strength as compared to normal muscle up to 8 weeks following denervation. IGF-1-injected denervated muscle, on the other hand, sustained muscle diameter and muscle weight, maintained a smaller number of NMJs, and relatively sustained fast-twitch and maximum tetanic strength as compared to normal muscle over 8 weeks. These data suggest that IGF-1 may help prevent muscle atrophy and secondary functional compromise after denervation.

Animals↗
PubMed1990

[A histological study on changes in the attachment of deep layer of the masseter muscle after denervation of the masseteric nerve].

To elucidate the histological and histochemical changes on the deep layer of masseter muscle at the site of their attachment to the mandible after masseteric denervation, 35-day old male rats were used in this experiment. The rats were sacrificed at 10 and 20 days after the left masseteric nerve was resected at the mandibular notch. The attachment of deep layer of masseter muscle to the mandible was examined using light microscopy, lead labelling technique (Okada and Mimura 1938), and histochemical analysis for ALPase activity. Electromyographic techniques were used for detecting masseter muscle function. 1. 10 days after denervation, the number, shape and ALPase activities of osteoblasts in the transitional zone and fibrocartilage cells in the tendinous attachment on the operated side were seen to decrease compared with those in control. Bone formation on the operated side was less than that in control. The electrophysical activity of masseter muscle was not detected on the operated side. 2. 20 days after surgery, ALPase activities of osteoblasts and fibrocartilage cells, and bone formation in the transitional zone and tendinous attachment on the operated side increased, compared with those observed in the same area on the operated side in the 10-day group, but still less than those in control. Low levels of electrophysical activity in masseter muscle were detected on the operated side. 3. In the periosteal attachment and inferior surface of the mandible on the operated side, bone formation was also seen to be less than that in control. These results suggested that the function of masseter muscle seems to exert the greatest influence on the proliferation and differentiation of both fibrocartilage cells and osteoblasts, and bone growth in the attachment of masseter muscle.

Alkaline Phosphatase↗
PubMed1981

An assessment of the spread of the signal for terminal sprouting within and between muscles.

Certain muscles of the mouse and rat have been studied in order to assess how far a signal from denervated muscle can spread to elicit terminal sprouting from intact endplates. Denervation of the muscles surrounding the rat foot 4th lumbrical muscle caused no terminal sprouting in the 4th lumbrical itself. In the hemidenervated mouse gluteus maximus terminal sprouting was restricted to the central region of the muscle where innervated and denervated fibres intermingle. There was no enhancement of such sprouting if the underlying and closely apposed gluteus medius was simultaneously denervated. Hemidenervation of the mouse diaphragm and interscutularis, where intact endplates lie near to denervated muscle fibres, produced no terminal sprouting. Hemidenervation of the mouse platysma, where intact endplates often lie adjacent to denervated muscle fibres, similarly produced no significant response. However, all muscles were capable of producing extensive terminal sprouting in response to paralysis induced by botulinum toxin. The stimulus for terminal sprouting produced by an inactive muscle fibre must therefore be effective only on the fibre's own terminal or immediately adjacent terminals.

Animals↗
PubMed2004

Morphometric ultrastructural analysis of satellite cells in denervated rat soleus muscle.

Morphometric analysis of the ultrastructural changes of satellite cells (SCs) of rat soleus muscle 7, 14, and 36 days post denervation was performed. Denervation caused decrease of surface area cross-section of SCs and their nuclei, volume fractions of nucleus and Golgi complex elements and ribosomes number. In contrast, the increase of surface area/volume ratio of SCs and nucleus, volume fraction and number of endosome/lysosome-like structures, and number of caveolae-like structures was noticed. Ultrastructural changes of SCs in denervated muscles strongly suggest decline of cell activity accompanied by increased processes of degradation of material of endo-, and/or egzogenous origin.

Animals↗
PubMed1993

Potassium channels from normal and denervated mouse skeletal muscle fibers.

The properties of singles K+ channels in normal and denervated muscles were compared using the "patch-clamp" technique. Single channels were recorded from vesicles obtained by stretching bundles of normal and denervated extensor digitorium longus (EDL) muscles. The most frequently observed channel in normal muscles was a high conductance (266 pS) Ca++ activated K+ channel. Although channel density, as estimated by patch recording, showed a significant decrease in denervated muscles, no differences were found in conductance and gating properties. Another voltage-dependent K+ channel (81 pS) was only recorded from normal muscles, but never from denervated ones. In addition, a 35 pS conductance was recorded from both normal and denervated fibers. This channel displayed neither voltage dependence nor sensitivity to tetraethylammonium (TEA). In contrast, another TEA-insensitive (16 pS) channel was recorded only from denervated muscles. We conclude that denervation induces significant changes in the distribution and expression of K+ channels in mammalian skeletal muscles.

Animals↗
PubMed1990

Enzymes of transmitter and energy metabolism in cat middle ear muscles.

Activities of the enzymes choline acetyltransferase (ChAT) and acetylcholinesterase (AChE), which metabolize the neuromuscular transmitter acetylcholine, and malate and lactate dehydrogenase (MDH and LDH), enzymes of oxidative and glycolytic energy metabolism, respectively, were measured in the middle ear muscles of the cat. For comparison, the same enzyme activities were measured in extraocular muscle tissue and in three hindlimb muscles rich in either slow oxidative (soleus), fast glycolytic (white part of vastus lateralis), or fast oxidative glycolytic (plantaris) muscle fibers. ChAT and AChE activities were much higher in middle ear muscles than in hindlimb muscles, consistent with a denser neuromuscular innervation, as in extraocular muscles. By contrast, MDH and LDH activities were remarkably low in the middle ear muscles, lower than in any of the hindlimb muscles or the extraocular muscles. Denervation of the stapedius muscle by peripheral transection of the facial nerve resulted in decreases in all four enzyme activities without associated changes in the tensor tympani. Surgical ablation of the peripheral facial nerve supply to the stapedius muscle appears to be a feasible option for producing its denervation. The results suggest some rather specialized chemical characteristics for the middle ear muscles.

Acetylcholinesterase↗
PubMed1998

Agrin and acetylcholine receptor distribution following electrical stimulation.

Electrical stimulation is a therapeutic modality available for the preservation of muscle function following peripheral nerve injury. Agrin, a synaptic basal lamina protein, induces accumulation of acetylcholine receptors (AChRs) and other molecules at the neuromuscular junction. Electrical stimulation of denervated muscle does not alter agrin and AChR distribution at abandoned synaptic sites, supporting the hypothesis that the existing aggregation of synaptic molecules, which may be necessary for successful reinnervation, is unaltered by electrical stimulation of denervated muscle.

Animals↗
PubMed1988

Comparison of injury and development in the neuromuscular system.

Comparisons of development and regeneration have suggested that axotomized motoneurons and denervated muscles undergo dedifferentiation to an embryonic state with recovery of adult properties after reinnervation. Using electrophysiological and radioligand-binding techniques to monitor axonal size and numbers of extrajunctional acetylcholine receptors in axotomized motoneurons and denervated muscles respectively, we have demonstrated that this dedifferentiation is limited. We suggest that this limited dedifferentiation may be adaptive for survival, regeneration and reinnervation. Correlative physiological and histochemical studies of reinnervated motor units in cat and rat hindlimb muscles show that the processes of regeneration and reinnervation differ in a number of fundamental ways from developmental processes of axonal growth and muscle innervation. Enlargement of motor units after partial nerve injuries does not appear to be limited to the size of the neonatal motor unit as originally suggested but may be influenced by factors operating at the level of axonal branching. Regeneration after complete and partial nerve injuries is a random process in contrast to the specific nature of the innervation of targets during development. Regenerating axons frequently fail to make connections with their original muscles and newly reinnervated motor units contain muscle fibres which formerly belonged to several different motor units. Despite this misdirection of regenerating nerve fibres, neuromuscular plasticity restores neuromuscular properties to the extent that these are appropriate at the single motor unit level for the gradation of force by the orderly recruitment of units during movement.

Animals↗
PubMed1989

The onset of myogenesis in denervated mouse skeletal muscle regenerating after injury.

Denervation of skeletal muscle stimulates increased turnover of muscle nuclei and connective tissue cells. The present investigation tests whether denervation "primes" myogenic precursor cells, so that the onset of DNA synthesis in muscle precursors after traumatic injury occurs earlier than in innervated muscle. The left legs of 29 male BALBc mice were denervated, and 1 week later small incisions were made in the tibialis anterior muscles of both legs (denervated and innervated). At specific times after injury each mouse was injected once with tritiated thymidine to label replicating muscle precursors. Muscle lesions were sampled 10 days after injury (when all precursors had fused to form myotubes) and prepared for auto-radiography. The presence of labelled myotube nuclei showed that muscle precursors had been synthesizing DNA at the time when [3H]thymidine had been injected. Our data suggest that very few precursors were proliferating in denervated muscle within 30 h after injury, and the onset of myogenesis at 30 h was essentially the same in denervated and innervated muscle. The retrospective analysis indicates that there were also similar proportions of muscle precursors proliferating at different times after injury, in regenerating lesions of both denervated and innervated muscle. Thus, denervation does not stimulate an earlier regenerative response in injured skeletal muscle.

Animals↗
PubMed2002

[Ultrastructural changes after denervation of different muscles].

OBJECTIVE: To observe the ultrastructural changes and number of satellite cells in different muscles with different denervation interval and investigate the mechanism of denervation atrophy. METHODS: Muscles of different denervation interval were harvested, which were 6 biceps brachii and 6 abductor digiti minimi. The ultrastructure of the samples were observed under transmission electron microscope. The number of nucleus and satellite cells were counted to calculate the percentage content of satellite cells. RESULTS: In early stage of denervation, the myofilament and sarcomere of the majority were well oriented. The nucleoli of some muscle cell nucleus were enlarged and pale. Vacuolarization was also seen in some mitochondria. There was no obvious proliferation of collagen fiber around myofibers. After denervation of half a year, rupture and disorientation of myofilament was seen. The nucleus became smaller, dark stained, and some of them were condensed. There was proliferation of fibroblasts, adipose cells and collagen fibers around myofibers. Motor endplate was not recognized one year after denervation. In the early stage of denervation, satellite cell percentage of the two muscles was relatively high. It then declined with time. One year after denervation, satellite cells were scarcely detected. Comparison of the curves for satellite cell declination in two muscles revealed that the declination of the abductor digiti minimi was faster than that of biceps brachii. Decrease of the former started 3 months after denervation, while the latter started after 6 months. CONCLUSION: Disappearing of motor endplate and proliferation of collagen fibers are main factors that affect the treatment outcome in late cases. Decrease of satellite cell number is another cause. The correlation of less satellite cell in abductor digiti minimi and poorer recovery of hand intrinsic muscles indicates that increment of satellite cells in long-term denervated muscles may be one of the effective measures to improve treatment outcome.

Brachial Plexus↗
PubMed1978

Automatic quantitative analysis of the electromyogram in partially denervated distal muscles: comparison with motor unit counting.

Using the ANOPS-101 minicomputer, an automatic analysis of muscle action potentials (MAP) was performed on a total of 53 extensor digitorum brevis (EDB) and abductor pollicis brevis (APB) muscles in patients with other evidence of denervation. The results were compared with observations made on 27 control subjects and they were also matched against estimates of functioning motor units made by the incremental stimulating technique of McComas et al (1971). Although the incidence of abnormal MAP parameters was relatively low in both muscles, it was possible to improve the diagnostic yield by combining the results for individual muscles. The relative merits and disadvantages of motor unit counting and automatic MAP analysis are discussed.

Adult↗
PubMed2005

Expression patterns of initiator and effector caspases in denervated human skeletal muscle.

There is evidence that apoptotic cell death contributes to the loss of denervated muscle fibers. In 17 patients with neurogenic muscular atrophy, we studied the expression of the apoptosis mediators APAF-1/caspase-9 and degrading caspases-2, -3, and -7 by immunohistochemical and western blot analyses. Muscle with neurogenic atrophy showed distinct upregulation of caspase-9 and -7 and no expression for APAF-1 (apoptosis protease-activating factor-1) and caspase-2 and -3. Expression of caspase-7 was restricted to atrophic fibers, but caspase-9 was also found in normal-sized muscle fibers where its expression was often confined to single fiber segments. These findings indicate that upregulated expression of caspase-9 can initiate the proteolytic cascade involving the downstream executioner caspase-7, which mediates degradation of denervated muscle fibers. However, apoptotic events may be restricted to single muscle-fiber segments, where apoptotic cell degradation contributes to the long-term process of atrophy. Pharmacological inhibition of caspases may be a therapeutic strategy in diminishing muscle atrophy.

Adult↗
PubMed1979

Neural control of skeletal muscle cholinesterase: a study using organ-cultured rat muscle.

1. It has been proposed that the influence of innervation on the cholinesterase activity (ChE) of skeletal muscle and on end-plate ChE in particular is mediated by trophic substance(s) moved by axonal transport and released from nerve. We have tested this hypothesis using rat extensor digitorum longus (e.d.l.) and diaphragm muscles denervated in vitro for several days and then maintained in organ culture to assay putative trophic substance(s). 2. The cholinesterase activity (ChE) of rat extensor digitorum longus (e.d.l.) muscles decreased dramatically after 5 days of denervation in vivo as previously reported. The ChE of rat e.d.l. muscles denervated in vivo for 3 days and then maintained in organ culture for 2 days was essentially identical to that of muscles denervated 5 days in vivo. 3. The ChE OF E.D.L. MUSCLES DENERVATED IN VIVO FOR 3 DAYS AND THEN MAINTAINED FOR 2 DAYS IN CULTURE MEDIUM SUPPLEMENTED WITH SCIATIC NERVE OR INNERVATED MUSCLE EXTRACT WAS SIGNIFICANTLY HIGHER THAN THAT OF MUSCLES DENERVATED IN VIVO FOR 5 DAYS OR DENERVATED IN VIVO FOR 3 DAYS AND THEN CULTURED FOR 2 DAYS IN CULTURE MEDIUM ALONE. Supplementing the culture medium with brain or spinal cord extract also significantly increased the ChE of organ-cultured e.d.l. muscles. 4. Supplementing the culture medium with liver or spleen extract or with the extract of muscle denervated for 3--7 days in vivo before extraction did not increase the ChE or organ-cultured e.d.l. muscles. 5. The effect of muscle extract on the ChE of organ-cultured e.d.l. muscles was dose dependent and occurred gradually reaching a maximum after approximately 24 h of culture. 6. Substance(s) which increased the ChE of organ-cultured e.d.l. muscles were found to accumulate in transected sciatic nerve in the region just proximal to the site of transection where substances moved by axonal transport are known to accumulate. 7. Media conditioned with neurally stimulated e.d.l. or diaphragm muscles caused a substantial and highly significant increase in the ChE of e.d.l. or diaphragm muscles denervated in vivo and then maintained in organ culture. Media conditioned in the same way with unstimulated muscles did not increase the ChE OF ORGAN-CULTURED MUSCLES. 8. The active substance(s) released by neural stimulation continued to be released when muscle contraction was blocked by adding D-tubocurarine to the medium during conditioning but the release of these substance(s) was significantly reduced when magnesium (10mM) was added to the medium during conditioning. 9 The substance(s) released by neural stimulation selectively increased ChE in the end-plate region. In diaphragm segments denervated in vivo and then maintained in medium conditioned with neurally stimulated muscle, there was a 102% increase in end-plate ChE but no detectable increase in background ChE. 10...

Animals↗
PubMed1996

Identification and characterization of a 47 base pair activity-dependent enhancer of the rat nicotinic acetylcholine receptor delta-subunit promoter.

Nicotinic acetylcholine receptor (nAChR) genes are regulated by muscle electrical activity. E-box sequences found in their promoters are necessary for this regulation. However, many muscle genes contain E-boxes, yet are not regulated by muscle depolarization. This suggests that other elements are necessary, perhaps working in conjunction with E-boxes, to confer depolarization-dependent control onto promoter activity. We have used direct DNA injection into muscle as an in vivo assay to identify and characterize these additional elements. Mutagenesis and expression assays identified multiple elements within the first 81 base pairs (bp) of the nAChR delta-subunit promoter that contribute to its regulation by muscle electrical activity. Within this 81 bp sequence, two regions of DNA were identified that were capable of conferring activity-dependent regulation onto a heterologous promoter. The stronger of these two putative enhancers was characterized further. It is a 47 bp sequence that contains an E-box along with sequences similar to the SV40 core enhancer and an SP1 site. Site-directed mutagenesis identified residues within each of these sequences that were necessary for enhancer activity. Furthermore, methylation interference DNA footprinting assays showed increased nuclear protein binding to sequences within both these enhancers after muscle denervation, and this pattern of binding was very similar to that observed with nuclear protein isolated from myotube extracts. These latter results suggest that similar mechanisms may mediate increased nAChR expression during muscle development and after muscle denervation.

Animals↗
PubMed1998

Myotrophic effects of muscle extracts obtained at different intervals after denervation.

A study was made of the myotrophic effects of denervated muscle extracts on normal Wistar rat soleus muscle. Extracts obtained 1 h, 2, 4 and 7 days after sectioning of the sciatic nerve were administered intraperitoneally over five consecutive days. Soleus muscles were routinely processed for morphological and morphometrical analysis using light microscopic techniques. Quantitative differences were observed in the effects of different extracts on total muscle area, fibre cross-sectional area, mean minimum diameter and number of fibres/ area. The greatest myotrophic response was elicited by extracts obtained at 2 and 4 days; differences with respect to controls and extracts obtained at 1 day were significant (P < 0.05) for all parameters studied. Statistically significant differences (P < 0.05) were also recorded for fibre cross-sectional area and mean minimum diameter between the 2- and 4-day groups and the 7-day group. It may thus be concluded that the time elapsing between denervation and extract obtention influences the effect of the extract on normal rat muscle.

Animals↗
PubMed1977

Subunit structure of the acetylcholine receptor from denervated rat skeletal muscle.

Acetylcholine receptor from denervated rat leg muscle was purified 48,000-fold by affinity chromatography on concanavalin A/Sepharose and cobrotoxin/Sepharose. A control preparation containing only contaminants was made by performing a parallel purification in which alpha-bungarotoxin was added to the preparation prior to the cobrotoxin/Sepharose step. Comparison of the receptor was greater than 90% pure. Examination of the purified receptor by polyacrylamide gel electrophoresis in sodium dodecyl sulfate revealed two major polypeptide chains with apparent weights of 45,000 and 51,000, along with minor components of 49,000, 56,000, 62,000, and 110,000. Polypeptides of the same molecular weight were found when purified acetylcholine receptor was radioiodinated and further purified by sucrose gradient sedimentation. Analysis of receptor purified from denervated muscles that had been incubated with [35S]methionine in organ culture showed that all of the identified polypeptide chains were radioactive, indicating that they had been synthesized de novo after denervation.

Acetylcholine↗