RESUMEN
BACKGROUND: Cerebral Palsy (CP) is a major cause of motor and cognitive disability in children due to injury to the developing brain. Early intensive sensorimotor rehabilitation has been shown to change brain structure and reduce CP symptoms severity. We combined environmental enrichment (EE) and treadmill training (TT) to observe the effects of a one-week program of sensorimotor stimulation (EETT) in animals exposed to a CP model and explored possible mechanisms involved in the functional recovery. METHODS: Pregnant Wistar rats were injected with Lipopolysaccharide (LPS - 200 µg/kg) intraperitoneally at embryonic days 18 and 19. At P0, pups of both sexes were exposed to 20' anoxia at 37 °C. From P2 to P21, hindlimbs were restricted for 16 h/day during the dark cycle. EETT lasted from P21 to P27. TT - 15 min/day at 7 cm/s. EE - 7 days in enriched cages with sensorimotor stimulus. Functional 3D kinematic gait analysis and locomotion were analyzed. At P28, brains were collected for ex-vivo MRI and histological assessment. Neurotrophins and key proteins involved in CNS function were assessed by western blotting. RESULTS: CP model caused gross and skilled locomotor disruption and altered CNS neurochemistry. EETT reversed locomotor dysfunction with minor effects over gait kinematics. EETT also decreased brain inflammation and glial activation, preserved myelination, upregulated BDNF signaling and modulated the expression of proteins involved in excitatory synaptic function in the brain and spinal cord. CONCLUSIONS: Using this translational approach based on intensive sensorimotor rehabilitation, we highlight pathways engaged in the early developmental processes improving neurological recovery observed in CP.
Asunto(s)
Parálisis Cerebral , Modelos Animales de Enfermedad , Locomoción , Plasticidad Neuronal , Ratas Wistar , Animales , Parálisis Cerebral/rehabilitación , Parálisis Cerebral/fisiopatología , Plasticidad Neuronal/fisiología , Ratas , Femenino , Locomoción/fisiología , Masculino , Encéfalo/metabolismo , Encéfalo/fisiopatología , Embarazo , Recuperación de la Función/fisiología , Encefalitis/metabolismo , Encefalitis/fisiopatología , Encefalitis/rehabilitación , Marcha/fisiología , Condicionamiento Físico Animal/fisiología , Condicionamiento Físico Animal/métodos , Enfermedades Neuroinflamatorias/metabolismo , Enfermedades Neuroinflamatorias/fisiopatologíaRESUMEN
Axon regeneration can be induced across anatomically complete spinal cord injury (SCI), but robust functional restoration has been elusive. Whether restoring neurological functions requires directed regeneration of axons from specific neuronal subpopulations to their natural target regions remains unclear. To address this question, we applied projection-specific and comparative single-nucleus RNA sequencing to identify neuronal subpopulations that restore walking after incomplete SCI. We show that chemoattracting and guiding the transected axons of these neurons to their natural target region led to substantial recovery of walking after complete SCI in mice, whereas regeneration of axons simply across the lesion had no effect. Thus, reestablishing the natural projections of characterized neurons forms an essential part of axon regeneration strategies aimed at restoring lost neurological functions.
Asunto(s)
Axones , Regeneración Nerviosa , Parálisis , Recuperación de la Función , Traumatismos de la Médula Espinal , Caminata , Animales , Ratones , Axones/fisiología , Regeneración Nerviosa/genética , Regeneración Nerviosa/fisiología , Neuronas/fisiología , Parálisis/fisiopatología , Traumatismos de la Médula Espinal/fisiopatología , ConectomaRESUMEN
A spinal cord injury interrupts pathways from the brain and brainstem that project to the lumbar spinal cord, leading to paralysis. Here we show that spatiotemporal epidural electrical stimulation (EES) of the lumbar spinal cord1-3 applied during neurorehabilitation4,5 (EESREHAB) restored walking in nine individuals with chronic spinal cord injury. This recovery involved a reduction in neuronal activity in the lumbar spinal cord of humans during walking. We hypothesized that this unexpected reduction reflects activity-dependent selection of specific neuronal subpopulations that become essential for a patient to walk after spinal cord injury. To identify these putative neurons, we modelled the technological and therapeutic features underlying EESREHAB in mice. We applied single-nucleus RNA sequencing6-9 and spatial transcriptomics10,11 to the spinal cords of these mice to chart a spatially resolved molecular atlas of recovery from paralysis. We then employed cell type12,13 and spatial prioritization to identify the neurons involved in the recovery of walking. A single population of excitatory interneurons nested within intermediate laminae emerged. Although these neurons are not required for walking before spinal cord injury, we demonstrate that they are essential for the recovery of walking with EES following spinal cord injury. Augmenting the activity of these neurons phenocopied the recovery of walking enabled by EESREHAB, whereas ablating them prevented the recovery of walking that occurs spontaneously after moderate spinal cord injury. We thus identified a recovery-organizing neuronal subpopulation that is necessary and sufficient to regain walking after paralysis. Moreover, our methodology establishes a framework for using molecular cartography to identify the neurons that produce complex behaviours.
Asunto(s)
Neuronas , Parálisis , Traumatismos de la Médula Espinal , Médula Espinal , Caminata , Animales , Humanos , Ratones , Neuronas/fisiología , Parálisis/genética , Parálisis/fisiopatología , Parálisis/terapia , Médula Espinal/citología , Médula Espinal/fisiología , Médula Espinal/fisiopatología , Traumatismos de la Médula Espinal/genética , Traumatismos de la Médula Espinal/fisiopatología , Traumatismos de la Médula Espinal/terapia , Caminata/fisiología , Estimulación Eléctrica , Región Lumbosacra/inervación , Rehabilitación Neurológica , Análisis de Secuencia de ARN , Perfilación de la Expresión GénicaRESUMEN
A spinal cord injury usually spares some components of the locomotor circuitry. Deep brain stimulation (DBS) of the midbrain locomotor region and epidural electrical stimulation of the lumbar spinal cord (EES) are being used to tap into this spared circuitry to enable locomotion in humans with spinal cord injury. While appealing, the potential synergy between DBS and EES remains unknown. Here, we report the synergistic facilitation of locomotion when DBS is combined with EES in a rat model of severe contusion spinal cord injury leading to leg paralysis. However, this synergy requires high amplitudes of DBS, which triggers forced locomotion associated with stress responses. To suppress these undesired responses, we link DBS to the intention to walk, decoded from cortical activity using a robust, rapidly calibrated unsupervised learning algorithm. This contingency amplifies the supraspinal descending command while empowering the rats into volitional walking. However, the resulting improvements may not outweigh the complex technological framework necessary to establish viable therapeutic conditions.
Asunto(s)
Estimulación Encefálica Profunda/métodos , Modelos Animales de Enfermedad , Vértebras Lumbares/fisiopatología , Corteza Motora/fisiopatología , Traumatismos de la Médula Espinal/terapia , Médula Espinal/fisiopatología , Caminata/fisiología , Animales , Estimulación Eléctrica/métodos , Femenino , Humanos , Locomoción/fisiología , Mesencéfalo/fisiopatología , Neuronas/fisiología , Ratas Endogámicas Lew , Traumatismos de la Médula Espinal/fisiopatologíaRESUMEN
Severe spinal cord contusions interrupt nearly all brain projections to lumbar circuits producing leg movement. Failure of these projections to reorganize leads to permanent paralysis. Here we modeled these injuries in rodents. A severe contusion abolished all motor cortex projections below injury. However, the motor cortex immediately regained adaptive control over the paralyzed legs during electrochemical neuromodulation of lumbar circuits. Glutamatergic reticulospinal neurons with residual projections below the injury relayed the cortical command downstream. Gravity-assisted rehabilitation enabled by the neuromodulation therapy reinforced these reticulospinal projections, rerouting cortical information through this pathway. This circuit reorganization mediated a motor cortex-dependent recovery of natural walking and swimming without requiring neuromodulation. Cortico-reticulo-spinal circuit reorganization may also improve recovery in humans.
Asunto(s)
Corteza Motora/fisiología , Recuperación de la Función/fisiología , Traumatismos de la Médula Espinal/patología , Traumatismos de la Médula Espinal/fisiopatología , Médula Espinal/fisiología , Núcleo Vestibular Lateral/fisiología , 8-Hidroxi-2-(di-n-propilamino)tetralin/farmacología , Animales , Encéfalo/anatomía & histología , Encéfalo/efectos de los fármacos , Channelrhodopsins/genética , Channelrhodopsins/metabolismo , Modelos Animales de Enfermedad , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Corteza Motora/efectos de los fármacos , Desempeño Psicomotor/efectos de los fármacos , Quipazina/farmacología , Ratas , Ratas Endogámicas Lew , Recuperación de la Función/efectos de los fármacos , Recuperación de la Función/genética , Agonistas de Receptores de Serotonina/farmacología , Médula Espinal/efectos de los fármacos , Traumatismos de la Médula Espinal/diagnóstico por imagen , Traumatismos de la Médula Espinal/tratamiento farmacológico , Antígenos Thy-1/administración & dosificación , Antígenos Thy-1/genética , Antígenos Thy-1/metabolismo , Núcleo Vestibular Lateral/efectos de los fármacosRESUMEN
Robotic exoskeletons provide programmable, consistent and controllable active therapeutic assistance to patients with neurological disorders. Here we introduce a prototype and preliminary experimental evaluation of a rehabilitative gait exoskeleton that enables compliant yet effective manipulation of the fragile limbs of rats. To assist the displacements of the lower limbs without impeding natural gait movements, we designed and fabricated soft pneumatic actuators (SPAs). The exoskeleton integrates two customizable SPAs that are attached to a limb. This configuration enables a 1 N force load, a range of motion exceeding 80 mm in the major axis, and speed of actuation reaching two gait cycles/s. Preliminary experiments in rats with spinal cord injury validated the basic features of the exoskeleton. We propose strategies to improve the performance of the robot and discuss the potential of SPAs for the design of other wearable interfaces.
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Miembros Artificiales/veterinaria , Dispositivo Exoesqueleto/veterinaria , Trastornos Neurológicos de la Marcha/fisiopatología , Trastornos Neurológicos de la Marcha/rehabilitación , Rehabilitación Neurológica/instrumentación , Robótica/instrumentación , Animales , Módulo de Elasticidad , Diseño de Equipo/veterinaria , Análisis de Falla de Equipo , Estudios de Factibilidad , Femenino , Trastornos Neurológicos de la Marcha/diagnóstico , Proyectos Piloto , Ratas , Ratas Endogámicas Lew , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Resultado del TratamientoRESUMEN
Medium-sized rings are widely considered to be under-represented in biological screening libraries for lead identification in medicinal chemistry. To help address this, a library of medium-sized lactams has been generated by using a simple, scalable and versatile ring-expansion protocol. Analysis of the library by using open-access computational tool LLAMA suggested that these lactams and their derivatives have highly promising physicochemical and 3D spatial properties and thus have much potential in drug discovery.
Asunto(s)
Lactamas/química , Aminoácidos/química , Ciclización , Diseño de Fármacos , Lactamas/síntesis químicaRESUMEN
OBJECTIVES: We aimed to develop a robotic interface capable of providing finely-tuned, multidirectional trunk assistance adjusted in real-time during unconstrained locomotion in rats and mice. APPROACH: We interfaced a large-scale robotic structure actuated in four degrees of freedom to exchangeable attachment modules exhibiting selective compliance along distinct directions. This combination allowed high-precision force and torque control in multiple directions over a large workspace. We next designed a neurorobotic platform wherein real-time kinematics and physiological signals directly adjust robotic actuation and prosthetic actions. We tested the performance of this platform in both rats and mice with spinal cord injury. MAIN RESULTS: Kinematic analyses showed that the robotic interface did not impede locomotor movements of lightweight mice that walked freely along paths with changing directions and height profiles. Personalized trunk assistance instantly enabled coordinated locomotion in mice and rats with severe hindlimb motor deficits. Closed-loop control of robotic actuation based on ongoing movement features enabled real-time control of electromyographic activity in anti-gravity muscles during locomotion. SIGNIFICANCE: This neurorobotic platform will support the study of the mechanisms underlying the therapeutic effects of locomotor prosthetics and rehabilitation using high-resolution genetic tools in rodent models.
Asunto(s)
Diseño de Equipo/métodos , Locomoción/fisiología , Prótesis Neurales , Robótica/métodos , Animales , Femenino , Miembro Posterior/inervación , Miembro Posterior/fisiopatología , Miembro Posterior/cirugía , Ratones , Ratones Endogámicos C57BL , Prótesis Neurales/tendencias , Ratas , Ratas Endogámicas Lew , Robótica/tendencias , Traumatismos de la Médula Espinal/fisiopatología , Traumatismos de la Médula Espinal/rehabilitación , Traumatismos de la Médula Espinal/cirugíaRESUMEN
Electrical neuromodulation of lumbar segments improves motor control after spinal cord injury in animal models and humans. However, the physiological principles underlying the effect of this intervention remain poorly understood, which has limited the therapeutic approach to continuous stimulation applied to restricted spinal cord locations. Here we developed stimulation protocols that reproduce the natural dynamics of motoneuron activation during locomotion. For this, we computed the spatiotemporal activation pattern of muscle synergies during locomotion in healthy rats. Computer simulations identified optimal electrode locations to target each synergy through the recruitment of proprioceptive feedback circuits. This framework steered the design of spatially selective spinal implants and real-time control software that modulate extensor and flexor synergies with precise temporal resolution. Spatiotemporal neuromodulation therapies improved gait quality, weight-bearing capacity, endurance and skilled locomotion in several rodent models of spinal cord injury. These new concepts are directly translatable to strategies to improve motor control in humans.
Asunto(s)
Potenciales Evocados Motores/fisiología , Retroalimentación Sensorial/fisiología , Miembro Posterior/fisiopatología , Locomoción/fisiología , Neuronas Motoras/fisiología , Músculo Esquelético/fisiopatología , Traumatismos de la Médula Espinal/fisiopatología , Estimulación de la Médula Espinal , Raíces Nerviosas Espinales/fisiopatología , Animales , Fenómenos Biomecánicos , Simulación por Computador , Femenino , Miembro Posterior/inervación , Cinética , Músculo Esquelético/inervación , Ratas , Ratas Endogámicas Lew , Médula Espinal/fisiología , Traumatismos de la Médula Espinal/patología , Traumatismos de la Médula Espinal/rehabilitación , Factores de Tiempo , Microtomografía por Rayos XRESUMEN
INTRODUCTION: Parkinson's disease (PD) is a progressive neurodegenerative disorder characterized by the loss of dopaminergic neurons as well as the presence of proteinaceous inclusions named Lewy bodies. α-synuclein (α-syn) is a major constituent of Lewy bodies, and the first disease-causing protein characterized in PD. Several α-syn-based animal models of PD have been developed to investigate the pathophysiology of PD, but none of them recapitulate the full picture of the disease. Ageing is the most compelling and major risk factor for developing PD but its impact on α-syn toxicity remains however unexplored. In this study, we developed and exploited a recombinant adeno-associated viral (AAV) vector of serotype 9 overexpressing mutated α-syn to elucidate the influence of ageing on the dynamics of PD-related neurodegeneration associated with α-syn pathology in different mammalian species. RESULTS: Identical AAV pseudotype 2/9 vectors carrying the DNA for human mutant p.A53T α-syn were injected into the substantia nigra to induce neurodegeneration and synucleinopathy in mice, rats and monkeys. Rats were used first to validate the ability of this serotype to replicate α-syn pathology and second to investigate the relationship between the kinetics of α-syn-induced nigrostriatal degeneration and the progressive onset of motor dysfunctions, strikingly reminiscent of the impairments observed in PD patients. In mice, AAV2/9-hα-syn injection into the substantia nigra was associated with accumulation of α-syn and phosphorylated hα-syn, regardless of mouse strain. However, phenotypic mutants with either accelerated senescence or resistance to senescence did not display differential susceptibility to hα-syn overexpression. Of note, p-α-syn levels correlated with nigrostriatal degeneration in mice. In monkeys, hα-syn-induced degeneration of the nigrostriatal pathway was not affected by the age of the animals. Unlike mice, monkeys did not exhibit correlations between levels of phosphorylated α-syn and neurodegeneration. CONCLUSIONS: In conclusion, AAV2/9-mediated hα-syn induces robust nigrostriatal neurodegeneration in mice, rats and monkeys, allowing translational comparisons among species. Ageing, however, neither exacerbated nigrostriatal neurodegeneration nor α-syn pathology per se. Our unprecedented multi-species investigation thus favours the multiple-hit hypothesis for PD wherein ageing would merely be an aggravating, additive, factor superimposed upon an independent disease process.