Innervated Muscle Fibers Research Articles

175 - Effect of Partial Denervation on Mitochondrial ROS Generation in Skeletal Muscle; a Role in Sarcopenia?

With advancing age there is an increased occurrence of denervated skeletal muscle fibres, which is associated with morphological breakdown of neuromuscular junctions (NMJs). Dramatic changes in mitochondrial generation of hydrogen peroxide (H 2 O 2 ) have been reported in skeletal muscle of rodents following experimental nerve transection 1 . We hypothesised that denervation of even a limited number of fibers, such as that observed during ageing, could influence H 2 O 2 generation by both the denervated and neighbouring innervated fibers within a muscle. Thy1-YFP mice (expressing YFP in neuronal cells and so allowing visualisation of these cells only) underwent surgical transection of 1 of the 3 nerve branches entering the anterior tibialis (AT) muscle. Following recovery for up to 7 days, the AT muscles were quickly excised and samples taken for morphological assessment or for assessment of mitochondrial H 2 O 2 generation. Confocal imaging of skeletal muscle preparations allowed visualisation and mapping of NMJ integrity. Analysis revealed that at 7 days post-surgery the innervation status of individual NMJs varied across the muscle and four regions were identified in which the muscle fibres were either fully innervated, fully denervated or had a mix of innervated and denervated fibres. From each region, small bundles of fibres were removed, saponin permeablised and mitochondrial peroxide generation assessed using the Amplex Red assay. Peroxide release was assessed in state 1 respiration and in the presence of some electron transport chain substrates. Following partial denervation, oxidation of Amplex red was increased by 40-70 fold compared to sham in the bundles of fibers from all regions of the AT muscle. An increase comparable to those observed in fully denervated muscles in our previous experiments. These data show that innervated muscle fibers respond to denervation of adjacent fibers by increasing mitochondrial peroxide generation. Thus data support our hypothesis that loss of innervation in a proportion of muscle fibres is sufficient to disrupt mitochondrial H 2 O 2 generation with consequences for the entire muscle and we propose that this process is relevant to understanding muscle peroxide changes in sarcopenia. Reference Muller FL et al 2007. AM J. Physiol. Supported by the UK BBSRC and NIA (AG-20591).

Brain-derived neurotrophic factor inhibits neuromuscular junction maturation mediated by inTracellular Ca(2+) and Ca(2+)/calmodulin-dependent kinase.

Brain-derived neurotrophic factor (BDNF) inhibits neuromuscular junction (NMJ) maturation. In this study we investigated the underlying molecular mechanisms of this process. We used a patch-clamp technique to measure spontaneous synaptic currents (SSCs) from innervated muscle cells in Xenopus nerve-muscle cocultures. In the presence of Ca(2+)/calmodulin-dependent kinase (CaMK) inhibitor KN93, SSC amplitude (226.3 ± 26.5 pA), frequency (30.9 ± 10.1 events/min), and percentage of bell-shaped amplitude distributions (47.1%) were reversed to control levels (286.7 ± 48.2 pA, 26.2 ± 5.8 events/min, and 47.1%, respectively). Depletion of intracellular Ca(2+) by BAPTA-AM or thapsigargin had similar reversal effects to KN93. In addition, cotreatment with both 2-APB (IP3 receptor inhibitor) and TMB-8 (ryanodine receptor inhibitor) also reversed the inhibitory effects of BDNF, as shown by the physiological parameters. CaMK mediates the inhibitory effects of BDNF on NMJ maturation. Ca(2+) released from intracellular stores through either IP3 receptors or ryanodine receptors regulates neurotrophic actions on NMJ maturation.

Elderly mouse skeletal muscle fibres have a diminished capacity to upregulate NCAM production in response to denervation.

Sarcopenia is a major contributor to the loss of independence and deteriorating quality of life in elderly individuals, it manifests as a decline in skeletal muscle mass and strength beyond the age of 65. Muscle fibre atrophy is a major contributor to sarcopenia and the most severely atrophic fibres are commonly found in elderly muscles to have permanently lost their motor nerve input. By contrast with elderly fibres, when fibres in young animals lose their motor input they normally mount a response to induce restoration of nerve contact, and this is mediated in part by upregulated expression of the nerve cell adhesion molecule (NCAM). Therefore, skeletal muscles appear to progressively lose their ability to become reinnervated, and here we have investigated whether this decline occurs via loss of the muscle's ability to upregulate NCAM in response to denervation. We performed partial denervation (by peripheral nerve crush) of the extensor digitorum longus muscle of the lower limb in both young and elderly mice. We used immunohistochemistry to compare relative NCAM levels at denervated and control innervated muscle fibres, focused on measurements at neuromuscular junctional, extra-junctional and cytoplasmic locations. Muscle fibres in young animals responded to denervation with significant (32.9%) increases in unpolysialylated NCAM at extra-junctional locations, but with no change in polysialylated NCAM. The same partial denervation protocol applied to elderly animals resulted in no significant change in either polysialylated or unpolysialylated NCAM at junctional, extra-junctional or cytoplasmic locations, therefore muscle fibres in young mice upregulated NCAM in response to denervation but fibres in elderly mice failed to do so. Elevation of NCAM levels is likely to be an important component of the muscle fibre's ability to attract or reattract a neural input, so we conclude that the presence of increasing numbers of long-term denervated fibres in elderly muscles is due, at least in part, to the fibre's declining ability to mount a normal response to loss of motor input.

Physical Exercise in Aging: Nine Weeks of Leg Press or Electrical Stimulation Training in 70 Years Old Sedentary Elderly People.

Sarcopenia is the age-related loss of muscle mass and function, reducing force generation and mobility in the elderlies. Contributing factors include a severe decrease in both myofiber size and number as well as a decrease in the number of motor neurons innervating muscle fibers (mainly of fast type) which is sometimes accompanied by reinnervation of surviving slow type motor neurons (motor unit remodeling). Reduced mobility and functional limitations characterizing aging can promote a more sedentary lifestyle for older individuals, leading to a vicious circle further worsening muscle performance and the patients’ quality of life, predisposing them to an increased risk of disability, and mortality. Several longitudinal studies have shown that regular exercise may extend life expectancy and reduce morbidity in aging people. Based on these findings, the Interreg IVa project aimed to recruit sedentary seniors with a normal life style and to train them for 9 weeks with either leg press (LP) exercise or electrical stimulation (ES). Before and at the end of both training periods, all the subjects were submitted to mobility functional tests and muscle biopsies from the Vastus Lateralis muscles of both legs. No signs of muscle damage and/or of inflammation were observed in muscle biopsies after the training. Functional tests showed that both LP and ES induced improvements of force and mobility of the trained subjects. Morphometrical and immunofluorescent analyses performed on muscle biopsies showed that ES significantly increased the size of fast type muscle fibers (p<0.001), together with a significant increase in the number of Pax7 and NCAM positive satellite cells (p<0.005). A significant decrease of slow type fiber diameter was observed in both ES and LP trained subjects (p<0.001). Altogether these results demonstrate the effectiveness of physical exercise either voluntary (LP) or passive (ES) to improve the functional performances of aging muscles. Here ES is demonstrated to be a safe home-based method to counteract fast type fiber atrophy, typically associated with aging skeletal muscle.

Effects of IgG anti-GM1 monoclonal antibodies on neuromuscular transmission and calcium channel binding in rat neuromuscular junctions.

Guillain-Barré syndrome is a type of acute inflammatory neuropathy that causes ataxia and is associated with the IgG anti-GM1 antibody. However, the pathogenic role of the IgG anti-GM1 antibody and calcium channels in neuromuscular junctions (NMJs) remains unclear. Thus, the aim of the present study was to investigate the effects of the IgG anti-GM1 monoclonal antibody (mAb) on spontaneous muscle action potentials (SMAPs), and the effects of calcium channel blockers, in a rat spinal cord-muscle co-culture system. In addition, the binding of IgG anti-GM1 mAb to calcium channels was investigated in the rat hemidiaphragm. The frequency of SMAPs in the innervated muscle cells was acutely inhibited by the IgG anti-GM1 mAb; however, this effect was blocked by the N-type calcium channel blocker, ω-conotoxin GVIA (30 nM). Furthermore, the P/Q-type calcium channel blocker, ω-agatoxin IVA (10 nM), was found to partially block the IgG anti-GM1 mAb-induced inhibitory effect in the spinal cord-muscle co-culture system. Immunohistochemical analysis of the rat hemidiaphragm indicated that IgG anti-GM1 mAb binding overlapped with anti-Cav2.2 (α1B) antibody binding in the nerve terminal. In addition, IgG anti-GM1 mAb binding partially overlapped with anti-Cav2.1 (α1A) antibody binding. Thus, the results demonstrated that the IgG anti-GM1 mAb binds to calcium channels in the nerve terminals of NMJs. Therefore, the inhibitory effect of IgG anti-GM1 mAb on SMAPs may involve N-type and P/Q-type calcium channels in motor nerve terminals at the NMJ.

A central mesencephalic reticular formation projection to the supraoculomotor area in macaque monkeys.

The central mesencephalic reticular formation is physiologically implicated in oculomotor function and anatomically interwoven with many parts of the oculomotor system's premotor circuitry. This study in Macaca fascicularis monkeys investigates the pattern of central mesencephalic reticular formation projections to the area in and around the extraocular motor nuclei, with special emphasis on the supraoculomotor area. It also examines the location of the cells responsible for this projection. Injections of biotinylated dextran amine were stereotaxically placed within the central mesencephalic reticular formation to anterogradely label axons and terminals. These revealed bilateral terminal fields in the supraoculomotor area. In addition, dense terminations were found in both the preganglionic Edinger-Westphal nuclei. The dense terminations just dorsal to the oculomotor nucleus overlap with the location of the C-group medial rectus motoneurons projecting to multiply innervated muscle fibers suggesting they may be targeted. Minor terminal fields were observed bilaterally within the borders of the oculomotor and abducens nuclei. Injections including the supraoculomotor area and oculomotor nucleus retrogradely labeled a tight band of neurons crossing the central third of the central mesencephalic reticular formation at all rostrocaudal levels, indicating a subregion of the nucleus provides this projection. Thus, these experiments reveal that a subregion of the central mesencephalic reticular formation may directly project to motoneurons in the oculomotor and abducens nuclei, as well as to preganglionic neurons controlling the tone of intraocular muscles. This pattern of projections suggests an as yet undetermined role in regulating the near triad.

Effect of Muscle Fiber Denervation on Mitochondrial Hydrogen Peroxide Generation in Adjacent Innervated Muscle Fibres: Role in Sarcopenia?

Nerve transection causes dramatic increases in mitochondrial generation of hydrogen peroxide (H2O2) in the denervated skeletal muscle of rodents. We hypothesized that denervation of a limited number of fibers, such as that observed in muscles during aging, could influence H2O2 generation by both denervated and innervated fibers throughout the muscle. Thy1‐YFP mice (expressing YFP in neurons) underwent full transection of the peroneal nerve or transection of only one of the 3 branches entering the anterior tibialis (AT) muscle. Following up to 10days recovery, generation of H2O2 by mitochondria in permeablised fibers with intact mitochondra from different portions of the AT was assessed using Amplex Red. Results demonstrated increased H2O2 generation by 3days after full transection compared with sham control, which remained elevated at 10days. In situ imaging showed loss of innervation of endplates with no evidence of post‐synaptic changes. Immuno‐fluorescent staining for markers of denervation showed neuronal cell adhesion molecule positive muscle fibers by 7days post‐surgery. Transection of the single branch of the peroneal nerve caused denervation of a portion of the AT and mitochondrial H2O2 generation in the denervated portion was elevated by 3days post‐transection in a similar manner to the fully denervated muscle. Denervation of some just fibers was associated with increased mitochondrial H2O2 generation in all areas of the muscle. Thus data support the hypothesis that the effect of partial denervation on muscle H2O2 generation is mirrored in fully innervated fibers of the same muscle, an effect which may be mirrored in sarcopenia. Supported by the UK Biotechnology and Biological Sciences Research Council.

The extraction of neural strategies from the surface EMG: an update.

A surface EMG signal represents the linear transformation of motor neuron discharge times by the compound action potentials of the innervated muscle fibers and is often used as a source of information about neural activation of muscle. However, retrieving the embedded neural code from a surface EMG signal is extremely challenging. Most studies use indirect approaches in which selected features of the signal are interpreted as indicating certain characteristics of the neural code. These indirect associations are constrained by limitations that have been detailed previously (Farina D, Merletti R, Enoka RM. J Appl Physiol 96: 1486-1495, 2004) and are generally difficult to overcome. In an update on these issues, the current review extends the discussion to EMG-based coherence methods for assessing neural connectivity. We focus first on EMG amplitude cancellation, which intrinsically limits the association between EMG amplitude and the intensity of the neural activation and then discuss the limitations of coherence methods (EEG-EMG, EMG-EMG) as a way to assess the strength of the transmission of synaptic inputs into trains of motor unit action potentials. The debated influence of rectification on EMG spectral analysis and coherence measures is also discussed. Alternatively, there have been a number of attempts to identify the neural information directly by decomposing surface EMG signals into the discharge times of motor unit action potentials. The application of this approach is extremely powerful, but validation remains a central issue.

The Neuromuscular Junction: Aging at the Crossroad between Nerves and Muscle.

Aging is associated with a progressive loss of muscle mass and strength and a decline in neurophysiological functions. Age-related neuromuscular junction (NMJ) plays a key role in musculoskeletal impairment that occurs with aging. However, whether changes in the NMJ precede or follow the decline of muscle mass and strength remains unresolved. Many factors such as mitochondrial dysfunction, oxidative stress, inflammation, changes in the innervation of muscle fibers, and mechanical properties of the motor units probably perform an important role in NMJ degeneration and muscle mass and strength decline in late life. This review addresses the primary events that might lead to NMJ dysfunction with aging, including studies on biomarkers, signaling pathways, and animal models. Interventions such as caloric restriction and exercise may positively affect the NMJ through this mechanism and attenuate the age-related progressive impairment in motor function.

P437: Different types of fibrillation potentials in human needle EMG

Rhythmic fibrillation potentials are the hallmark of denervated muscle fibres in needle EMG of a striated muscle (Conrad et al. 1972, Heckmann & Ludin 1982). They are readily activated by the insertion of an EMG needle electrode (Kugelberg & Petersen 1949). Irregular fibrillation potentials may also be present (Buchthal & Rosenfalck 1966, Purves & Sakmann 1974). There is, however, an obvious difficulty to discriminate between true irregular fibrillation potentials of a denervated muscle and end plate spikes, which occur in a normal muscle. This may lead to the conclusion that only rhythmic fibrillation potentials matter (Stohr 1977). We have pointed out that fibrillation potentials, whether regular or irregular, have longer minimum interpotential intervals than end plate spikes (Partanen & Danner 1982). During long sequences, the mean interval between successive end plate spikes and rhythmic fibrillation potential tends to increase, whereas irregular fibrillations do not show this type of “self-inhibition” (Partanen & Danner 1982, Partanen & Nousiainen 1983). The aim of this chapter is to describe fibrillation potentials of different categories in either completely or partially denervated human limb muscles, or after a muscle injury. We also compare the characteristics of fibrillation potentials to neurally driven sequences, such as “myokymic” fibrillation potentials and end plate spikes (Brown & Varkey 1981, Partanen & Nousiainen 1983, Partanen 1999). “Myokymic” fibrillation potentials are a rare phenomenon of innervated muscle fibres. They have not been described earlier and are readily confused with end plate spikes. The term “myokymic” fibrillations is descriptive. The pathophysiology of “myokymic fibrillation” is different from true myokymia of whole motor units (see Willison 1982, Stalberg & Trontelj 1982).

Autologous minced muscle grafts: a tissue engineering therapy for the volumetric loss of skeletal muscle

Volumetric muscle loss (VML) results in a large void deficient in the requisite materials for regeneration for which there is no definitive clinical standard of care. Autologous minced muscle grafts (MG), which contain the essential components for muscle regeneration, may embody an ideal tissue engineering therapy for VML. The purpose of this study was to determine if orthotopic transplantation of MG acutely after VML in the tibialis anterior muscle of male Lewis rats promotes functional tissue regeneration. Herein we report that over the first 16 wk postinjury, MG transplantation 1) promotes remarkable regeneration of innervated muscle fibers within the defect area (i.e., de novo muscle fiber regeneration); 2) reduced evidence of chronic injury in the remaining muscle mass compared with nonrepaired muscles following VML (i.e., transplantation attenuated chronically upregulated transforming growth factor-β1 gene expression and the presence of centrally located nuclei in 30% of fibers observed in nonrepaired muscles); and 3) significantly improves net torque production (i.e., ∼55% of the functional deficit in nonrepaired muscles was restored). Additionally, voluntary wheel running was shown to reduce the heightened accumulation of extracellular matrix deposition observed within the regenerated tissue of MG-repaired sedentary rats 8 wk postinjury (collagen 1% area: sedentary vs. runner, ∼41 vs. 30%), which may have been the result of an augmented inflammatory response [i.e., M1 (CCR7) and M2 (CD163) macrophage expression was significantly greater in runner than sedentary MG-repaired muscles 2 wk postinjury]. These findings support further exploration of autologous minced MGs for the treatment of VML.

Neurophysiological and immunohistochemical studies of IgG anti-GM1 monoclonal antibody on neuromuscular transmission: effects in rat neuromuscular junctions

Guillain-Barré syndrome, which is a variant of acute inflammatory neuropathy, is associated with anti-GM1 antibodies and causes ataxia. We investigated the effects of IgG anti-GM1 monoclonal antibody (IgG anti-GM1 mAb) on spontaneous muscle action potentials in a rat spinal cord-muscle co-culture system and the localization of IgG anti-GM1 mAb binding in the rat hemi-diaphragm. The frequency of spontaneous muscle action potentials in innervated muscle cells was acutely inhibited by IgG anti-GM1 mAb. When cultures were pretreated with GM2 synthase antisense oligodeoxynucleotide, IgG anti-GM1 mAb failed to inhibit spontaneous muscle action potentials, demonstrating the importance of the GM1 epitope in the action of IgG anti-GM1 mAb. Immunohistochemistry of rat hemi-diaphragm showed that IgG anti-GM1 mAb binding overlapped with neurofilament 200 (NF200) antibodies staining, but not α-bungarotoxin (α-BuTx) staining, demonstrating that IgG anti-GM1 mAb was localized at the presynaptic nerve terminal. IgG anti-GM1 mAb binding overlapped with syntaxin antibody and S-100 antibody in the nerve terminal. After collagenase treatment, IgG anti-GM1 mAb and NF200 antibodies did not show staining, but α-BuTx selectively stained the hemi-diaphragm. IgG anti-GM1 mAb binds to the presynaptic nerve terminal of neuromuscular junctions. Therefore, we suggest that the inhibitory effect of IgG anti-GM1 mAb on spontaneous muscle action potentials is related to the GM1 epitope in presynaptic motor nerve terminals at the NMJs.

Palisade endings and proprioception in extraocular muscles: a comparison with skeletal muscles

This article describes current views on motor and sensory control of extraocular muscles (EOMs) based on anatomical data. The special morphology of EOMs, including their motor innervation, is described in comparison to classical skeletal limb and trunk muscles. The presence of proprioceptive organs is reviewed with emphasis on the palisade endings (PEs), which are unique to EOMs, but the function of which is still debated. In consideration of the current new anatomical data about the location of cell bodies of PEs, a hypothesis on the function of PEs in EOMs and the multiply innervated muscle fibres they are attached to is put forward.

Muscle Precursor Cells for the Restoration of Irreversibly Damaged Sphincter Function

Multiple modalities, including injectable bulking agents and surgery, have been used to treat stress urinary incontinence. However, none of these methods is able to fully restore normal striated sphincter muscle function. In this study, we explored the possibility of achieving functional recovery of the urinary sphincter muscle using autologous muscle precursor cells (MPCs) as an injectable, cell-based therapy. A canine model of striated urinary sphincter insufficiency was created by microsurgically removing part of the sphincter muscle in 24 dogs. Autologous MPCs were obtained, expanded in culture, and injected into the damaged sphincter muscles of 12 animals. The animals were followed for up to 6 months after injection, and urodynamic studies, functional organ bath studies, ultrastructural and histological examinations were performed. Animals receiving MPC injections demonstrated sphincter pressures of approximately 80% of normal values, while the pressures in the control animals without cells dropped and remained at 20% of normal values. Histological analysis indicated that the implanted cells survived and formed tissue, including new innervated muscle fibers, within the injected region of the sphincter. These results indicate that autologous muscle precursor cells may be able to restore otherwise irreversibly damaged urinary sphincter function clinically.

Prevalence and elimination of sibling neurite convergence in motor units supplying neonatal and adult mouse skeletal muscle

During development, neurons form supernumerary synapses, most of which are selectively pruned leading to stereotyped patterns of innervation. During the development of skeletal muscle innervation, or its regeneration after nerve injury, each muscle fiber is transiently innervated by multiple motor axon branches but eventually by a single branch. The selective elimination of all but one branch is the result of competition between the converging arbors. It is thought that motor neurons initially innervate muscle fibers randomly, but that axon branches from the same neuron (sibling branches) do not converge to innervate the same muscle fiber. However, random innervation would result in many neonatal endplates that are co-innervated by sibling branches. To investigate whether this occurs we examined neonatal levator auris longus (LAL) and 4th deep lumbrical (4DL) muscles, as well as adult reinnervated deep lumbrical muscles (1-4) in transgenic mice expressing yellow fluorescent protein (YFP) as a reporter. We provide direct evidence of convergence of sibling neurites within single fluorescent motor units, both during development and during regeneration after nerve crush. The incidence of sibling neurite convergence was 40% lower in regeneration and at least 75% lower during development than expected by chance. Therefore, there must be a mechanism that decreases the probability of its occurrence. As sibling neurite convergence is not seen in normal adults, or at later timepoints in regeneration, synapse elimination must also remove convergent synaptic inputs derived from the same motor neuron. Mechanistic theories of synaptic competition should now accommodate this form of isoaxonal plasticity.

Denervation Causes Fiber Atrophy and Myosin Heavy Chain Co-Expression in Senescent Skeletal Muscle

Although denervation has long been implicated in aging muscle, the degree to which it is causes the fiber atrophy seen in aging muscle is unknown. To address this question, we quantified motoneuron soma counts in the lumbar spinal cord using choline acetyl transferase immunhistochemistry and quantified the size of denervated versus innervated muscle fibers in the gastrocnemius muscle using the in situ expression of the denervation-specific sodium channel, Nav1.5, in young adult (YA) and senescent (SEN) rats. To gain insights into the mechanisms driving myofiber atrophy, we also examined the myofiber expression of the two primary ubiquitin ligases necessary for muscle atrophy (MAFbx, MuRF1). MN soma number in lumbar spinal cord declined 27% between YA (638±34 MNs×mm−1) and SEN (469±13 MNs×mm−1). Nav1.5 positive fibers (1548±70 μm2) were 35% smaller than Nav1.5 negative fibers (2367±78 μm2; P<0.05) in SEN muscle, whereas Nav1.5 negative fibers in SEN were only 7% smaller than fibers in YA (2553±33 μm2; P<0.05) where no Nav1.5 labeling was seen, suggesting denervation is the primary cause of aging myofiber atrophy. Nav1.5 positive fibers had higher levels of MAFbx and MuRF1 (P<0.05), consistent with involvement of the proteasome proteolytic pathway in the atrophy of denervated muscle fibers in aging muscle. In summary, our study provides the first quantitative assessment of the contribution of denervation to myofiber atrophy in aging muscle, suggesting it explains the majority of the atrophy we observed. This striking result suggests a renewed focus should be placed on denervation in seeking understanding of the causes of and treatments for aging muscle atrophy.

Reinnervation by the contralateral facial nerve in patients with peripheral facial palsy

Reinnervation activity is triggered after complete unilateral peripheral facial palsy (PFP). In 27 patients with PFP we recorded electromyographic activity with a concentric needle electrode inserted 1 cm lateral to the oral commissure of the affected side. We applied electrical stimuli to the unaffected (contralateral) facial nerve from the tragus to the mid-lower lip and measured the response latency variability and segmental conduction velocity. Responses to electrical stimulation of the unaffected facial nerve were found in all patients. Mean conduction velocity was 49.6 ± 6.2 m/s between tragus and oral commissure, and 6.0 ± 1.9 m/s between oral commissure and mid-lower lip. Latency variability was 0.27 ms to facial nerve stimulation and 0.08 ms to oral commissure stimulation. Short distance sprouting of axons that innervate muscle fibers, which originate from the unaffected facial nerve, results in propagation of impulses to muscle fibers in the midline.

Is there any sense in the Palisade endings of eye muscles?

Palisade endings (PEs), which are unique to the eye muscles, are associated with multiply innervated muscle fibers. They lie at the myotendinous junctions and form a cap around the muscle fiber tip. They are found in all animals investigated so far, but their function is not known. Recently, we demonstrated that cell bodies of PEs and tendon organs lie around the periphery of the oculomotor nucleus in the C- and S-groups. A morphological analysis of these peripheral neurons revealed the existence of different populations within the C-group. We propose that a small group of round or spindle-shaped cells gives rise to PEs, and another group of multipolar neurons provide the multiple motor endings. If PEs have a sensory function, then their cell body location close to motor neurons would be in an ideal location to control tension in extraocular muscles; in the case of the C-group, its proximity to the preganglionic neurons of the Edinger-Westphal nucleus would permit its participation in the near response. Despite their unusual properties, PEs may have a sensory function.

Motor nucleus activity fails to predict extraocular muscle forces in ocular convergence

For a given eye position, firing rates of abducens neurons (ABNs) generally (Mays et al. 1984), and lateral rectus (LR) motoneurons (MNs) in particular (Gamlin et al. 1989a), are higher in converged gaze than when convergence is relaxed, whereas LR and medial rectus (MR) muscle forces are slightly lower (Miller et al. 2002). Here, we confirm this finding for ABNs, report a similarly paradoxical finding for neurons in the MR subdivision of the oculomotor nucleus (MRNs), and, for the first time, simultaneously confirm the opposing sides of these paradoxes by recording physiological LR and MR forces. Four trained rhesus monkeys with binocular eye coils and custom muscle force transducers on the horizontal recti of one eye fixated near and far targets, making conjugate saccades and symmetric and asymmetric vergence movements of 16-27°. Consistent with earlier findings, we found in 44 ABNs that the slope of the rate-position relationship for symmetric vergence (k(V)) was lower than that for conjugate movement (k(C)) at distance, i.e., mean k(V)/k(C) = 0.50, which implies stronger LR innervation in convergence. We also found in 39 MRNs that mean k(V)/k(C) = 1.53, implying stronger MR innervation in convergence as well. Despite there being stronger innervation in convergence at a given eye position, we found both LR and MR muscle forces to be slightly lower in convergence, -0.40 and -0.20 g, respectively. We conclude that the relationship of ensemble MN activity to total oculorotary muscle force is different in converged gaze than when convergence is relaxed. We conjecture that LRMNs with k(V) < k(C) and MRMNs with k(V) > k(C) innervate muscle fibers that are weak, have mechanical coupling that attenuates their effective oculorotary force, or serve some nonoculorotary, regulatory function.

Evidence that the extraocular motor nuclei innervate monkey palisade endings

Palisade endings are found in the extraocular muscles (EOMs) of almost every mammalian species, including primates. These nerve specializations surrounding the muscle fiber insertion have been postulated to be the proprioceptors of the EOMs. However, it was recently demonstrated that palisade endings have a cholinergic nature, which reopened the question of whether palisade endings are motor or sensory structures. In this work, we examined whether the cell bodies of palisade endings lie in EOM motor nuclei by injecting an anterograde tracer, biotinylated dextran amine, into the abducens nucleus of a macaque monkey. Tracer visualization in the lateral rectus muscle was combined with choline acetyltransferase (ChAT) and α-bungarotoxin staining. Analysis of the samples was performed by conventional light microscopy and confocal laser scanning microscopy. About 30% of the nerve fibers innervating the muscle were tracer positive. These were ChAT positive as well. Tracer positive nerve fibers established motor contacts on singly and multiply innervated muscle fibers, which were confirmed by α-bungarotoxin staining. At the transition between muscle and distal tendon, we found palisade endings that contained tracer. Palisade endings exhibited the classic morphology: axons arising from the muscle extend onto the tendon, then turn back 180° and terminate in a cuff of terminals around an individual muscle fiber tip. This finding suggests that the cell bodies of palisade endings lie in the EOM motor nuclei, which complements prior studies demonstrating a cholinergic, and possibly motor, phenotype for palisade endings.