Motor Neuron Subtypes Research Articles

Patterns of Spinal Sensory-Motor Connectivity Prescribed by a Dorsoventral Positional Template

Sensory-motor circuits in the spinal cord are constructed with a fine specificity that coordinates motor behavior, but the mechanisms that direct sensory connections with their motor neuron partners remain unclear. The dorsoventral settling position of motor pools in the spinal cord is known to match the distal-to-proximal position of their muscle targets in the limb, but the significance of invariant motor neuron positioning is unknown. An analysis of sensory-motor connectivity patterns in FoxP1 mutant mice, where motor neuron position has been scrambled, shows that the final pattern of sensory-motor connections is initiated by the projection of sensory axons to discrete dorsoventral domains of the spinal cord without regard for motor neuron subtype or, indeed, the presence of motor neurons. By implication, the clustering and dorsoventral settling position of motor neuron pools serve as a determinant of the pattern of sensory input specificity and thus motor coordination.

GDE2 Regulates Subtype-Specific Motor Neuron Generation through Inhibition of Notch Signaling

The specification of spinal interneuron and motor neuron identities initiates within progenitor cells, while motor neuron subtype diversification is regulated by hierarchical transcriptional programs implemented postmitotically. Here we find that mice lacking GDE2, a six-transmembrane protein that triggers motor neuron generation, exhibit selective losses ofdistinct motor neuron subtypes, specifically in defined subsets of limb-innervating motor pools that correlate with the loss of force-generating alpha motor neurons. Mechanistically, GDE2 is expressed by postmitotic motor neurons but utilizes extracellular glycerophosphodiester phosphodiesterase activity to induce motor neuron generation by inhibiting Notch signaling in neighboring motor neuron progenitors. Thus, neuronal GDE2 controls motor neuron subtype diversity through a non-cell-autonomous feedback mechanism that directly regulates progenitor cell differentiation, implying that subtype specification initiates within motor neuron progenitor populations prior to their differentiation into postmitotic motor neurons.

Motoneuron subtypes show specificity in glycine receptor channel abnormalities in a transgenic mouse model of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by selective loss of motoneurons. Recently we studied glycine receptors (GlyRs) in motoneurons in an ALS mouse model expressing a mutant form of human superoxide dismutase-1 with a Gly93→Ala substitution (G93A-SOD1). Living motoneurons in dissociated spinal cord cultures were identified by using transgenic mice expressing eGFP driven by the Hb9 promoter. We showed that GlyR-mediated currents were reduced in large-sized (diameter > 28 μm) Hb9-eGFP+ motoneurons from G93A-SOD1 embryonic mice. Here we analyze GlyR currents in a morphologically distinct subgroup of medium-sized (diameter 10-28 μm) Hb9-eGFP+ motoneurons, presumably gamma or slow-type alpha motoneurons. We find that glycine-induced current densities were not altered in medium-sized G93A-SOD1 motoneurons. No significant differences in glycinergic mIPSCs were observed between G93A-SOD1 and control medium-sized motoneurons. These results indicate that GlyR deficiency early in the disease process of ALS is specific for large alpha motoneurons.

Overlapping Roles and Collective Requirement for the Coreceptors GAS1, CDO, and BOC in SHH Pathway Function

Secreted Hedgehog (HH) ligands signal through the canonical receptor Patched (PTCH1). However, recent studies implicate three additional HH-binding, cell-surface proteins, GAS1, CDO, and BOC, as putative coreceptors for HH ligands. A central question is to what degree these coreceptors function similarly and what their collective requirement in HH signal transduction is. Here we provide evidence that GAS1, CDO, and BOC play overlapping and essential roles during HH-mediated ventral neural patterning of the mammalian neural tube. Specifically, we demonstrate two important roles for these molecules: an early role in cell fate specification of multiple neural progenitors and a later role in motor neuron progenitor maintenance. Most strikingly, genetic loss-of-function experiments indicate an obligatory requirement for GAS1, CDO, and BOC in HH pathway activity in multiple tissues.

Phosphorylation Regulates OLIG2 Cofactor Choice and the Motor Neuron-Oligodendrocyte Fate Switch

SummaryA fundamental feature of central nervous system development is that neurons are generated before glia. In the embryonic spinal cord, for example, a group of neuroepithelial stem cells (NSCs) generates motor neurons (MNs), before switching abruptly to oligodendrocyte precursors (OLPs). We asked how transcription factor OLIG2 participates in this MN-OLP fate switch. We found that Serine 147 in the helix-loop-helix domain of OLIG2 was phosphorylated during MN production and dephosphorylated at the onset of OLP genesis. Mutating Serine 147 to Alanine (S147A) abolished MN production without preventing OLP production in transgenic mice, chicks, or cultured P19 cells. We conclude that S147 phosphorylation, possibly by protein kinase A, is required for MN but not OLP genesis and propose that dephosphorylation triggers the MN-OLP switch. Wild-type OLIG2 forms stable homodimers, whereas mutant (unphosphorylated) OLIG2S147A prefers to form heterodimers with Neurogenin 2 or other bHLH partners, suggesting a molecular basis for the switch.

Retinoid-independent motor neurogenesis from human embryonic stem cells reveals a medial columnar ground state.

A major challenge in neurobiology is to understand mechanisms underlying human neuronal diversification. Motor neurons (MNs) represent a diverse collection of neuronal subtypes, displaying differential vulnerability in different human neurodegenerative diseases. The ability to manipulate cell subtype diversification is critical to establish accurate, clinically relevant in vitro disease models. Retinoid signalling contributes to caudal precursor specification and subsequent MN subtype diversification. Here we investigate the necessity for retinoic acid in motor neurogenesis from human embryonic stem cells. We show that activin/nodal signalling inhibition, followed by sonic hedgehog agonist treatment, is sufficient for MN precursor specification, which occurs even in the presence of retinoid pathway antagonists. Importantly, precursors mature into HB9/ChAT-expressing functional MNs. Furthermore, retinoid-independent motor neurogenesis results in a ground state biased to caudal, medial motor columnar identities from which a greater retinoid-dependent diversity of MNs, including those of lateral motor columns, can be selectively derived in vitro.

MicroRNA miR-9 Modifies Motor Neuron Columns by a Tuning Regulation of FoxP1 Levels in Developing Spinal Cords

The precise organization of motor neuron subtypes in a columnar pattern in developing spinal cords is controlled by cross-interactions of multiple transcription factors and segmental expressions of Hox genes and their accessory proteins. Accurate expression levels and domains of these regulators are essential for organizing spinal motor neuron columns and axonal projections to target muscles. Here, we show that microRNA miR-9 is transiently expressed in a motor neuron subtype and displays overlapping expression with its target gene FoxP1. Overexpression or knockdown of miR-9 alters motor neuron subtypes, switches columnar identities, and changes axonal innervations in developing chick spinal cords. miR-9 modifies spinal columnar organization by specifically regulating FoxP1 protein levels, which in turn determine distinct motor neuron subtypes. Our findings demonstrate that miR-9 is an essential regulator of motor neuron specification and columnar formation. Moreover, the overlapping expression of miR-9 and its target FoxP1 further illuminates the importance of fine-tuning regulation by microRNAs in motor neuron development.

Presenilin-Dependent Receptor Processing Is Required for Axon Guidance

The Alzheimer's disease-linked gene presenilin is required for intramembrane proteolysis of amyloid-β precursor protein, contributing to the pathogenesis of neurodegeneration that is characterized by loss of neuronal connections, but the role of Presenilin in establishing neuronal connections is less clear. Through a forward genetic screen in mice for recessive genes affecting motor neurons, we identified the Columbus allele, which disrupts motor axon projections from the spinal cord. We mapped this mutation to the Presenilin-1 gene. Motor neurons and commissural interneurons in Columbus mutants lacking Presenilin-1 acquire an inappropriate attraction to Netrin produced by the floor plate because of an accumulation of DCC receptor fragments within the membrane that are insensitive to Slit/Robo silencing. Our findings reveal that Presenilin-dependent DCC receptor processing coordinates the interplay between Netrin/DCC and Slit/Robo signaling. Thus, Presenilin is a key neural circuit builder that gates the spatiotemporal pattern of guidance signaling, thereby ensuring neural projections occur with high fidelity.

Stochastic Mechanisms of Cell Fate Specification that Yield Random or Robust Outcomes

Although cell fate specification is tightly controlled to yield highly reproducible results and avoid extreme variation, developmental programs often incorporate stochastic mechanisms to diversify cell types. Stochastic specification phenomena are observed in a wide range of species and an assorted set of developmental contexts. In bacteria, stochastic mechanisms are utilized to generate transient subpopulations capable of surviving adverse environmental conditions. In vertebrate, insect, and worm nervous systems, stochastic fate choices are used to increase the repertoire of sensory and motor neuron subtypes. Random fate choices are also integrated into developmental programs controlling organogenesis. Although stochastic decisions can be maintained to produce a mosaic of fates within a population of cells, they can also be compensated for or directed to yield robust and reproducible outcomes.

Functional Diversity of ESC-Derived Motor Neuron Subtypes Revealed through Intraspinal Transplantation

Cultured ESCs can form different classes of neurons, but whether these neurons can acquire specialized subtype features typical of neurons in vivo remains unclear. We show here that mouse ESCs can be directed to form highly specific motor neuron subtypes in the absence of added factors, through a differentiation program that relies on endogenous Wnts, FGFs, and Hh-mimicking the normal program of motor neuron subtype differentiation. Molecular markers that characterize motor neuron subtypes anticipate the functional properties of these neurons in vivo: ESC-derived motor neurons grafted isochronically into chick spinal cord settle in appropriate columnar domains and select axonal trajectories with a fidelity that matches that of their in vivo generated counterparts. ESC-derived motor neurons can therefore be programmed in a predictive manner to acquire molecular and functional properties that characterize one of the many dozens of specialized motor neuron subtypes that exist in vivo.

Developmental regulation of subtype‐specific motor neuron excitability

At early embryonic stages, zebrafish spinal neuron subtypes can be distinguished and accessed for physiological studies. This provides the opportunity to determine electrophysiological properties of different spinal motor neuron subtypes. Such differences have the potential to then regulate, in a subtype-specific manner, activity-dependent developmental events such as axonal outgrowth and pathfinding. The zebrafish spinal cord contains a population of early born neurons. Our recent work has revealed that primary motor neuron (PMN) subtypes in the zebrafish spinal cord differ with respect to electrical properties during early important periods when PMNs extend axons to their specific targets. Here, we review recent findings regarding the development of electrical properties in PMN subtypes. Moreover, we consider the possibility that electrical activity in PMNs may play a cell nonautonomous role and thus influence the development of later developing motor neurons. Further, we discuss findings that support a role for a specific sodium channel isoform, Nav1.6, expressed by specific subtypes of spinal neurons in activity-dependent processes that impact axonal outgrowth and pathfinding.

Identification of novel spinal cholinergic genetic subtypes disclose Chodl and Pitx2 as markers for fast motor neurons and partition cells

Spinal cholinergic neurons are critical for motor function in both the autonomic and somatic nervous systems and are affected in spinal cord injury and in diseases such as amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy. Using two screening approaches and in situ hybridization, we identified 159 genes expressed in typical cholinergic patterns in the spinal cord. These include two general cholinergic neuron markers, one gene exclusively expressed in motor neurons, and nine genes expressed in unknown subtypes of somatic motor neurons. Further, we present evidence that chondrolectin (Chodl) is expressed by fast motor neurons and that estrogen-related receptor beta (ERRbeta) is a candidate marker for slow motor neurons. In addition, we suggest paired-like homeodomain transcription factor 2 (Pitx2) as a marker for cholinergic partition cells.

A Conditioning Lesion Provides Selective Protection in a Rat Model of Amyotrophic Lateral Sclerosis

BackgroundAmyotrophic Lateral Sclerosis (ALS) is neurodegenerative disease characterized by muscle weakness and atrophy due to progressive motoneuron loss. The death of motoneuron is preceded by the failure of neuromuscular junctions (NMJs) and axonal retraction. Thus, to develop an effective ALS therapy you must simultaneously preserve motoneuron somas, motor axons and NMJs. A conditioning lesion has the potential to accomplish this since it has been shown to enhance neuronal survival and recovery from trauma in a variety of contexts.Methodology/Principal FindingsTo test the effects of a conditioning lesion in a model of familial ALS we administered a tibial nerve crush injury to presymptomatic fALSG93A rats. We examined its effects on motor function, motoneuron somas, motor axons, and NMJs. Our experiments revealed a novel paradigm for the conditioning lesion effect. Specifically we found that the motor functional decline in fALSG93A rats that received a conditioning lesion was delayed and less severe. These improvements in motor function corresponded to greater motoneuron survival, reduced motor axonopathy, and enhanced NMJ maintenance at disease end-stage. Furthermore, the increased NMJ maintenance was selective for muscle compartments innervated by the most resilient (slow) motoneuron subtypes, but was absent in muscle compartments innervated by the most vulnerable (fast fatigable) motoneuron subtypes.Conclusions/SignificanceThese findings support the development of strategies aimed at mimicking the conditioning lesion effect to treat ALS as well as underlined the importance of considering the heterogeneity of motoneuron sub-types when evaluating prospective ALS therapeutics.

Dendritic targeting in the leg neuropil of Drosophila: the role of midline signalling molecules in generating a myotopic map.

Author SummaryDuring development the axons of sensory neurons generate highly ordered ”sensory maps„ within the nervous system that represent specific qualities of the environment. Much less is known about the anatomical organization and development of motor systems. Here, we show that the leg motoneurons of Drosophila organize their dendrites within the central nervous system in a way that reflects the position of the muscles they innervate. These motoneurons generate a ‘myotopic map’ by targeting the growth of their dendrites (sites of synaptic input) into discrete territories during development. The precise targeting of dendrites along the mediolateral axis is controlled by the signaling molecules Slit and Netrin, which are secreted by midline cells. These proteins act as global guidance cues and exert their effects via distinct signaling pathways using receptors called Roundabout and Frazzled, respectively. Previous studies have shown that Slit also helps to position the termini of axons (sites of synaptic output), independent of their synaptic partners. We suggest that the coordinated targeting of both input and output elements of a neural system into a common space using shared global guidance cues could be a simple way of establishing the specificity of synaptic connections within neural networks.

Zebrafish Motor Neuron Subtypes Differ Electrically Prior to Axonal Outgrowth

Different muscle targets and transcription factor expression patterns reveal the presence of motor neuron subtypes. However, it is not known whether these subtypes also differ with respect to electrical membrane properties. To address this question, we studied primary motor neurons (PMNs) in the spinal cord of zebrafish embryos. PMN genesis occurs during gastrulation and gives rise to a heterogeneous set of motor neurons that differ with respect to transcription factor expression, muscle targets, and soma location within each spinal cord segment. The unique subtype-specific soma locations and axonal trajectories of two PMNs-MiP (middle) and CaP (caudal)-allowed their identification in situ as early as 17 h postfertilization (hpf), prior to axon genesis. Between 17 and 48 hpf, CaPs and MiPs displayed subtype-specific electrical membrane properties. Voltage-dependent inward and outward currents differed significantly between MiPs and CaPs. Moreover, by 48 hpf, CaPs and MiPs displayed subtype-specific firing behaviors. Our results demonstrate that motor neurons that differ with respect to muscle targets and transcription factor expression acquire subtype-specific electrical membrane properties. Moreover, the differences are evident prior to axon genesis and persist to the latest stage studied, 2 days postfertilization.

Gamma and alpha motor neurons distinguished by expression of transcription factor Err3

Spinal motor neurons are specified to innervate different muscle targets through combinatorial programs of transcription factor expression. Whether transcriptional programs also establish finer aspects of motor neuron subtype identity, notably the prominent functional distinction between alpha and gamma motor neurons, remains unclear. In this study, we identify DNA binding proteins with complementary expression profiles in alpha and gamma motor neurons, providing evidence for molecular distinctions in these two motor neuron subtypes. The transcription factor Err3 is expressed at high levels in gamma but not alpha motor neurons, whereas the neuronal DNA binding protein NeuN marks alpha but not gamma motor neurons. Signals from muscle spindles are needed to support the differentiation of Err3(on)/NeuN(off) presumptive gamma motor neurons, whereas direct proprioceptive sensory input to a motor neuron pool is apparently dispensable. Together, these findings provide evidence that transcriptional programs define functionally distinct motor neuron subpopulations, even within anatomically defined motor pools.

Hox-directed motoneuron subtype development and the role of the Hoxd11 homeodomain

Hox-directed motoneuron subtype development and the role of the Hoxd11 homeodomain

‘Runxs and regulations’ of sensory and motor neuron subtype differentiation: Implications for hematopoietic development

Runt-related (RUNX) transcription factors are evolutionarily conserved regulators of a number of developmental mechanisms. RUNX proteins often control the balance between proliferation and differentiation and alterations of their functions are associated with different types of cancer and other human pathologies. Moreover, RUNX factors control important steps during the developmental acquisition of mature phenotypes. A number of investigations are beginning to shed light on the involvement of RUNX family members in the development of the nervous system. This review summarizes recent progress in the study of the roles of mammalian RUNX proteins during the differentiation of sensory and motor neurons in the peripheral and central nervous system, respectively. The implications of those findings for RUNX-mediated regulation of hematopoietic development will also be discussed.

A role for motoneuron subtype–selective ER stress in disease manifestations of FALS mice

The mechanisms underlying disease manifestations in neurodegeneration remain unclear, but their understanding is critical to devising effective therapies. We carry out a longitudinal analysis in vivo of identified motoneurons selectively vulnerable (VUL) or resistant (RES) to motoneuron disease (amyotrophic lateral sclerosis, ALS) and show that subtype-selective endoplasmic reticulum (ER) stress responses influence disease manifestations. VUL motoneurons were selectively prone to ER stress and showed gradually upregulated ER stress markers from birth on in three mouse models of familial ALS (FALS). 25-30 days before the earliest denervations, ubiquitin signals increased in both VUL and RES motoneurons, but an unfolded protein response coupled with microglial activation was initiated selectively in VUL motoneurons. This transition was followed by selective axonal degeneration and spreading stress. The ER stress-protective agent salubrinal attenuated disease manifestations and delayed progression, whereas chronic enhancement of ER stress promoted disease. Thus, whereas all motoneurons are preferentially affected in ALS, ER stress responses in specific motoneuron subtypes influence the progressive manifestations of weakening and paralysis.

Restricted patterns of Hoxd10 and Hoxd11 set segmental differences in motoneuron subtype complement in the lumbosacral spinal cord

During normal vertebrate development, Hoxd10 and Hoxd11 are expressed by differentiating motoneurons in restricted patterns along the rostrocaudal axis of the lumbosacral (LS) spinal cord. To assess the roles of these genes in the attainment of motoneuron subtypes characteristic of LS subdomains, we examined subtype complement after overexpression of Hoxd10 or Hoxd11 in the embryonic chick LS cord and in a Hoxd10 loss-of-function mouse embryo. Data presented here provide evidence that Hoxd10 defines the position of the lateral motor column (LMC) as a whole and, in rostral LS segments, specifically promotes the development of motoneurons of the lateral subdivision of the lateral motor column (LMCl). In contrast, Hoxd11 appears to impart a caudal and medial LMC (LMCm) identity to some motoneurons and molecular profiles suggestive of a suppression of LMC development in others. We also provide evidence that Hoxd11 suppresses the expression of Hoxd10 and the retinoic acid synthetic enzyme, retinaldehyde dehydrogenase 2 (RALDH2). In a normal chick embryo, Hoxd10 and RALDH2 are expressed throughout the LS region at early stages of motoneuron differentiation but their levels decline in Hoxd11-expressing caudal LS segments that ultimately contain few LMCl motoneurons. We hypothesize that one of the roles played by Hoxd11 is to modulate Hoxd10 and local retinoic acid levels and thus, perhaps define the caudal boundaries of the LMC and its subtype complement.