RESUMO
The basic rhythmic activity that underlies stepping is generated by a neural network, situated in the spinal cord, known as the locomotor central pattern generator (CPG). While a series of lesion experiments have demonstrated that the mammalian locomotor CPG is distributed throughout the ventral portion of the caudal spinal cord, the specific transverse distribution of this neural network is unclear. Here we evoke fictive locomotor activity of various frequencies in upright spinal cords prepared from male and female neonatal mice. This preparation enables us to use an imaging approach to identify locomotor-related cells across the transverse plane of the spinal cord. Results indicate that there is a clear shift in the recruitment of cells toward the ventromedial, and away from the ventrolateral, spinal cord as the frequency of fictive locomotion increases. Surprisingly, the analysis of multiple frequencies of fictive locomotion in the same spinal cord indicates that few neurons are involved in locomotor outputs across multiple speeds. Collectively, these experiments allow us to map the transverse distribution of the locomotor CPG and highlight the pattern of dynamic recruitment that occurs within this neural circuit as the frequency is altered. Our findings are consistent with data indicating that there is a speed-dependent recruitment of interneuronal populations during locomotion and suggest that the locomotor CPG is not a static network, but rather the specific cells recruited vary extensively based on demand.SIGNIFICANCE STATEMENT In this article, we use an imaging approach to identify all those cells that are rhythmically active at the same frequency as fictive locomotion recorded from the ventral roots of the isolated spinal cord. These experiments allow us to map the distribution of locomotor-related cells across the transverse plane of the spinal cord and identify the recruitment pattern of these cells as the frequency of locomotor outputs is altered. Our results indicate that there are drastic changes in the specific neurons activated at different frequencies and provide support for the concept that the locomotor central pattern generator is a modular network with speed-dependent recruitment of interneuronal components.
Assuntos
Geradores de Padrão Central/fisiologia , Locomoção/fisiologia , Neurônios/fisiologia , Medula Espinal/fisiologia , Animais , Animais Recém-Nascidos , Feminino , Masculino , Camundongos , Neurônios Motores/fisiologia , Técnicas de Patch-ClampRESUMO
In order for locomotion to occur, a complex pattern of muscle activation is required. For more than a century, it has been known that the timing and pattern of stepping movements in mammals are generated by neural networks known as central pattern generators (CPGs), which comprise multiple interneuron cell types located entirely within the spinal cord. A genetic approach has recently been successful in identifying several populations of spinal neurons that make up this neural network, as well as the specific role they play during stepping. In spite of this progress, the identity of the neurons responsible for generating the locomotor rhythm and the manner in which they are interconnected have yet to be deciphered. In this review, we summarize key features considered to be expressed by locomotor rhythm-generating neurons and describe the different genetically defined classes of interneurons which have been proposed to be involved.
Assuntos
Interneurônios/fisiologia , Locomoção/fisiologia , Músculo Esquelético/inervação , Rede Nervosa/fisiologia , Medula Espinal/fisiologia , Animais , Humanos , Músculo Esquelético/fisiologia , Medula Espinal/citologiaRESUMO
The basic pattern of activity underlying stepping in mammals is generated by a neural network located in the caudal spinal cord. Within this network, the specific circuitry coordinating left-right alternation has been shown to involve several groups of molecularly defined interneurons. Here we characterize a population of spinal neurons that express the Wilms' tumor 1 (WT1) gene and investigate their role during locomotor activity in mice of both sexes. We demonstrate that WT1-expressing cells are located in the ventromedial region of the spinal cord of mice and are also present in the human spinal cord. In the mouse, these cells are inhibitory, project axons to the contralateral spinal cord, terminate in close proximity to other commissural interneuron subtypes, and are essential for appropriate left-right alternation during locomotion. In addition to identifying WT1-expressing interneurons as a key component of the locomotor circuitry, this study provides insight into the manner in which several populations of molecularly defined interneurons are interconnected to generate coordinated motor activity on either side of the body during stepping.SIGNIFICANCE STATEMENT In this study, we characterize WT1-expressing spinal interneurons in mice and demonstrate that they are commissurally projecting and inhibitory. Silencing of this neuronal population during a locomotor task results in a complete breakdown of left-right alternation, whereas flexor-extensor alternation was not significantly affected. Axons of WT1 neurons are shown to terminate nearby commissural interneurons, which coordinate motoneuron activity during locomotion, and presumably regulate their activity. Finally, the WT1 gene is shown to be present in the spinal cord of humans, raising the possibility of functional homology between these species. This study not only identifies a key component of the locomotor circuitry but also begins to unravel the connectivity among the growing number of molecularly defined interneurons that comprise this neural network.
Assuntos
Geradores de Padrão Central/citologia , Interneurônios Comissurais/citologia , Locomoção/fisiologia , Proteínas Repressoras/metabolismo , Medula Espinal/citologia , Animais , Geradores de Padrão Central/fisiologia , Interneurônios Comissurais/fisiologia , Feminino , Masculino , Camundongos , Vias Neurais/citologia , Vias Neurais/fisiologia , Medula Espinal/fisiologia , Proteínas WT1RESUMO
Locomotion is common to all animals and is essential for survival. Neural circuits located in the spinal cord have been shown to be necessary and sufficient for the generation and control of the basic locomotor rhythm by activating muscles on either side of the body in a specific sequence. Activity in these neural circuits determines the speed, gait pattern, and direction of movement, so the specific locomotor pattern generated relies on the diversity of the neurons within spinal locomotor circuits. Here, we review findings demonstrating that developmental genetics can be used to identify populations of neurons that comprise these circuits and focus on recent work indicating that many of these populations can be further subdivided into distinct subtypes, with each likely to play complementary functions during locomotion. Finally, we discuss data describing the manner in which these populations interact with each other to produce efficient, task-dependent locomotion.
Assuntos
Interneurônios/fisiologia , Locomoção/fisiologia , Rede Nervosa/anatomia & histologia , Rede Nervosa/fisiologia , Medula Espinal/citologia , Medula Espinal/fisiologia , Animais , Humanos , Neurônios Motores/fisiologia , Rede Nervosa/diagnóstico por imagem , Rede Nervosa/crescimento & desenvolvimento , Medula Espinal/diagnóstico por imagem , Medula Espinal/crescimento & desenvolvimentoRESUMO
Since the early 1900's it has been known that a neural network, situated entirely within the spinal cord, is capable of generating the movements required for coordinated locomotion in limbed vertebrates. Due the number of interneurons in the spinal cord, and the extent to which neurons with the same function are intermingled with others that have divergent functions, the components of this neural circuit (now referred to as the locomotor central pattern generator-CPG) have long proven to be difficult to identify. Over the past 20 years a molecular approach has been incorporated to study the locomotor CPG. This approach has resulted in new information regarding the identity of its component interneurons, and their specific role during locomotor activity. In this mini review the role of the inhibitory interneuronal populations that have been shown to be involved in locomotor activity are described, and their specific role in securing left-right, and flexor extensor alternation is outlined. Understanding how these interneuronal populations are activated, modulated, and interact with one another will help us understand how locomotor behavior is produced. In addition, a deeper understanding of the structure and mechanism of function of the locomotor CPG has the potential to assist those developing strategies aimed at enhancing recovery of motor function in spinal cord injured patients.
Assuntos
Interneurônios , Locomoção , Animais , Humanos , Locomoção/fisiologia , Interneurônios/fisiologia , Medula Espinal/fisiologia , NeurôniosRESUMO
Locomotor movements in mammals are generated by neural networks, situated in the spinal cord, known as central pattern generators (CPGs). Recently, significant strides have been made in the genetic identification of interneuronal components of the locomotor CPG and their specific function. Despite this progress, a population of interneurons that is required for locomotor rhythmogenesis has yet to be identified, and it has been suggested that subsets of interneurons belonging to several genetically-defined populations may be involved. In this study, rather than hunt for rhythmogenic neurons, we take a different approach and attempt to identify the specific region of the spinal cord in which they are located. Focal application of 5-hydroxytryptamine creatine sulfate complex (5-HT) and N-methyl-D-aspartate (NMDA) to the central canal of the rostral lumbar segments of newborn male and female mouse spinal cords quickly generates a robust pattern of fictive locomotion, while inhibition or ablation of neurons in this region disrupts the locomotor rhythm in both rostral and caudal lumbar segments. When applied to the central canal at caudal lumbar levels a higher volume of 5-HT and NMDA are required to elicit fictive locomotion, while inhibition of neurons surrounding the central canal at caudal levels again interrupts rhythmic activity at local segmental levels with minimal effects rostrally. The results of this study indicate that interneurons in the most medial laminae of the neonatal mouse spinal cord are both necessary and sufficient for the generation of locomotor activity, and suggests that this is the region where the rhythm generating core of the locomotor CPG resides.
Assuntos
Geradores de Padrão Central , N-Metilaspartato , Animais , Camundongos , Feminino , Masculino , Animais Recém-Nascidos , N-Metilaspartato/farmacologia , Serotonina/farmacologia , Medula Espinal , Mamíferos , Locomoção , Interneurônios/fisiologiaRESUMO
Antisense oligonucleotide-mediated (AO-mediated) therapy is a promising strategy to treat several neurological diseases, including spinal muscular atrophy (SMA). However, limited delivery to the CNS with AOs administered intravenously or subcutaneously is a major challenge. Here, we demonstrate a single subcutaneous administration of cell-penetrating peptide DG9 conjugated to an AO called phosphorodiamidate morpholino oligomer (PMO) reached the CNS and significantly prolonged the median survival compared with unconjugated PMO and R6G-PMO in a severe SMA mouse model. Treated mice exhibited substantially higher expression of full-length survival of motor neuron 2 in both the CNS and systemic tissues compared with nontreated and unmodified AO-treated mice. The treatment ameliorated the atrophic musculature and improved breathing function accompanied by improved muscle strength and innervation at the neuromuscular junction with no signs of apparent toxicity. We also demonstrated DG9-conjugated PMO localized in nuclei in the spinal cord and brain after subcutaneous injections. Our data identify DG9 peptide conjugation as a powerful way to improve the efficacy of AO-mediated splice modulation. Finally, DG9-PMO is a promising therapeutic option to treat SMA and other neurological diseases, overcoming the necessity for intrathecal injections and treating body-wide tissues without apparent toxicity.
Assuntos
Atrofia Muscular Espinal , Splicing de RNA , Camundongos , Animais , Morfolinos/genética , Atrofia Muscular Espinal/genética , Oligonucleotídeos Antissenso/farmacologia , FenótipoRESUMO
Our understanding of the neural control of locomotion has been greatly enhanced by the ability to identify and manipulate genetically defined populations of interneurons that comprise the locomotor central pattern generator (CPG). To date, the dI6 interneurons are one of the few populations that settle in the ventral region of the postnatal spinal cord that have not been investigated. In the present study, we utilized a novel transgenic mouse line to electrophysiologically characterize dI6 interneurons located close to the central canal and study their function during fictive locomotion. The majority of dI6 cells investigated were found to be rhythmically active during fictive locomotion and could be divided into two electrophysiologically distinct populations of interneurons. The first population fired rhythmic trains of action potentials that were loosely coupled to ventral root output and contained several intrinsic membrane properties of rhythm-generating neurons, raising the possibility that these cells may be involved in the generation of rhythmic activity in the locomotor CPG. The second population fired rhythmic trains of action potentials that were tightly coupled to ventral root output and lacked intrinsic oscillatory mechanisms, indicating that these neurons may be driven by a rhythm-generating network. Together these results indicate that dI6 neurons comprise an important component of the locomotor CPG that participate in multiple facets of motor behavior.
Assuntos
Interneurônios/fisiologia , Medula Espinal/fisiologia , Potenciais de Ação/fisiologia , Animais , Animais Recém-Nascidos , Feminino , Proteínas de Homeodomínio/genética , Locomoção/fisiologia , Masculino , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Transgênicos , Técnicas de Patch-Clamp/métodos , Periodicidade , Proteínas/genética , RNA não Traduzido , Medula Espinal/citologia , Raízes Nervosas Espinhais/fisiologiaRESUMO
The neuronal networks that generate vertebrate movements such as walking and swimming are embedded in the spinal cord. These networks, which are referred to as central pattern generators (CPGs), are ideal systems for determining how ensembles of neurons generate simple behavioural outputs. In spite of efforts to address the organization of the locomotor CPG in walking animals, little is known about the identity and function of the spinal interneuron cell types that contribute to these locomotor networks. Here we use four complementary genetic approaches to directly address the function of mouse V1 neurons, a class of local circuit inhibitory interneurons that selectively express the transcription factor Engrailed1. Our results show that V1 neurons shape motor outputs during locomotion and are required for generating 'fast' motor bursting. These findings outline an important role for inhibition in regulating the frequency of the locomotor CPG rhythm, and also suggest that V1 neurons may have an evolutionarily conserved role in controlling the speed of vertebrate locomotor movements.
Assuntos
Locomoção/fisiologia , Neurônios Motores/fisiologia , Medula Espinal/citologia , Medula Espinal/fisiologia , Potenciais de Ação , Animais , Proteínas do Olho/genética , Deleção de Genes , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Interneurônios/fisiologia , Locomoção/genética , Camundongos , Fator de Transcrição PAX6 , Fatores de Transcrição Box Pareados/deficiência , Fatores de Transcrição Box Pareados/genética , Receptores de Neuropeptídeos/metabolismo , Proteínas Repressoras/genética , Fatores de Tempo , Transgenes/genética , Caminhada/fisiologiaRESUMO
In the past two decades we have learned an enormous amount of information regarding the identity of functional components of the neural circuitry responsible for generating locomotor activity in mammals. Molecular techniques, combined with classic electrophysiological and anatomical approaches, have resulted in the identification of a handful of classes of genetically defined interneuronal populations, and a delineation of the specific function of many of these during stepping. What lags behind at this point is a clear picture of the synaptic connectivity of each population, this information is key if we are to understand how the interneuronal components that are responsible for locomotor activity work together to form a functional circuit. In this mini review I will summarize what is, and what is not, known regarding the synaptic connectivity of each genetically defined interneuronal population that is involved in locomotion.
Assuntos
Geradores de Padrão Central , Animais , Geradores de Padrão Central/fisiologia , Medula Espinal/fisiologia , Locomoção/fisiologia , Interneurônios/fisiologia , Fenômenos Eletrofisiológicos , MamíferosRESUMO
Calnexin is a molecular chaperone and a component of the quality control of the secretory pathway. We have generated calnexin gene-deficient mice (cnx(-/-)) and showed that calnexin deficiency leads to myelinopathy. Calnexin-deficient mice were viable with no discernible effects on other systems, including immune function, and instead they demonstrated dysmyelination as documented by reduced conductive velocity of nerve fibers and electron microscopy analysis of sciatic nerve and spinal cord. Myelin of the peripheral and central nervous systems of cnx(-/-) mice was disorganized and decompacted. There were no abnormalities in neuronal growth, no loss of neuronal fibers, and no change in fictive locomotor pattern in the absence of calnexin. This work reveals a previously unrecognized and important function of calnexin in myelination and provides new insights into the mechanisms responsible for myelin diseases.
Assuntos
Calnexina/genética , Calnexina/fisiologia , Doenças Desmielinizantes/metabolismo , Bainha de Mielina/metabolismo , Animais , Animais Recém-Nascidos , Calnexina/metabolismo , Membrana Celular/metabolismo , Eletrofisiologia/métodos , Retículo Endoplasmático/metabolismo , Feminino , Genótipo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Dobramento de Proteína , Nervo Isquiático/metabolismo , Nervo Isquiático/ultraestrutura , Medula Espinal/metabolismo , Medula Espinal/ultraestruturaRESUMO
Genetic strategies for perturbing activity of selected neurons hold great promise for understanding circuitry and behavior. Several such strategies exist, but there has been no direct demonstration of reversible inactivation of mammalian neurons in vivo. We previously reported quickly reversible inactivation of neurons in vitro using expression of the Drosophila allatostatin receptor (AlstR). Here, adeno-associated viral vectors are used to express AlstR in vivo in cortical and thalamic neurons of rats, ferrets, and monkeys. Application of the receptor's ligand, allatostatin (AL), leads to a dramatic reduction in neural activity, including responses of visual neurons to optimized visual stimuli. Additionally, AL eliminates activity in spinal cords of transgenic mice conditionally expressing AlstR. This reduction occurs selectively in AlstR-expressing neurons. Inactivation can be reversed within minutes upon washout of the ligand and is repeatable, demonstrating that the AlstR/AL system is effective for selective, quick, and reversible silencing of mammalian neurons in vivo.
Assuntos
Proteínas de Drosophila/fisiologia , Inibição Neural/fisiologia , Neurônios/metabolismo , Receptores Acoplados a Proteínas G/fisiologia , Receptores de Neuropeptídeos/fisiologia , Potenciais de Ação/fisiologia , Animais , Córtex Cerebral/metabolismo , Proteínas de Drosophila/biossíntese , Proteínas de Drosophila/genética , Feminino , Furões , Macaca mulatta , Masculino , Camundongos , Camundongos Transgênicos , Neuropeptídeos/metabolismo , Ratos , Receptores Acoplados a Proteínas G/biossíntese , Receptores Acoplados a Proteínas G/genética , Receptores de Neuropeptídeos/biossíntese , Receptores de Neuropeptídeos/genéticaRESUMO
Walking in our complex environment requires continual higher order integrated spatiotemporal information. This information is processed in the somatosensory cortex, and it has long been presumed that it influences movement via descending tracts originating from the motor cortex. Here we show that neuronal activity in the primary somatosensory cortex tightly correlates with the onset and speed of locomotion in freely moving mice. Using optogenetics and pharmacogenetics in combination with in vivo and in vitro electrophysiology, we provide evidence for a direct corticospinal pathway from the primary somatosensory cortex that synapses with cervical excitatory neurons and modulates the lumbar locomotor network independently of the motor cortex and other supraspinal locomotor centers. Stimulation of this pathway enhances speed of locomotion, while inhibition decreases locomotor speed and ultimately terminates stepping. Our findings reveal a novel pathway for neural control of movement whereby the somatosensory cortex directly influences motor behavior, possibly in response to environmental cues.
Assuntos
Locomoção/fisiologia , Córtex Somatossensorial/fisiologia , Animais , Camundongos , Camundongos Endogâmicos C57BL , Movimento/fisiologia , Células Piramidais/fisiologiaRESUMO
Rhythmic limb movements are controlled by pattern-generating neurons within the ventral spinal cord, but little is known about how these locomotor circuits are assembled during development. At early stages of embryogenesis, motor neurons are spontaneously active, releasing acetylcholine that triggers the depolarization of adjacent cells in the spinal cord. To investigate whether acetylcholine-driven activity is required for assembly of the central pattern-generating (CPG) circuit, we studied mice lacking the choline acetyltransferase (ChAT) enzyme. Our studies show that a rhythmically active spinal circuit forms in ChAT mutants, but the duration of each cycle period is elongated, and right-left and flexor-extensor coordination are abnormal. In contrast, blocking acetylcholine receptors after the locomotor network is wired does not affect right-left or flexor-extensor coordination. These findings suggest that the cholinergic neurotransmitter pathway is involved in configuring the CPG during a transient period of development.
Assuntos
Acetilcolina/metabolismo , Colina O-Acetiltransferase/deficiência , Neurônios Motores/metabolismo , Vias Neurais/embriologia , Medula Espinal/embriologia , Potenciais de Ação/fisiologia , Animais , Ataxia/etiologia , Eletrofisiologia , Embrião de Mamíferos , Imuno-Histoquímica , Hibridização In Situ , Camundongos , Camundongos Mutantes , Atividade Motora/fisiologia , Vias Neurais/enzimologia , Técnicas de Cultura de ÓrgãosRESUMO
The basic rhythmic activity characteristic of locomotion in mammals is generated by a neural network, located in the spinal cord, known as the locomotor central pattern generator (CPG). Although a great deal of effort has gone into the study of this neural circuit over the past century, identification and characterization of its component interneurons has proven to be challenging, largely due to their location and distribution. Recent work incorporating a molecular approach has provided a great deal of insight into the genetic identity of interneurons that make up this neural circuit, as well as the specific roles that they play during stepping. Despite this progress we still know relatively little regarding the manner in which these neuronal populations are interconnected. In this article we review the interneuronal populations shown to be involved in locomotor activity, briefly summarize their specific function, and focus on experimental work that provides insight into their synaptic connectivity. Finally, we discuss how recently developed viral approaches can potentially be incorporated to provide further insight into the network structure of this neural circuit.
RESUMO
BACKGROUND: The basic rhythmicity underlying stepping in mammals is generated by a neural network, situated in the spinal cord, known as the locomotor central pattern generator (CPG). While a molecular approach has provided information regarding neuronal populations that participate in locomotor activity and their specific function, the distributed nature of the locomotor CPG has made it difficult to identify and characterize the specific neurons belonging to each population that are rhythmically-active during stepping. NEW METHOD: We describe a preparation in which we isolate the spinal cord from a neonatal mouse, section it at a lumbar segment, situate it in an upright orientation under the objective lens of a 2- photon microscope, and evoke fictive locomotion. RESULTS: This preparation allows us to image rhythmic Ca2+ oscillations in spinal neurons, and visually identify those that are involved in fictive locomotor activity. We can then characterize unique features of these neurons. COMPARISON WITH EXISTING METHODS: This builds on existing fictive locomotor preparations and is the first which allows for the visual identification of locomotor related neurons spanning the transverse plane of the spinal cord, facilitating their electrophysiological and anatomical characterization CONCLUSIONS: This approach promises to provide new information regarding the distribution of the locomotor CPG in the transverse plane, the characteristics of its component interneurons, as well as the cellular mechanisms and network properties which underlie rhythm generation. By altering the location of Ca2+ indicator application it can also be used to identify and characterize neurons involved in other facets of sensorimotor processing.
Assuntos
Comportamento Animal , Geradores de Padrão Central/citologia , Técnicas de Preparação Histocitológica/métodos , Locomoção , Medula Espinal/citologia , Animais , Animais Recém-Nascidos , Camundongos , Microscopia de Fluorescência por Excitação Multifotônica , Técnicas de Patch-ClampRESUMO
The sequential stepping of left and right limbs is a fundamental motor behavior that underlies walking movements. This relatively simple locomotor behavior is generated by the rhythmic activity of motor neurons under the control of spinal neural networks known as central pattern generators (CPGs) that comprise multiple interneuron cell types. Little, however, is known about the identity and contribution of defined interneuronal populations to mammalian locomotor behaviors. We show a discrete subset of commissural spinal interneurons, whose fate is controlled by the activity of the homeobox gene Dbx1, has a critical role in controlling the left-right alternation of motor neurons innervating hindlimb muscles. Dbx1 mutant mice lacking these ventral interneurons exhibit an increased incidence of cobursting between left and right flexor/extensor motor neurons during drug-induced locomotion. Together, these findings identify Dbx1-dependent interneurons as key components of the spinal locomotor circuits that control stepping movements in mammals.
Assuntos
Interneurônios/metabolismo , Atividade Motora/genética , Medula Espinal/metabolismo , Caminhada/fisiologia , Animais , Genes fos/genética , Genes fos/fisiologia , Proteínas de Homeodomínio/biossíntese , Proteínas de Homeodomínio/genética , Técnicas In Vitro , Camundongos , Camundongos Mutantes , Atividade Motora/fisiologia , Desempenho Psicomotor/fisiologiaRESUMO
The locomotor central pattern generator is a neural network located in the ventral aspect of the caudal spinal cord that underlies stepping in mammals. While many genetically defined interneurons that are thought to comprise this neural network have been identified and characterized, the dI6 cells- which express the transcription factors WT1 and/or DMRT3- are one population that settle in this region, are active during locomotion, whose function is poorly understood. These cells were originally hypothesized to be commissural premotor interneurons, however evidence in support of this is sparse. Here we characterize this population of cells using the TgDbx1Cre;R26EFP;Dbx1LacZ transgenic mouse line, which has been shown to be an effective marker of dI6 interneurons. We show dI6 cells to be abundant in laminae VII and VIII along the entire spinal cord and provide evidence that subtypes outside the WT1/DMRT3 expressing dI6 cells may exist. Retrograde tracing experiments indicate that the majority of dI6 cells project descending axons, and some make monosynaptic or disynaptic contacts onto motoneurons on either side of the spinal cord. Analysis of their activity during non-resetting deletions, which occur during bouts of fictive locomotion, suggests that these cells are involved in both locomotor rhythm generation and pattern formation. This study provides a thorough characterization of the dI6 cells labeled in the TgDbx1Cre;R26EFP;Dbx1LacZ transgenic mouse, and supports previous work suggesting that these cells play multiple roles during locomotor activity.
Assuntos
Interneurônios/citologia , Interneurônios/fisiologia , Medula Espinal/citologia , Medula Espinal/fisiologia , Animais , Animais Recém-Nascidos , Geradores de Padrão Central/citologia , Geradores de Padrão Central/fisiologia , Lateralidade Funcional , Imuno-Histoquímica , Locomoção/fisiologia , Camundongos Transgênicos , Neurônios Motores/citologia , Neurônios Motores/fisiologia , Técnicas de Rastreamento Neuroanatômico , Técnicas de Patch-Clamp , Medula Espinal/crescimento & desenvolvimento , Técnicas de Cultura de TecidosRESUMO
The V0 interneuronal population is derived from Dbx1 expressing progenitors. Initial studies on these interneurons in the mouse spinal cord demonstrated that they project commissural axons and are involved in coordinating left-right alternation during locomotion. Subsequent work has indicated that the V0 population can be divided into genetically distinct ventral (V0V) and dorsal (V0D) subpopulations, and experimental evidence suggests that each is responsible for left-right alternation at different locomotor speeds. In this study, we perform a series of experiments to probe the location and connectivity of these subpopulations in neonatal mice and demonstrate that they are more diverse than previously predicted. While the distribution of either subpopulation remains consistent along the extent of the lumbar spinal cord, a cluster of V0D cells lateral to the central canal receive substantial input from primary afferents. Retrograde tracing and activity dependent labeling experiments demonstrate that a group of V0 interneurons located in this same region preferentially project axons towards contralateral motoneurons via an oligosynaptic pathway, and are active during fictive locomotion. Our results suggest that this subset of V0 interneurons may be primarily responsible for coordination of left-right alternation during locomotion. Furthermore these experiments indicate that while genetic identity is one determinant of the function of a neuron during locomotion, the specific position in which the cell is located may also play a key role.
Assuntos
Interneurônios/fisiologia , Locomoção/fisiologia , Medula Espinal/fisiologia , Vias Aferentes/citologia , Vias Aferentes/crescimento & desenvolvimento , Vias Aferentes/fisiologia , Animais , Animais Recém-Nascidos , Lateralidade Funcional/fisiologia , Glicina/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Imuno-Histoquímica , Interneurônios/citologia , Vértebras Lombares , Camundongos Transgênicos , Neurônios Motores/citologia , Neurônios Motores/fisiologia , Células-Tronco Neurais/fisiologia , Técnicas de Rastreamento Neuroanatômico , Proteínas Proto-Oncogênicas c-fos/metabolismo , Serotonina/metabolismo , Medula Espinal/citologia , Medula Espinal/crescimento & desenvolvimentoRESUMO
V1 and V2b interneurons (INs) are essential for the production of an alternating flexor-extensor motor output. Using a tripartite genetic system to selectively ablate either V1 or V2b INs in the caudal spinal cord and assess their specific functions in awake behaving animals, we find that V1 and V2b INs function in an opposing manner to control flexor-extensor-driven movements. Ablation of V1 INs results in limb hyperflexion, suggesting that V1 IN-derived inhibition is needed for proper extension movements of the limb. The loss of V2b INs results in hindlimb hyperextension and a delay in the transition from stance phase to swing phase, demonstrating V2b INs are required for the timely initiation and execution of limb flexion movements. Our findings also reveal a bias in the innervation of flexor- and extensor-related motor neurons by V1 and V2b INs that likely contributes to their differential actions on flexion-extension movements.