RESUMEN
Axonal transport of organelles has emerged as a key process in the regulation of neuronal differentiation and survival. Several components of this specialised transport machinery, their regulators and vesicular cargoes are mutated or altered in many neurodegenerative conditions. The molecular characterisation of these mechanisms has furthered our understanding of neuronal homeostasis, providing insights into the spatio-temporal control of membrane traffic and signalling in neurons with a precision not achievable in other cellular systems. Here, we summarise the recent advances in the field of axonal trafficking of different organelles, and the essential role of motor and adaptor proteins in this process.
Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Transporte Axonal , Axones/fisiología , Proteínas Motoras Moleculares/fisiología , Enfermedades Neurodegenerativas/metabolismo , Orgánulos/fisiología , Animales , Humanos , Enfermedades Neurodegenerativas/patologíaRESUMEN
Spinal and bulbar muscular atrophy (SBMA), or Kennedy's disease, is a late-onset motor neuron disease (MND) caused by an abnormal expansion of the CAG repeat in the androgen receptor (AR) gene on the X-chromosome, encoding a polyglutamine (poly-Q) sequence in the protein product. Mutant poly-Q-expanded AR protein is widely expressed but leads to selective lower motoneuron death. Although the mechanisms that underlie SBMA remain unclear, defective axonal transport has been implicated in MND and other forms of poly-Q disease. Transcriptional dysregulation may also be involved in poly-Q repeat pathology. We therefore examined axonal transport in a mouse model of SBMA recapitulating many aspects of the human disease. We found no difference in the expression levels of motor and the microtubule-associated protein tau, in the spinal cord and sciatic nerve of wild-type (WT) and SBMA mice at various stages of disease progression. Furthermore, we found no alteration in binding properties of motor proteins and tau to microtubules. Moreover, analysis of axonal transport rates both in cultured primary motoneurons in vitro and in vivo in the sciatic nerve of adult WT and mutant SBMA mice demonstrated no overt axonal transport deficits in these systems. Our results therefore indicate that unlike other motoneuron and poly-Q diseases, axonal transport deficits do not play a significant role in the pathogenesis of SBMA.
Asunto(s)
Transporte Axonal , Atrofia Muscular Espinal/metabolismo , Atrofia Muscular/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Expresión Génica , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuronas Motoras/metabolismo , Atrofia Muscular/genética , Atrofia Muscular Espinal/genética , Proteínas tau/genética , Proteínas tau/metabolismoRESUMEN
ALS is a fatal neurodegenerative disease characterized by selective motor neuron death resulting in muscle paralysis. Mutations in superoxide dismutase 1 (SOD1) are responsible for a subset of familial cases of ALS. Although evidence from transgenic mice expressing human mutant SOD1(G93A) suggests that axonal transport defects may contribute to ALS pathogenesis, our understanding of how these relate to disease progression remains unclear. Using an in vivo assay that allows the characterization of axonal transport in single axons in the intact sciatic nerve, we have identified clear axonal transport deficits in presymptomatic mutant mice. An impairment of axonal retrograde transport may therefore represent one of the earliest axonal pathologies in SOD1(G93A) mice, which worsens at an early symptomatic stage. A deficit in axonal transport may therefore be a key pathogenic event in ALS and an early disease indicator of motor neuron degeneration.
Asunto(s)
Esclerosis Amiotrófica Lateral/fisiopatología , Transporte Axonal/fisiología , Neuronas Motoras/patología , Superóxido Dismutasa/genética , Animales , Progresión de la Enfermedad , Quimografía , Ratones , Mitocondrias/fisiología , Neuronas Motoras/fisiología , Mutación/genética , Nervio Ciático/citología , Superóxido Dismutasa-1RESUMEN
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease resulting in the selective death of motor neurons in the brain and spinal cord. Some familial cases of ALS are caused by dominant mutations in the gene encoding superoxide dismutase (SOD1). The emergence of interfering RNA (RNAi) for specific gene silencing could be therapeutically beneficial for the treatment of such dominantly inherited diseases. We generated a lentiviral vector to mediate expression of RNAi molecules specifically targeting the human SOD1 gene (SOD1). Injection of this vector into various muscle groups of mice engineered to overexpress a mutated form of human SOD1 (SOD1(G93A)) resulted in an efficient and specific reduction of SOD1 expression and improved survival of vulnerable motor neurons in the brainstem and spinal cord. Furthermore, SOD1 silencing mediated an improved motor performance in these animals, resulting in a considerable delay in the onset of ALS symptoms by more than 100% and an extension in survival by nearly 80% of their normal life span. These data are the first to show a substantial extension of survival in an animal model of a fatal, dominantly inherited neurodegenerative condition using RNAi and provide the highest therapeutic efficacy observed in this field to date.
Asunto(s)
Esclerosis Amiotrófica Lateral/genética , Degeneración Nerviosa , Interferencia de ARN , Superóxido Dismutasa/genética , Esclerosis Amiotrófica Lateral/terapia , Animales , Modelos Animales de Enfermedad , Terapia Genética , Vectores Genéticos , Humanos , Lentivirus/genética , Ratones , Ratones Transgénicos , Mutación , ARN Interferente Pequeño , Tasa de Supervivencia , TransfecciónRESUMEN
Axonal transport is responsible for the movement of signals and cargo between nerve termini and cell bodies. Pathogens also exploit this pathway to enter and exit the central nervous system. In this study, we characterised the binding, endocytosis and axonal transport of an adenovirus (CAV-2) that preferentially infects neurons. Using biochemical, cell biology, genetic, ultrastructural and live-cell imaging approaches, we show that interaction with the neuronal membrane correlates with coxsackievirus and adenovirus receptor (CAR) surface expression, followed by endocytosis involving clathrin. In axons, long-range CAV-2 motility was bidirectional with a bias for retrograde transport in nonacidic Rab7-positive organelles. Unexpectedly, we found that CAR was associated with CAV-2 vesicles that also transported cargo as functionally distinct as tetanus toxin, neurotrophins, and their receptors. These results suggest that a single axonal transport carrier is capable of transporting functionally distinct cargoes that target different membrane compartments in the soma. We propose that CAV-2 transport is dictated by an innate trafficking of CAR, suggesting an unsuspected function for this adhesion protein during neuronal homeostasis.
Asunto(s)
Adenoviridae/metabolismo , Transporte Axonal , Axones/virología , Neuronas Motoras/virología , Receptores Citoplasmáticos y Nucleares/metabolismo , Factores de Transcripción/metabolismo , Animales , Carbocianinas/metabolismo , Células Cultivadas , Vesículas Cubiertas por Clatrina/ultraestructura , Vesículas Cubiertas por Clatrina/virología , Invaginaciones Cubiertas de la Membrana Celular/ultraestructura , Invaginaciones Cubiertas de la Membrana Celular/virología , Receptor de Androstano Constitutivo , Endocitosis , Endosomas/metabolismo , Endosomas/virología , Colorantes Fluorescentes/metabolismo , Ganglios Espinales/metabolismo , Ganglios Espinales/ultraestructura , Ganglios Espinales/virología , Concentración de Iones de Hidrógeno , Ratones , Ratones Endogámicos C57BL , Microscopía Fluorescente , Neuronas Motoras/metabolismo , Neuronas Motoras/ultraestructura , Proteínas del Tejido Nervioso/metabolismo , Ratas , Nervio Ciático/metabolismo , Proteínas de Transporte Vesicular/metabolismoRESUMEN
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by motoneuron degeneration resulting in paralysis and eventual death. ALS is regarded as a motoneuron-specific disorder but increasing evidence indicates non-neuronal cells play a significant role in disease pathogenesis. Although the precise aetiology of ALS remains unclear, mutations in the superoxide dismutase (SOD1) gene are known to account for approximately 20% of familial ALS. We examined the influence of SOD1(G93A) expression in astrocytes on mitochondrial homeostasis in motoneurons in a primary astrocyte : motoneuron co-culture model. SOD1(G93A) expression in astrocytes induced changes in mitochondrial function of both SOD1(G93A) and wild-type motoneurons. In the presence of SOD1(G93A) astrocytes, mitochondrial redox state of both wild-type and SOD1(G93A) motoneurons was more reduced and mitochondrial membrane potential decreased. While intra-mitochondrial calcium levels [Ca(2+)](m) were elevated in SOD1(G93A) motoneurons, changes in mitochondrial function did not correlate with [Ca(2+)](m). Thus, expression of SOD1(G93A) in astrocytes directly alters mitochondrial function even in embryonic motoneurons, irrespective of genotype. These early deficits in mitochondrial function induced by surrounding astrocytes may increase the vulnerability of motoneurons to other neurotoxic mechanisms involved in ALS pathogenesis.
Asunto(s)
Astrocitos/metabolismo , Mitocondrias/fisiología , Neuronas Motoras/metabolismo , Superóxido Dismutasa/metabolismo , Animales , Astrocitos/citología , Astrocitos/efectos de los fármacos , Calcio/metabolismo , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Técnicas de Cocultivo/métodos , Fluoresceínas/metabolismo , Expresión Génica/efectos de los fármacos , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Hidroliasas/metabolismo , Inmunohistoquímica , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Potencial de la Membrana Mitocondrial/fisiología , Ratones , Ratones Transgénicos , Microscopía Confocal , Proteínas Asociadas a Microtúbulos/metabolismo , Mitocondrias/efectos de los fármacos , Neuronas Motoras/citología , Neuronas Motoras/efectos de los fármacos , Oxidación-Reducción/efectos de los fármacos , Médula Espinal/citología , Superóxido Dismutasa/genética , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiónico/farmacologíaRESUMEN
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by the selective loss of motoneurons in the spinal cord, brain stem, and motor cortex. However, despite intensive research, an effective treatment for this disease remains elusive. In this study we show that treatment of postsymptomatic, 90-day-old SOD1G93A mice with a synthetic cannabinoid, WIN55,212-2, significantly delays disease progression. Furthermore, genetic ablation of the Faah enzyme, which results in raised levels of the endocannabinoid anandamide, prevented the appearance of disease signs in 90-day-old SOD1G93A mice. Surprisingly, elevation of cannabinoid levels with either WIN55,212-2 or Faah ablation had no effect on life span. Ablation of the CB1 receptor, in contrast, had no effect on disease onset in SOD1(G93A) mice but significantly extended life span. Together these results show that cannabinoids have significant neuroprotective effects in this model of ALS and suggest that these beneficial effects may be mediated by non-CB1 receptor mechanisms.
Asunto(s)
Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Esclerosis Amiotrófica Lateral/genética , Cannabinoides/metabolismo , Morfolinas/farmacología , Morfolinas/uso terapéutico , Naftalenos/farmacología , Naftalenos/uso terapéutico , Superóxido Dismutasa/genética , Amidohidrolasas/genética , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Benzoxazinas , Moléculas de Adhesión Celular Neuronal/genética , Progresión de la Enfermedad , Femenino , Humanos , Longevidad/efectos de los fármacos , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Neuronas Motoras/metabolismo , Fatiga Muscular/efectos de los fármacos , Fatiga Muscular/genética , Músculo Esquelético/citología , Músculo Esquelético/efectos de los fármacos , Neuropéptidos/genética , Protocadherinas , Receptores de Superficie Celular/genética , Superóxido Dismutasa/metabolismoRESUMEN
Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative condition characterised by the selective loss of motor neurons from the spinal cord, brainstem and motor cortex. Although the pathogenic mechanisms that underlie ALS are not yet fully understood, there is significant evidence that several neurotoxic mechanisms including excitotoxicity, inflammation and oxidative stress, all contribute to disease pathogenesis. Furthermore, recent results have established that although primarily a motor neuron specific disorder, ALS is not cell-autonomous and non-neuronal cells including astroglia and microglia play a critical role in mechanism of disease. Currently the only licensed therapy available for the treatment of ALS is the anti-glutamatergic agent Riluzole, which has limited therapeutic effects. However, there is increasing evidence that cannabinoids and manipulation of the endocannabinoid system may have therapeutic value in ALS, in addition to other neurodegenerative conditions. Cannabinoids exert anti-glutamatergic and anti-inflammatory actions through activation of the CB(1) and CB(2) receptors, respectively. Activation of CB(1) receptors may therefore inhibit glutamate release from presynaptic nerve terminals and reduce the postsynaptic calcium influx in response to glutamate receptor stimulation. Meanwhile, CB(2) receptors may influence inflammation, whereby receptor activation reduces microglial activation, resulting in a decrease in microglial secretion of neurotoxic mediators. Finally, cannabinoid agents may also exert anti-oxidant actions by a receptor-independent mechanism. Therefore the ability of cannabinoids to target multiple neurotoxic pathways in different cell populations may increase their therapeutic potential in the treatment of ALS. Recent studies investigating this potential in models of ALS, in particular those that focus on strategies that activate CB(2) receptors, are discussed in this review.