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The cellular basis of age-related behavioral decline remains obscure but alterations in synapses are likely candidates. Accordingly, the beneficial effects on neural function of caloric restriction and exercise, which are among the most effective anti-aging treatments known, might also be mediated by synapses. As a starting point in testing these ideas, we studied the skeletal neuromuscular junction (NMJ), a large, accessible peripheral synapse. Comparison of NMJs in young adult and aged mice revealed a variety of age-related structural alterations, including axonal swellings, sprouting, synaptic detachment, partial or complete withdrawal of axons from some postsynaptic sites, and fragmentation of the postsynaptic specialization. Alterations were significant by 18 mo of age and severe by 24 mo. A life-long calorie-restricted diet significantly decreased the incidence of pre- and postsynaptic abnormalities in 24-mo-old mice and attenuated age-related loss of motor neurons and turnover of muscle fibers. One month of exercise (wheel running) in 22-mo-old mice also reduced age-related synaptic changes but had no effect on motor neuron number or muscle fiber turnover. Time-lapse imaging in vivo revealed that exercise partially reversed synaptic alterations that had already occurred. These results demonstrate a critical effect of aging on synaptic structure and provide evidence that interventions capable of extending health span and lifespan can partially reverse these age-related synaptic changes. aging | neuromuscular junction | muscle | motor neuron | sarcopenia doi/ 10.1073/pnas.1002220107

Defective RNA metabolism is an emerging mechanism involved in ALS pathogenesis and possibly in other neurodegenerative disorders. Here, we show that microRNA (miRNA) activity is essential for long-term survival of postmitotic spinal motor neurons (SMNs) in vivo. Thus, mice that do not process miRNA in SMNs exhibit hallmarks of spinal muscular atrophy (SMA), including sclerosis of the spinal cord ventral horns, aberrant end plate architecture, and myofiber atrophy with signs of denervation. Furthermore, a neurofilament heavy subunit previously implicated in motor neuron degeneration is specifically up-regulated in miRNA-deficient SMNs. We demonstrate that the heavy neurofilament subunit is a target of miR-9, a miRNA that is specifically down-regulated in a genetic model of SMA. These data provide evidence for miRNA function in SMN diseases and emphasize the potential role of miR-9--based regulatory mechanisms in adult neurons and neurodegenerative states. ALS | Dicer | microRNA | motor neuron | neurodegeneration doi/ 10.1073/pnas.1006151107

For the promise of human induced pluripotent stem cells (iPSCs) to be realized, it is necessary to ask if and how efficiently they may be differentiated to functional cells of various lineages. Here, we have directly compared the neural-differentiation capacity of human iPSCs and embryonic stem cells (ESCs). We have shown that human iPSCs use the same transcriptional network to generate neuroepithelia and functionally appropriate neuronal types over the same developmental time course as hESCs in response to the same set of morphogens; however, they do it with significantly reduced efficiency and increased variability. These results were consistent across iPSC lines and independent of the set of reprogramming transgenes used to derive iPSCs as well as the presence or absence of reprogramming transgenes in iPSCs. These findings, which show a need for improving differentiation potency of iPSCs, suggest the possibility of employing human iPSCs in pathological studies, therapeutic screening, and autologous cell transplantation. neural development | regeneration | reprogramming | transplantation | motor neurons doi/10.1073/pnas.0910012107

Production of voluntary movements relies critically on the functional integration of several motor cortical areas, such as the primary motor cortex, and the spinal circuitries. Surprisingly, after almost 40 years of research, how the motor cortices specify descending neural signals destined for the downstream interneurons and motoneurons has remained elusive. In light of the many recent experimental demonstrations that the motor system may coordinate muscle activations through a linear combination of muscle synergies, we hypothesize that the motor cortices may function to select and activate fixed muscle synergies specified by the spinal or brainstem networks. To test this hypothesis, we recorded electromyograms (EMGs) from 12-16 upper arm and shoulder muscles from both the unaffected and the stroke-affected arms of stroke patients having moderate-to-severe unilateral ischemic lesions in the frontal motor cortical areas. Analyses of EMGs using a nonnegative matrix factorization algorithm revealed that in seven of eight patients the muscular compositions of the synergies for both the unaffected and the affected arms were strikingly similar to each other despite differences in motor performance between the arms, and differences in cerebral lesion sizes and locations between patients. This robustness of muscle synergies that we observed supports the notion that descending cortical signals represent neuronal drives that select, activate, and flexibly combine muscle synergies specified by networks in the spinal cord and/or brainstem. Our conclusion also suggests an approach to stroke rehabilitation by focusing on those synergies with altered activations after stroke. motor control | motor modules | neurorehabilitation | motor primitives | electromyography www.pnas.org/cgi/doi/10.1073/pnas.0910114106

Pompe disease is a severe form of muscular dystrophy due to glycogen accumulation in all tissues, especially striated muscle. Disease severity is directly related to the deficiency of acid [alpha]-glucosidase (GAA), which degrades glycogen in the lysosome. Respiratory dysfunction is a hallmark of the disease, muscle weakness has been viewed as the underlying cause, and the possibility of an associated neural contribution has not been evaluated previously. Therefore, we examined behavioral and neurophysiological aspects of breathing in 2 animal models of Pompe disease--the [Gaa.sup.-/-] mouse and a transgenic line (MTP) expressing GAA only in skeletal muscle, as well as a detailed analysis of the CNS in a Pompe disease patient. Glycogen content was elevated in the [Gaa.sup.-/-] mouse cervical spinal cord. Retrograde labeling of phrenic motoneurons showed significantly greater soma size in [Gaa.sup.-/-] mice vs. isogenic controls, and glycogen was observed in [Gaa.sup.-/-] phrenic motoneurons. Ventilation, assessed via plethysmography, was attenuated during quiet breathing and hypercapnic challenge in [Gaa.sup.-/-] mice (6 to >21 months of age) vs. controls. We confirmed that MTP mice had normal diaphragmatic contractile properties; however, MTP mice had ventilation similar to the [Gaa.sup.-/-] mice during quiet breathing. Neurophysiological recordings indicated that efferent phrenic nerve inspiratory burst amplitudes were substantially lower in [Gaa.sup.-/-] and MTP mice vs. controls. In human samples, we demonstrated similar pathology in the cervical spinal cord and greater accumulation of glycogen in spinal cord compared with brain. We conclude that neural output to the diaphragm is deficient in [Gaa.sup.-/-] mice, and therapies targeting muscle alone may be ineffective in Pompe disease. glycogenosis | motor neuron | muscular dystrophy | myopathy

This paper presents an algorithm to resolve superimposed action potentials encountered during the decomposition of electromyographic signals. The algorithm uses particle swarm optimization with a variety of features including randomization, crossover, and multiple swarms. In a simulation study involving realistic superpositions of two to five motor-unit action potentials, the algorithm had an accuracy of 98%. Index Terms--Alignment, decomposition, electromyography, particle swarm optimization, superposition.

The extensive alternative splicing in higher eukaryotes has initiated a debate whether alternative mRNA isoforms are generated by an inaccurate spliceosome or are the consequence of highly degenerate splice sites within the human genome. Here, we established a quantitative assay to evaluate the accuracy of splicesite pairing by determining the number of incorrect exon-skipping events made from constitutively spliced pre-mRNA transcripts. We demonstrate that the spliceosome pairs exons with an astonishingly high degree of accuracy that may be limited by the quality of pre-mRNAs generated by RNA pol II. The error rate of exon pairing is increased by the effects of the neurodegenerative disorder spinal muscular atrophy because of reduced levels of Survival of Motor Neuron, a master assembler of spliceosomal components. We conclude that all multi-intron-containing genes are alternatively spliced and that the reduction of SMN results in a general splicing defect that is mediated through alterations in the fidelity of splice-site pairing. pre-mRNA splicing | spinal muscular atrophy | splice-site pairing | splicing fidelity | survival of motor neuron

[Ca.sup.2+] can stimulate cyclic nucleotide synthesis, but it is not known whether this signaling occurs in nerve terminals in response to activity. Here, in vivo imaging of Drosophila motoneuron terminals shows that activity rapidly induces a long-lasting signal from a transgenically expressed optical indicator based on the epac1 (exchange protein directly activated by cAMP 1) cAMP-binding domain. The epac1-cAMP sensor (camps) response in synaptic boutons depends on extracellular [Ca.sup.2+] and ryanodine receptor-mediated [Ca.sup.2+]-induced [Ca.sup.2+] release from the endoplasmic reticulum. However, mutations that inhibit rutabaga [Ca.sup.2+]-stimulated adenylyl cyclase and dunce cAMP-specific phosphodiesterase (PDE) have no effect. Instead, the activity-dependent presynaptic epac1-camps signal reflects elevation of cGMP in response to nitric oxide-activated guanylyl cyclase. Posttetanic presynaptic cGMP is long-lived because of limited PDE activity. Thus, nerve terminal biochemical signaling induced by brief bouts of activity temporally summates on a time scale orders of magnitude longer than fast transmission. cyclases | cyclic nucleotides | nerve terminal | plasticity | neuromuscular junction

Dominant mutations in cytoplasmic dynein (Loa or Cra) have been reported to provoke selective, age-dependent killing of motor neurons, while paradoxically slowing degeneration and death of motor neurons in one mouse model of an inherited form of ALS. Examination of Loa animals reveals no degeneration of large caliber [alpha]-motor neurons beyond an age-dependent loss (initiating only after 18 months) that was comparable in Loa and wild-type littermates. Absence of Loa-mediated [alpha]-motor neuron loss contrasted with dramatic, sustained, mutant dynein-mediated postnatal loss of lumbar proprioceptive sensory axons, accompanied by decreased excitatory glutamatergic inputs to motor neurons. In mouse models of inherited ALS caused by mutations in superoxide dismutase (SOD1), mutant dynein modestly prolonged survival in the one mouse model with the most extensive motor neuron loss ([SOD.sup.G93A]) while showing marginal ([SOD.sup.G85R]) or no ([SOD.sup.G37R]) benefit in models with higher numbers of surviving motor neurons at end stage. These findings support a noncell autonomous, excitotoxic contribution from proprioceptive sensory neurons that modestly accelerates disease onset in inherited ALS. cytoplasmic dynein | excitotoxicity | noncell autonomous | sensory neurons

A large number of cytokines and growth factors support the development and subsequent maintenance of postnatal motor neurons. RegIII[beta] also known as Reg2 in rat and HIP/PAP1 in humans, is a member of a family of growth factors found in many areas of the body and previously shown to play an important role in both the development and regeneration of subsets of motor neurons. It has been suggested that RegIII[beta] expressed by motor neurons is both an obligatory intermediate in the downstream signaling of the leukemia inhibitory factor/ciliary neurotrophic factor (CNTF) family of cytokines, maintaining the integrity of motor neurons during development, as well as a powerful influence on Schwann cell growth during regeneration of the peripheral nerve. Here we report that in mice with a deletion of the Reglll[beta] gene, motor neuron survival was unaffected up to 28 weeks after birth. However, there was no CNTF-mediated rescue of neonatal facial motor neurons after axotomy in KO animals when compared with wild-type. In mice, Reglll[beta] positive motor neurons are concentrated in cranial motor nuclei that are involved in the patterning of swallowing and suckling. We found that suckling was impaired in Reglll[beta] KO mice and correlated this with a significant delay in myelination of the hypoglossal nerve. In summary, we propose that Reglll[beta] has an important role to play in the developmental fine-tuning of neonatal motor behaviors mediating the response to peripherally derived cytokines and growth factors and regulating the myelination of motor axons. Schwann cells | suckling | hypoglossal nerve | LIF

Individual spinal motor neuron identities are specified in large part by the intrinsic repertoire of transcription factors expressed by undifferentiated progenitors and maturing neurons. It is shown here that the transcription factor AML1/Runx1 (Runx1) is expressed in selected spinal motor neuron subtypes after the onset of differentiation and is both necessary and sufficient to suppress interneuron-specific developmental programs and promote maintenance of motor neuron characteristics. These findings show an important role for Runx1 during the consolidation of selected spinal motor neuron identities. Moreover, they suggest a requirement for a persistent suppression of interneuron genes within maturing motor neurons. lateral motor column | median motor column | runt | spinal cord spinal accessory column

Amyotrophic lateral sclerosis (ALS) is a neuro-degenerative disease where motor neurons within the brain and spinal cord are lost, leading to paralysis and death. Certain growth factors should, in principle, be able to protect dying motor neurons. However, targeted delivery to the spinal cord or brain has been a constant problem. There is also accumulating evidence that glial cells might play a crucial role in maintaining motor neuron function and survival in ALS. Stem cells isolated and expanded in culture can be modified to release growth factors and generate glial cells following transplantation into the spinal cord or brain. As such, they might be able to both detoxify the local environment around dying motor neurons and deliver trophic factors. Here we examine the feasibility of translating these findings into new treatments for ALS patients.

Understanding transdifferentiation--the conversion of one differentiated cell type into another--is important from both basic science and clinical perspectives. In Caenorhabditis elegans, an epithelial cell named Y is initially part of the rectum but later appears to withdraw, migrate, and then become a motor neuron named PDA. Here, we show that this represents a bona fide transdifferentiation event: Y has epithelial hallmarks without detectable neural characteristics, and PDA has no residual epithelial characteristics. Using available mutants and laser microsurgery, we found that transdifferentiation does not depend on fusion with a neighboring cell or require migration of Y away from the rectum, that other rectal epithelial cells are not competent to transdifferentiate, and that transdifferentiation requires the EGL-5 and SEM-4 transcription factors and LIN-12/Notch signaling. Our results establish Y-to-PDA transdifferentiation as a genetically tractable model for deciphering the mechanisms underlying cellular plasticity in vivo. cell plasticity | motor neuron | rectum | hindgut

Experiments in monkeys demonstrated that many parietal and premotor neurons coding a specific motor act (e.g., grasping) show a markedly different activation when this act is part of actions that have different goals (e.g., grasping for eating vs. grasping for placing). Many of these 'action-constrained' neurons have mirror properties firing selectively to the observation of the initial motor act of the actions to which they belong motorically. By activating a specific action chain from its very outset, this mechanism allows the observers to have an internal copy of the whole action before its execution, thus enabling them to understand directly the agent's intention. Using electromyographic recordings, we show that a similar chained organization exists in typically developing children, whereas it is impaired in children with autism. We propose that, as a consequence of this functional impairment, high-functioning autistic children may understand the intentions of others cognitively but lack the mechanism for understanding them experientially. mirror neurons | motor chains | motor acts | goal understanding | motor intention

Neurotrophins are potent survival factors for developing and injured neurons. However, they are not being used to treat neurodegenerarive diseases because of difficulties in administration and numerous side effects that have been encountered in previous clinical trials. Their biological activities use Trk (tropomyosin-related kinase) transmembrane tyrosine kinases. Therefore, one alternative approach is to use transactivation pathways such as adenosine 2A receptor agonists, which can activate Trk receptor signaling independent of neurotrophin binding. However, the relevance in vivo and applicability of these transactivation events during neurodegenerative and injury conditions have never been extensively studied. Here we demonstrate that motoneuron survival after facial nerve lesioning is significantly enhanced by transactivation of Trk receptor tyrosine kinases by adenosine agonists. Moreover, survival of motoneurons directly required the activation of the BDNF receptor TrkB and an increase in Akt (AKT8 virus oncogene cellular homolog) activity. The ability of small molecules to activate atrophic response by using Trk signaling provides a unique mechanism to promote survival signals in motoneurons and suggests new strategies for using transactivation in neurodegenerative diseases. brain-derived neurotrophic factor | transactivation

Recently, mutations in several genes have been identified as primary causes for the degeneration of motoneurons and their axons. Strikingly, mutations in the same genes were associated with clinically different motoneuron syndromes. The identity of these genes also shed light on the mechanisms of motoneuron degeneration and revealed that overlapping motoneuron phenotypes might be caused by heterogeneous molecular mechanisms. Overall, these findings have challenged the diagnostic classification system set by clinical judgement and triggered the notion of heterogeneity in motoneuron disease. It will now be especially relevant to identify the mechanisms and principles that motoneuron diseases have in common, as this will allow us to identify the most relevant therapeutic targets. On the other hand, heterogeneity in motoneuron disease also implies that finding a monotherapy cure for motoneuron disease will be challenging and that pre-clinical testing of therapeutic targets should not be limited to a single animal model.

The neural control of locomotion involves a constant interplay between the actions of a central pattern generator (CPG) and sensory input elicited by bodily movement. With respect to the CPG, recent analysis of fictive locomotion has shown that durations of flexion and extension tend to covary along specific lines in plots of phase duration versus cycle duration. The slopes of these lines evidently depend on internal states that vary among preparations, but, within a preparation, remain rather steady from one sequence to the next. These relationships can be reproduced in a simple oscillator model having two pairs of preset parameters, suggesting that steady internal drives to flexor and extensor half-centers determine how phase durations covary. Regarding the role of sensory inputs, previous experiments have revealed state-dependent rules that govern phase-switching independently of the CPG rhythm. In addition, sensory input is known to modulate motoneuronal activation through stretch reflexes. To explore how sensory input combines with the locomotor CPG, we used a neuromechanical model with muscle actuators, proprioceptive feedback, sensory phase-switching rules, and a CPG. Interestingly, sequences of stable locomotion were always associated with phase durations that conformed to an extensor-dominated phase-duration characteristic (where extension durations vary more than flexion durations). This is the characteristic seen in normal animals, but not necessarily in fictive locomotion, where movement and associated sensory input are absent. This suggests that to produce the biomechanical events required for stability, an extensor-dominated phase-duration characteristic is required. In the model, when the preset CPG phase durations were well matched to coincide the biomechanical requirements, CPG-mediated phase switching produced stable cycles. When CPG phase durations were too short, phases switched prematurely and the model soon fell. When CPG phase durations were too long, sensory rules fired and overrode the CPG, maintaining stability. We posit that under normal circumstances, descending input from higher centers continually adjusts the operating point of the CPG on the preset phase-duration characteristic according to anticipated biomechanical requirements. When the predictions are good, CPG-generated phase durations closely match those required by the kinetics and kinematics, and little or no sensory adjustment occurs. We propose the term 'neuromechanical tuning' to describe this process of matching the CPG to the biomechanical requirements.

Catecholaminergic neurons of the A2 area play a prominent role in brain stem vagal circuits. It is not clear, however, whether these neurons are noradrenergic or adrenergic, i.e., display tyrosine hydroxylase (TH) and dopamine-[beta]-hydroxylase (D[beta]H) immunoreactivity (-IR) or dopaminergic (i.e., TH- but not D[beta]H-IR). Our aims were to investigate whether a subpopulation of neurons in the A2 area was dopaminergic and, if so, to investigate the effects of dopamine (DA) on the membrane of gastricprojecting agal motoneurons. We observed that although the majority of A2 neurons were both TH- and D[beta]H-IR, a small percentage of nucleus tractus solitarius neurons were TH-IR only, suggesting that DA itself may play role in these circuits. Whole cell recordings from thin brain stem slices showed that 71% of identified gastric-projecting motoneurons responded to DA (1-300 [micro]-M) with either an excitation (28%) or an inhibition (43%) of the membrane; the remaining 29% of the neurons were unresponsive. The DA-induced depolarization was mimicked by SK 38393 and prevented by pretreatment with SCH 23390. Conversely, the DA-induced inhibition was mimicked by bromoergocryptine and prevented by pretreatment with L741626. When tested on the same neuron, the effects of DA and NE were not always similar. In fact, in neurons in which DA induced a membrane depolarization, 77% were inhibited by NE, whereas 75% of neurons unresponsive to DA were inhibited by NE. Our data suggest that DA modulates the membrane properties of gastric-projecting motoneurons via D1- and D2-like receptors, and DA may play different roles than norepinephrine in brain stem vagal circuits. brain stem; electrophysiology; gastric

Byline: T. Kh. Gainutdinova (1), V. V. Andrianov (1), R. R. Nazyrova (1), Kh. L. Gainutdinov (1) Author Affiliation: (1) Russian Academy of Sciences, Kazan' Physico-Technical Institute, Kazan Article History: Registration Date: 08/10/2004