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Optimal recovery of muscle function after proximal nerve injuries remains a complex and challenging problem. After a nerve injury, alterations in the affected muscles lead to atrophy, and later degeneration and replacement by fat-fibrous tissues. At present, several different strategies for the preservation of skeletal muscle have been reported, including various sets of physical exercises, muscle massage, physical methods (e.g. electrical stimulation, magnetic field and laser stimulation, low-intensity pulsed ultrasound), medicines (e.g. nutrients, natural and chemical agents, anti-inflammatory and antioxidants, hormones, enzymes and enzyme inhibitors), regenerative medicine (e.g. growth factors, stem cells and microbiota) and surgical procedures (e.g. supercharge end-to-side neurotization). The present review will focus on methods that aimed to minimize the damage to muscles after denervation based on our present knowledge., Competing Interests: Declaration of conflicting interestsThe authors declare no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Neuregulin 1 (NRG1) is a growth factor produced by both peripheral nerves and skeletal muscle. In muscle, it regulates neuromuscular junction gene expression, acetylcholine receptor number, muscle homeostasis and satellite cell survival. NRG1 signalling is mediated by the tyrosine kinase receptors ErbB3 and ErbB4 and their co-receptors ErbB1 and ErbB2. The NRG1/ErbB system is well studied in nerve tissue after injury, but little is known about this system in skeletal muscle after denervation/reinnervation processes. Here, we performed a detailed time-course expression analysis of several NRG1 isoforms and ErbB receptors in the rat superficial digitorum flexor muscle after three types of median nerve injuries of different severities. We found that ErbB receptor expression was correlated with the innervated state of the muscle, with upregulation of ErbB2 clearly associated with the denervation state. Interestingly, the NRG1 isoforms were differently regulated depending on the nerve injury type, leading to the hypothesis that both the NRG1α and NRG1β isoforms play a key role in the muscle reaction to injury. Indeed, in vitro experiments with C2C12 atrophic myotubes revealed that both NRG1α and NRG1β treatment influences the best-known atrophic pathways, suggesting that NRG1 might play an anti-atrophic role.

Muscle regeneration depends on satellite cells (SCs), quiescent precursors that, in consequence of injury or in pathological states such as muscular dystrophies, activate, proliferate, and differentiate to repair the damaged tissue. A subset of SCs undergoes self-renewal, thus preserving the SC pool and its regenerative potential. Unacylated ghrelin (UnAG) is a circulating hormone that protects muscle from atrophy, promotes myoblast differentiation, and enhances ischemia-induced muscle regeneration. Here we show that UnAG increases SC activity and stimulates Par polarity complex/p38-mediated asymmetric division, fostering both SC self-renewal and myoblast differentiation. Because of those activities on different steps of muscle regeneration, we hypothesized a beneficial effect of UnAG in mdx dystrophic mice, in which the absence of dystrophin leads to chronic muscle degeneration, defective muscle regeneration, fibrosis, and, at later stages of the pathology, SC pool exhaustion. Upregulation of UnAG levels in mdx mice reduces muscle degeneration, improves muscle function, and increases dystrophin-null SC self-renewal, maintaining the SC pool. Our results suggest that UnAG has significant therapeutic potential for preserving the muscles in dystrophies. Stem Cells 2017;35:1733-1746., (© 2017 The Authors STEM CELLS published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.)

Platelet-rich plasma (PRP) has received increasing interest in applied medicine, being widely used in clinical practice with the aim of stimulating tissue healing. Despite the reported clinical success, there is still a lack of knowledge when considering the biological mechanisms at the base of the activity of PRP during the process of muscle healing. The aim of the present study was to verify whether the local delivery of PRP modulates specific molecular events involved in the early stages of the muscle regeneration process. The right flexor sublimis muscle of anesthetized Wistar rats was mechanically injured and either treated with PRP or received no treatment. At day 2 and 5 after surgery, the animals were sacrificed and the muscle samples evaluated at molecular levels. PRP treatment increased significantly the mRNA level of the pro-inflammatory cytokines IL-1β, and TGF-β1. This phenomenon induced an increased expression at mRNA and/or protein levels of several myogenic regulatory factors such as MyoD1, Myf5 and Pax7, as well as the muscular isoform of insulin-like growth factor1 (IGF-1Eb). No effect was detected with respect to VEGF-A expression. In addition, PRP application modulated the expression of miR-133a together with its known target serum response factor (SRF); increased the phosphorylation of αB-cristallin, with a significant improvement in several apoptotic parameters (NF-κB-p65 and caspase 3), indexes of augmented cell survival. The results of the present study indicates that the effect of PRP in skeletal muscle injury repair is due both to the modulation of the molecular mediators of the inflammatory and myogenic pathways, and to the control of secondary pathways such as those regulated by myomiRNAs and heat shock proteins, which contribute to proper and effective tissue regeneration.

Sciatic nerve crush injury in rat animal model is one of the most common experimental models used in regenerative research. However, the availability of transgenic mouse for nerve regeneration studies is constantly increasing and, therefore, the shift from rat model to mouse model is, in some cases, necessary. Moreover, since most of the human nerve lesions occur in the upper limb, it is also advantageous to shift from sciatic nerve to median nerve. In this study we described an experimental model which involves lesions of the median nerve in the mouse. Data showed that the finger flexor muscle contraction strength, assessed to evaluate the motor function recovery, and reached values not different from the control already 20 days after injury. The degree of nerve regeneration evaluated with stereological methods in light microscopy showed that, 25 days after injury, the number of regenerated myelinated fibers was comparable to the control, but they were smaller with a thinner myelin thickness. Stereological analysis made in electron microscopy confirmed these results, although the total number of fibers quantified was significantly higher compared to light microscopy analysis, due to the very small size of some fibers that can be detected only in electron microscopy.

Background: There is abundant evidence suggesting that growth factors may play a key role in the healing process, especially in the early stages of inflammation. Despite the reported clinical successes with the use of growth factors there is still a lack of knowledge on the biological mechanism underlying the activity of platelet-rich plasma during the process of muscle healing. The aim of this study was to analyse the early effects of platelet- rich plasma in an easily reproducible animal model., Materials and Methods: Wistar male adult rats (n=102) were used in this study. The muscle lesion was created with a scalpel in the flexor sublimis muscles. Platelet-rich plasma was applied immediately after surgery. Treated, untreated and contralateral muscles were analysed by morphological evaluation and western blot assay., Results: Leucocyte infiltration was significantly greater in muscles treated with platelet-rich plasma than in both untreated and contralateral muscles. The latter showed greater leucocyte infiltration when compared to the untreated muscles. Platelet-rich plasma treatment also modified the cellular composition of the leucocyte infiltration leading to increased expression of CD3, CD8, CD19 and CD68 and to decreased CD4 antigen expression in both platelet-rich plasma treated and contralateral muscles. Blood vessel density and blood vessel diameters were not statistically significantly different between the three groups analysed., Discussion: The results of this study showed that treatment with platelet-rich plasma magnified the physiological early inflammatory response following a muscle injury, modifying the pattern of cellular recruitment. Local platelet-rich plasma treatment may exert a direct or, more plausibly, indirect systemic effect on healing processes, at least in the earliest inflammatory phase.

Preservation of biochemical processes in muscles is a major challenge in patients with severe peripheral nerve injury. In this chapter, we address the effects of laser irradiation and biochemical transformation in muscle, using in vitro and in vivo experimental models. The authors attempt to explain the possible mechanism of laser phototherapy applied on skeletal muscle on the basis of literature review and new results. A detailed knowledge of the evolution of endplates acetylcholine receptors and creatine kinase activity following laser irradiation can help to understand the therapeutic effect of laser phototherapy on muscle. This study showed that the laser phototherapy increases biochemical activity in intact muscle and thus could have direct therapeutic applications on muscle, especially during progressive atrophy resulting from peripheral nerve injury., (© 2013 Elsevier Inc. All rights reserved.)

Despite extensive research and surgical innovation, the treatment of peripheral nerve injuries remains a complex issue, particularly in nonsharp lesions. The aim of this study was to assess the clinical outcome in a group of 16 patients who underwent, in emergency, a primary repair for crush injury of sensory and mixed nerves of the upper limb with biological tubulization, namely, the muscle-vein-combined graft. The segments involved were sensory digital nerves in eight cases and mixed nerves in another eight cases (four median nerves and four ulnar nerves). The length of nerve defect ranged from 0.5 to 4 cm (mean 1.9 cm). Fifteen of 16 patients showed some degree of functional recovery. Six patients showed diminished light touch (3.61), six had protective sensation (4.31), and three showed loss of protective sensation (4.56) using Semmes-Weinstein monofilament test. All the patients who underwent digital nerve repair had favorable results graded as S4 in one case, S3+ in six cases, and S3 in one case. With respect to mixed nerve repair, we observed two S4, two S3+, two S3, one S2, and one S0 sensory recovery. Less favorable results were observed for motor function with three M4, one M3, two M2, and two M0 recoveries. Altogether, the results of this retrospective study demonstrates that tubulization nerve repair in emergency, in case of short nerve gaps, may restore the continuity of the nerve avoiding secondary nerve grafting. This technique preserves donor nerve and, in case of failure, does not preclude a delayed repair with a nerve graft., (Copyright © 2012 Wiley Periodicals, Inc.)

Neuromuscular recovery after peripheral nerve lesion depends on the regeneration of severed axons that re-establish their functional connection with the denervated muscle. The aim of this study was to determine the effects of electrical stimulation (ES) on the neuromuscular recovery after nerve crush injury in rats. Electrical stimulation was carried out on the tibialis anterior (TA) muscle after sciatic nerve crush injury in a rat model. Six ES sessions were administered every other day starting from day 3 postinjury until the end of the experiment (day 14). The sciatic functional index was calculated. Muscle excitability, neural cell adhesion molecule (N-CAM) expression, and muscle fiber cross-sectional area (CSA) were accessed from TA muscle. Regenerated sciatic nerves were analyzed by light and confocal microscopy. Both treated (crush+ES) and untreated (crush) groups had their muscle weight and CSA decreased compared with the normal group (P < 0.05). Electrical stimulation accentuated muscle fiber atrophy more in the crush+ES than in the crush group (P < 0.05). N-CAM expression increased in both crush and crush+ES groups compared with the normal group (P < 0.05). Regenerated nerves revealed no difference between the crush and crush+ES groups. Nevertheless, functional recovery at day 14 post-injury was significantly lower in crush+ES group compared with the crush group. In addition, the crush+ES group had chronaxie values significantly higher on days 7 and 13 compared with the crush group, which indicates a decrease in muscle excitability in the crush+ES animals. The results of this study do not support a benefit of the tested protocol of ES during the period of motor nerve recovery following injury.

Skeletal muscle atrophy represents one of the main causes of poor outcome of microsurgical nerve reconstruction. Recent studies have pointed to the importance of the neuregulin/ErbB signaling pathway in the development and regeneration of the neuromuscular system. Here, we show by immunohistochemistry, RT-PCR, and Western blotting analyses, in an in vivo model of adult skeletal muscle denervation/reinnervation, that expression of Neuregulin1 (NRG1) and ErbB receptors is regulated by the innervation condition. We found out that a significant upregulation of the alpha-, but not beta-, isoform of NRG1, as well as of ErbB2, ErbB3, and ErbB4-cyt1 isoform occurs as a consequence of denervation of flexor digitorum muscles of the rat forelimb by median nerve transection. Moreover, after tubulization median nerve repair, and consequent muscle reinnervation, all messengers of the NRG1/ErbB system are promptly downregulated. Therefore, our results suggest the existence of a alpha-NRG1-mediated autocrine and/or paracrine trophic loop in skeletal muscles that is activated after denervation and promptly deactivated after nerve reconstruction. This myotrophic loop is a promising therapeutic target for the prevention of muscle atrophy. Yet, the recent demonstration of a similar alpha-NRG1-mediated gliotrophic loop in denervated Schwann cells provides a possible explanation for the effectiveness of muscle conduits for tubulization nerve repair., (Copyright 2009 Wiley-Liss, Inc. Microsurgery 2009.)

Posttraumatic nerve repair and prevention of muscle atrophy represent a major challenge of restorative medicine. Considerable interest exists in the potential therapeutic value of laser phototherapy for restoring or temporarily preventing denervated muscle atrophy as well as enhancing regeneration of severely injured peripheral nerves. Low-power laser irradiation (laser phototherapy) was applied for treatment of rat denervated muscle in order to estimate biochemical transformation on cellular and tissue levels, as well as on rat sciatic nerve model after crush injury, direct or side-to-end anastomosis, and neurotube reconstruction. Nerve cells' growth and axonal sprouting were investigated in embryonic rat brain cultures. The animal outcome allowed clinical double-blind, placebo-controlled randomized study that measured the effectiveness of 780-nm laser phototherapy on patients suffering from incomplete peripheral nerve injuries for 6 months up to several years. In denervated muscles, animal study suggests that the function of denervated muscles can be partially preserved by temporary prevention of denervation-induced biochemical changes. The function of denervated muscles can be restored, not completely but to a very substantial degree, by laser treatment initiated at the earliest possible stage post injury. In peripheral nerve injury, laser phototherapy has an immediate protective effect. It maintains functional activity of the injured nerve for a long period, decreases scar tissue formation at the injury site, decreases degeneration in corresponding motor neurons of the spinal cord, and significantly increases axonal growth and myelinization. In cell cultures, laser irradiation accelerates migration, nerve cell growth, and fiber sprouting. In a pilot, clinical, double-blind, placebo-controlled randomized study in patients with incomplete long-term peripheral nerve injury, 780-nm laser irradiation can progressively improve peripheral nerve function, which leads to significant functional recovery. A 780-nm laser phototherapy temporarily preserves the function of a denervated muscle, and accelerates and enhances axonal growth and regeneration after peripheral nerve injury or reconstructive procedures. Laser activation of nerve cells, their growth, and axonal sprouting can be considered as potential treatment for neural injury. Animal and clinical studies show the promoting action of phototherapy on peripheral nerve regeneration, which makes it possible to suggest that the time for broader clinical trials has come.

Reorganization of the muscle endplate structures is an important parameter for the study of posttraumatic neuromuscular recovery. The aim of this study was to investigate the changes and the distribution of the acetylcholine receptors (AChRs) in flexor digitorum sublimis muscle after 30 days of denervation in the rat forelimb experimental model. In young male rats, the median and ulnar nerves of the right forelimb were surgically transected and a 1-cm-long segment was removed to avoid spontaneous regeneration. Along the postoperative, the presence of complete functional loss was assessed by the grasping test. After 30 days, rats were sacrificed and flexor digitorum sublimis muscles of both limbs were explanted. The muscles were analysed by light microscopy, to assess the degree of muscle atrophy, and by immunofluorescence after rodhamine-conjugated alpha-bungarotoxin incubation to investigate the reorganization of endplates. The occurrence of muscle denervation was established, prior to sacrifice, by complete loss of the grip function. Light microscopy showed that 30-day denervation is sufficient to induce severe muscle fiber atrophy. Fluorescence analysis at low resolution showed that background fluorescence was higher in denervated muscles possibly because of the presence of extrajunctional AChR. At higher resolution, the endplates were clearly visible as ribbon-like structures. In control fibres, AChR formed a compact and bright structure while in denervated samples it appeared more diffuse and dimmer. Quantitative analysis showed that endplate area was larger in denervated muscles than in control samples. A corresponding decrease in fluorescence intensity was observed after subtracting the basal fluorescence. In conclusion, results of the present study demonstrate that 30 days of denervation induce severe atrophy in rat flexor digitorum sublimis muscle that is accompanied by significant changes in acetylcholine receptor density and distribution. These results also suggest that the rat median nerve denervation experimental model can be an excellent approach for the study of the progression of endplate re-organization after muscle denervation, and reinnervation, considering also its relatively low impact on animal well being in comparison to other experimental models.

Denervation induces muscle fiber atrophy and changes in the gene expression rates of skeletal muscle. Electrical stimulation (ES) is a procedure generally used to treat denervated muscles in humans. This study evaluated the effect of ES based on chronaxie and rheobase on the expression of the myoD and atrogin-1 genes in denervated tibialis anterior (TA) muscle of Wistar rats. Five groups were examined: (1) denervated (D); (2) D+ES; (3) sham denervation; (4) normal (N); and (5) N+ES. Twenty muscle contractions were stimulated every 48 h using surface electrodes. After 28 days, ES significantly decreased the expression of myoD and atrogin-1 in D+ES compared to the D group. However, ES did not prevent muscle-fiber atrophy after denervation. Thus, ES based on chronaxie values and applied to denervated muscles using surface electrodes, as normally used in human rehabilitation, was able to reduce the myoD and atrogin-1 gene expressions, which are related to muscular growth and atrophy, respectively. The results of this study provide new information for the treatment of denervated skeletal muscle using surface ES.

The management of peripheral nerve injury continues to be a major clinical challenge. The most widely used technique for bridging defects in peripheral nerves is the use of autologous nerve grafts. This technique, however, necessitates a donor nerve and corresponding deficit. Many alternative techniques have thus been developed. The use of skeletal muscle tissue as graft material for nerve repair is one example. The rationale regarding the use of the skeletal muscle tissue technique is the availability of a longitudinally oriented basal lamina and extracellular matrix components that direct and enhance regenerating nerve fibers. These factors provide superiority over other bridging methods as vein grafts or (non)degradable nerve conduits. The main disadvantages of this technique are the risk that nerve fibers can grow out of the muscle tissue during nerve regeneration, and that a donor site is necessary to harvest the muscle tissue. Despite publications on nerve conduits as an alternative for peripheral nerve repair, autologous nerve grafting is still the standard care for treatment of a nerve gap in the clinical situation; however, the use of the skeletal muscle tissue technique can be added to the surgeon's arsenal of peripheral nerve repair tools, especially for bridging short nerve defects or when traditional nerve autografts cannot be employed. This technique has been investigated both experimentally and clinically and, in this article, an overview of the literature on skeletal muscle grafts for bridging peripheral nerve defects is presented.

Various tubulization techniques can be used to bridge peripheral nerve lesions with substance loss. Among the different materials that have been used so far in alternative to traditional fresh nerve autografts, fresh muscle-vein combined conduits (made by a vein segment filled with fresh skeletal muscle) proved to be particularly effective. In this study, nerve repair of 10-mm long nerve defects by means of muscle-vein combined tubes was compared with repair by means of traditional nerve autografts in the rat sciatic nerve experimental model. Results did not reveal any significant difference between the two groups of regenerated nerves with respect to the total number, mean density and mean size of myelinated nerve fibers. In addition, we also report the results of an experimental study in the rabbit sciatic nerve model, which showed that fresh skeletal muscle enrichment of the vein segment made it possible to bridge 55-mm long nerve gaps. These results provide further evidence of the effectiveness of fresh muscle-vein combined grafts and support the view that this type of conduit can be used also for repairing long nerve gaps.

Y-shaped vein conduits enriched with fresh skeletal muscle fibers were used to bridge a concomitant ulnar and median nerve transection with substance loss in rats. The proximal limb of the Y-chamber was sutured to the proximal ulnar nerve, while the two distal limbs were sutured to ulnar and median distal nerve stumps. Eight months after surgery, median nerve functional recovery was evaluated by means of the grasping test, and nerve fiber regeneration in both repaired nerves was assessed by means of design-based histomorphometry. Results showed that nerve fibers regenerated along both severed nerve trunks, and in the median nerve led to a recovery corresponding to 58% of normal nerve function. Quantitative analysis showed no significant morphological differences between myelinated nerve fibers regenerated along the two distal nerves except for the number of fibers, which was higher in the median nerve. Notably, the total number of regenerated nerve fibers in the two distal nerves was 4-fold the normal fiber number in the ulnar nerve. Besides their interest in relation to the long-lasting debate about the topographic specificity of nerve regeneration, the results of this study show an effective way to repair, in the rat experimental model, two transected nerve trunks innervating agonistic muscles in the case that the proximal stump of only one nerve is preserved., ((c) 2004 Wiley-Liss, Inc.)

Using RT-PCR, we have investigated expression of isoforms beta1 (the axonal isoform) and alpha2a-2b (the mesenchymal isoform) of neuregulin-1, one of the most important known trophic factors for Schwann cells, in the rat sciatic nerve repaired by muscle-enriched non-nervous conduits (made by a vein filled with fresh skeletal muscle). Repaired nerves were harvested 2, 6 and 13 days post-operatively. Results showed that while muscle-vein combined grafts were enriched in mRNA coding for alpha2a-2b since the very early regeneration stages, isoform beta1 mRNA was not detectable inside the tubes at day 2 and 6 post-operatively while its expression at day 13 was very slight. These results suggest that Schwann cell survival and activity inside a fresh muscle-enriched non-nervous conduit graft (a key factor for successful nerve regeneration along the graft) may be supported by the mesenchymal isoform of neuregulin-1 during very early repair phases, i.e. when axons are still not present along the tube.

Peripheral nerve researchers frequently use the rat sciatic nerve crush model in order to test different therapeutic approaches. The purpose of this study was to determine the sequence of changes after an axonotmetic injury by means of a biomechanical model of the foot and ankle, and compare them with walking track analysis, over a fixed period of time. A kinematic analysis program was used to acquire ankle motion data for further analysis. Although repeated measures analysis of variance showed significant cumulative changes induced by the crush lesion for both ankle kinematic parameters and sciatic functional index, post-hoc multiple comparisons by the Student-Neuman-Keuls test revealed significant differences between week 0 and week 8 only for ankle kinematics. These results are of importance in showing the superiority of ankle kinematics in detecting small biomechanical deficits related to hyperexcitability of the plantarflexor muscles, in contrast with walking track analysis, which showed full motor functional recovery 8 weeks after the crush lesion.

While it has been shown that terminolateral (end-to-side) neurorrhaphy leads to successful functional motor reinnervation of the peripheral territories belonging to the severed nerve, data on the morphology of terminolateral sprouting and on the voluntary control of the motor function restored by terminolateral neurorrhaphy are still partial. In this study, the severed rat median nerve was sutured in an end-to-side fashion to the intact ulnar nerve. The progression of recovery of the flexion of the fingers was assessed by means of the grasping test. Seven months after surgery, the rats were sacrificed, and morphologic and morphometric analysis was performed on the regenerated median nerve and on the donor ulnar nerve. Results of the functional assessment showed that voluntary motor control of the muscles innervated by the median nerve was partially and progressively recovered by terminolateral neurorrhaphy, with a mean strength in the flexion of the fingers that reached about 20 percent of normal before sacrifice. Morphologic and morphometric analysis showed that nerve-fiber regeneration occurred in all repaired median nerves. Signs of nerve fiber atrophy were detected in the ulnar nerve distal to the point of suture, suggesting the possible occurrence of secondary damage to the donor nerve after terminolateral neurorrhaphy that should be taken into consideration in a clinical perspective.

The search for good conduits for bridging nerve defects is a major challenge of today's tissue engineering research. In this paper we report on a laser confocal microscope study on early nerve regeneration inside a tissue engineered graft made by a poly(DLLA-epsilon-CL) conduit enriched with fresh skeletal muscle. The same biodegradable tubes filled with PBS solution were used as controls. The conduits were placed to bridge unilateral 1-cm-long rat sciatic nerve defects and analysed 10 days after surgery. Results showed that inside the muscle-enriched tubes axon regeneration, labelled by means of anti-neurofilament antibody, was already begun, whilst no axon regeneration was detectable along control tubes. In addition, a-GFAP (glial fibrillar acid protein) immuno-labelling of Schwann cells showed that progression inside muscle-enriched tubes, especially from the distal nerve stump, was much more evident than in control conduits. These results suggest that enrichment of synthetic tubes with fresh skeletal muscle promotes axon regeneration and Schwann cell migration in early nerve repair stages.

The aim of this study was to compare functional peripheral nerve recovery in the rat sciatic nerve model after reconstruction of a 10-mm gap with a biodegradable poly (DLLA-epsilon-CL) nerve guide, as filled with either fresh skeletal muscle or phosphate-buffered saline (PBS). During 24 weeks of recovery, motor and sensory functional evaluation was tested by extensor postural thrust (EPT) and withdrawal reflex latency (WRL), respectively. At the end of the experiment, anesthetized animals were prepared for motor nerve conduction velocity (MNCV) studies, followed by gastrocnemius and soleus muscle weight measurement. Motor functional recovery was greater in the muscle-grafted group, and reached a significant difference from weeks 8-12 (P < 0.05). The results of this investigation suggest that filling a nerve guide with fresh skeletal muscle induces faster maturation of regenerated nerve fibers in comparison with traditional tubular repair., (Copyright 2003 Wiley-Liss, Inc. MICROSURGERY 23:346-353 2003)

While it is well know that the percentage of stellite cells in relation to myonuclei rapidly decreases in aneurally regenerating adult muscle, the fate of satellite cells in neonatally denervated muscles has received little attention so far. In the present study, rat tibialis anterior muscles were denervated at birth and analysed after 5, 7 and 10 weeks. At least 400 myonuclei in each muscle were assessed by electron microscopy and the percentage of satellite cell nuclei in relation to the number of myonuclei was calculated. The results indicated that the percentage of satellite cells steeply declines after neonatal denervation and, after 10 weeks, satellite cells were practically lacking in the muscles under analysis. This process of exhaustion of the satellite cell pool appears to be more rapid in developing than in adult muscles.