RESUMEN
The pathophysiology of tremor in Parkinson's disease (PD) is evolving towards a complex alteration to monoaminergic innervation, and increasing evidence suggests a key role of the locus coeruleus noradrenergic system (LC-NA). However, the difficulties in imaging LC-NA in patients challenge its direct investigation. To this end, we studied the development of tremor in a reserpinized rat model of PD, with or without a selective lesioning of LC-NA innervation with the neurotoxin DSP-4. Eight male rats (Sprague Dawley) received DSP-4 (50 mg/kg) two weeks prior to reserpine injection (10 mg/kg) (DR-group), while seven male animals received only reserpine treatment (R-group). Tremor, rigidity, hypokinesia, postural flexion and postural immobility were scored before and after 20, 40, 60, 80, 120 and 180 min of reserpine injection. Tremor was assessed visually and with accelerometers. The injection of DSP-4 induced a severe reduction in LC-NA terminal axons (DR-group: 0.024 ± 0.01 vs. R-group: 0.27 ± 0.04 axons/um2, p < 0.001) and was associated with significantly less tremor, as compared to the R-group (peak tremor score, DR-group: 0.5 ± 0.8 vs. R-group: 1.6 ± 0.5; p < 0.01). Kinematic measurement confirmed the clinical data (tremor consistency (% of tremor during 180 s recording), DR-group: 37.9 ± 35.8 vs. R-group: 69.3 ± 29.6; p < 0.05). Akinetic-rigid symptoms did not differ between the DR- and R-groups. Our results provide preliminary causal evidence for a critical role of LC-NA innervation in the development of PD tremor and foster the development of targeted therapies for PD patients.
Asunto(s)
Enfermedad de Parkinson , Temblor , Humanos , Masculino , Animales , Ratas , Ratas Sprague-Dawley , Temblor/inducido químicamente , Reserpina/farmacología , Encéfalo , NorepinefrinaRESUMEN
The serotonin transporter (5-HTT) is a key molecule of serotoninergic neurotransmission and target of many anxiolytics and antidepressants. In humans, 5-HTT gene variants resulting in lower expression levels are associated with behavioral traits of anxiety. Furthermore, functional magnetic resonance imaging (fMRI) studies reported increased cerebral blood flow (CBF) during resting state (RS) and amygdala hyperreactivity. 5-HTT deficient mice as an established animal model for anxiety disorders seem to be well suited for investigating amygdala (re-)activity in an fMRI study. We investigated wildtype (5-HTT+/+), heterozygous (5-HTT+/-), and homozygous 5-HTT-knockout mice (5-HTT-/-) of both sexes in an ultra-high-field 17.6 Tesla magnetic resonance scanner. CBF was measured with continuous arterial spin labeling during RS, stimulation state (SS; with odor of rats as aversive stimulus), and post-stimulation state (PS). Subsequently, post mortem c-Fos immunohistochemistry elucidated neural activation on cellular level. The results showed that in reaction to the aversive odor CBF in total brain and amygdala of all mice significantly increased. In male 5-HTT+/+ mice amygdala RS CBF levels were found to be significantly lower than in 5-HTT+/- mice. From RS to SS 5-HTT+/+ amygdala perfusion significantly increased compared to both 5-HTT+/- and 5-HTT-/- mice. Perfusion level changes of male mice correlated with the density of c-Fos-immunoreactive cells in the amygdaloid nuclei. In female mice the perfusion was not modulated by the 5-Htt-genotype, but by estrous cycle stages. We conclude that amygdala reactivity is modulated by the 5-Htt genotype in males. In females, gonadal hormones have an impact which might have obscured genotype effects. Furthermore, our results demonstrate experimental support for the tonic model of 5-HTTLPR function.
Asunto(s)
Amígdala del Cerebelo/irrigación sanguínea , Ansiedad/diagnóstico por imagen , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas de Transporte de Serotonina en la Membrana Plasmática/deficiencia , Amígdala del Cerebelo/metabolismo , Animales , Ansiedad/genética , Circulación Cerebrovascular , Modelos Animales de Enfermedad , Femenino , Hormonas Gonadales/metabolismo , Homocigoto , Imagen por Resonancia Magnética , Masculino , Ratones , Ratones Noqueados , Ratas , Proteínas de Transporte de Serotonina en la Membrana Plasmática/genética , Caracteres SexualesRESUMEN
Since exposure therapy for anxiety disorders incorporates extinction of contextual anxiety, relapses may be due to reinstatement processes. Animal research demonstrated more stable extinction memory and less anxiety relapse due to vagus nerve stimulation (VNS). We report a valid human three-day context conditioning, extinction and return of anxiety protocol, which we used to examine effects of transcutaneous VNS (tVNS). Seventy-five healthy participants received electric stimuli (unconditioned stimuli, US) during acquisition (Day1) when guided through one virtual office (anxiety context, CTX+) but never in another (safety context, CTX-). During extinction (Day2), participants received tVNS, sham, or no stimulation and revisited both contexts without US delivery. On Day3, participants received three USs for reinstatement followed by a test phase. Successful acquisition, i.e. startle potentiation, lower valence, higher arousal, anxiety and contingency ratings in CTX+ versus CTX-, the disappearance of these effects during extinction, and successful reinstatement indicate validity of this paradigm. Interestingly, we found generalized reinstatement in startle responses and differential reinstatement in valence ratings. Altogether, our protocol serves as valid conditioning paradigm. Reinstatement effects indicate different anxiety networks underlying physiological versus verbal responses. However, tVNS did neither affect extinction nor reinstatement, which asks for validation and improvement of the stimulation protocol.
Asunto(s)
Trastornos de Ansiedad/fisiopatología , Ansiedad/fisiopatología , Condicionamiento Clásico/fisiología , Nervio Vago/fisiopatología , Adulto , Nivel de Alerta/fisiología , Extinción Psicológica/fisiología , Femenino , Humanos , Masculino , Memoria/fisiología , Reflejo de Sobresalto/fisiología , Estimulación Eléctrica Transcutánea del Nervio/métodos , Estimulación del Nervio Vago/métodos , Realidad Virtual , Adulto JovenRESUMEN
Adhesion-type G protein-coupled receptors (aGPCRs), a large molecule family with over 30 members in humans, operate in organ development, brain function and govern immunological responses. Correspondingly, this receptor family is linked to a multitude of diverse human diseases. aGPCRs have been suggested to possess mechanosensory properties, though their mechanism of action is fully unknown. Here we show that the Drosophila aGPCR Latrophilin/dCIRL acts in mechanosensory neurons by modulating ionotropic receptor currents, the initiating step of cellular mechanosensation. This process depends on the length of the extended ectodomain and the tethered agonist of the receptor, but not on its autoproteolysis, a characteristic biochemical feature of the aGPCR family. Intracellularly, dCIRL quenches cAMP levels upon mechanical activation thereby specifically increasing the mechanosensitivity of neurons. These results provide direct evidence that the aGPCR dCIRL acts as a molecular sensor and signal transducer that detects and converts mechanical stimuli into a metabotropic response.
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Potenciales de Acción , AMP Cíclico/metabolismo , Proteínas de Drosophila/metabolismo , Mecanorreceptores/fisiología , Receptores de Péptidos/metabolismo , Células Receptoras Sensoriales/fisiología , Animales , Drosophila , Fenómenos Electrofisiológicos , Imagen ÓpticaRESUMEN
BEACH domain proteins are involved in membrane protein traffic and human diseases, but their molecular mechanisms are not understood. The BEACH protein LRBA has been implicated in immune response and cell proliferation, and human LRBA mutations cause severe immune deficiency. Here, we report a first functional and molecular phenotype outside the immune system of LRBA-knockout mice: compromised olfaction, manifesting in reduced electro-olfactogram response amplitude, impaired food-finding efficiency, and smaller olfactory bulbs. LRBA is prominently expressed in olfactory and vomeronasal chemosensory neurons of wild-type mice. Olfactory impairment in the LRBA-KO is explained by markedly reduced concentrations (20-40% of wild-type levels) of all three subunits αolf, ß1 and γ13 of the olfactory heterotrimeric G-protein, Golf, in the sensory cilia of olfactory neurons. In contrast, cilia morphology and the concentrations of many other proteins of olfactory cilia are not or only slightly affected. LRBA is also highly expressed in photoreceptor cells, another cell type with a specialized sensory cilium and heterotrimeric G-protein-based signalling; however, visual function appeared unimpaired by the LRBA-KO. To our knowledge, this is the first observation that a BEACH protein is required for the efficient subcellular localization of a lipid-anchored protein, and of a ciliary protein.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Cilios/metabolismo , Subunidades alfa de la Proteína de Unión al GTP/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Electrorretinografía , Femenino , Regulación de la Expresión Génica , Aparato de Golgi/efectos de los fármacos , Aparato de Golgi/metabolismo , Masculino , Ratones Noqueados , Ratones Transgénicos , Trastornos del Olfato/genética , Bulbo Olfatorio/metabolismo , Bulbo Olfatorio/patología , Neuronas Receptoras Olfatorias/metabolismo , Dominios Proteicos , Retina/anomalías , Órgano Vomeronasal/citología , Órgano Vomeronasal/metabolismoRESUMEN
Current research supports the notion that the apparently innate trait Sensory Processing Sensitivity (SPS) may act as a modulator of development as function of the environment. Interestingly, the common serotonin transporter linked polymorphic region (5-HTTLPR) does the same. While neural mechanisms underlying SPS are largely unknown, those associated with the 5-HTTLPR have been extensively investigated. We perform a comparative analysis of research findings on sensory processing facets associated with the trait and polymorphism to: 1. detect shared phenotypes and frame a hypothesis towards neural mechanisms underlying SPS; 2. increase the understanding of 5-HTTLPR-associated behavioral patterns. Trait and polymorphism are both associated with differential susceptibility to environmental stimuli; additionally, both involve 1. having stronger emotional reactions, 2. processing of sensory information more deeply, 3. being more aware of environmental subtleties, and 4. being easily overstimulated. We discuss neural mechanisms and environmental conditions that may underlie these four facets. Besides urging the actual assessment of the link between the two, the conclusions of our analyses may guide and focus future research strategies.
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Serotonina/metabolismo , Cognición , Emociones , Genotipo , Humanos , Imagen por Resonancia Magnética , Proteínas de Transporte de Serotonina en la Membrana PlasmáticaRESUMEN
The cholinergic and dopaminergic innervation of the amygdala plays an important role in attention, emotional arousal, aversive forms of associative learning, conditioned responses, and stress responsivity. Roman High- (RHA) and Low-Avoidance (RLA) rats are an ideal model to study the potential impact of this innervation on behavioral responses, because they were selected bidirectionally for differences in their two-way active avoidance performance. RHA rats are known to quickly acquire two-way active avoidance and show indications of enhanced impulsive behavior, novelty seeking, and vulnerability to substance abuse, whereas RLA rats exhibit a passive coping style with high levels of immobility and enhanced stress responsivity. In the present study, the density of acetylcholine esterase (AchE)-positive cholinergic fibers and tyrosine hydroxylase immunoreactive (TH-ir) fibers were analyzed in various amygdala nuclei. In comparison to RLA rats, RHA rats displayed a significantly higher density of AchE-positive fibers in the lateral nucleus (La), the major sensory input area of the amygdala. In contrast, RLA rats showed a higher density of TH-ir fibers in the lateral division of the central nucleus (CeL), which modulates amygdala output and is known to contain more corticotropin-releasing hormone (CRH) positive neurons in RLA than in RHA rats. The findings suggest that a higher density of AchE-positive fibers in the La of RHA rats may facilitate attentional mechanisms and aversive forms of associative learning in RHA rats, whereas the increased density of TH-ir fibers in the CeL of RLA rats may be involved in the regulation of enhanced CRH expression and stress responsivity.
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Acetilcolinesterasa/metabolismo , Amígdala del Cerebelo/metabolismo , Reacción de Prevención/fisiología , Fibras Nerviosas/metabolismo , Tirosina 3-Monooxigenasa/metabolismo , Animales , Hormona Liberadora de Corticotropina/metabolismo , Conducta Impulsiva/fisiología , Ratas , Ratas EndogámicasRESUMEN
Deep brain stimulation of the subthalamic nucleus is an effective treatment option for Parkinson's disease. In our lab we established a protocol to screen different neurostimulation patterns in hemiparkinsonian (unilateral lesioned) rats. It consists of creating a unilateral Parkinson's lesion by injecting 6-hydroxydopamine (6-OHDA) into the right medial forebrain bundle, implanting chronic stimulation electrodes into the subthalamic nucleus and evaluating motor outcomes at the end of 24 hr periods of cable-bound external neurostimulation. The stimulation was conducted with constant current stimulation. The amplitude was set 20% below the individual threshold for side effects. The motor outcome evaluation was done by the assessment of spontaneous paw use in the cylinder test according to Shallert and by the assessment of skilled reaching in the staircase test according to Montoya. This protocol describes in detail the training in the staircase box, the cylinder test, as well as the use of both in hemiparkinsonian rats. The use of both tests is necessary, because the staircase test seems to be more sensitive for fine motor skill impairment and exhibits greater sensitivity to change during neurostimulation. The combination of the unilateral Parkinson model and the two behavioral tests allows the assessment of different stimulation parameters in a standardized way.
Asunto(s)
Estimulación Encefálica Profunda , Núcleo Subtalámico , Animales , Haz Prosencefálico Medial/efectos de los fármacos , Haz Prosencefálico Medial/patología , Oxidopamina/toxicidad , Enfermedad de Parkinson/etiología , Enfermedad de Parkinson/fisiopatología , Enfermedad de Parkinson/terapia , RatasRESUMEN
Synapse-associated protein 1 (Syap1/BSTA) is the mammalian homologue of Sap47 (synapse-associated protein of 47 kDa) in Drosophila. Sap47 null mutant larvae show reduced short-term synaptic plasticity and a defect in associative behavioral plasticity. In cultured adipocytes, Syap1 functions as part of a complex that phosphorylates protein kinase Bα/Akt1 (Akt1) at Ser(473) and promotes differentiation. The role of Syap1 in the vertebrate nervous system is unknown. Here, we generated a Syap1 knock-out mouse and show that lack of Syap1 is compatible with viability and fertility. Adult knock-out mice show no overt defects in brain morphology. In wild-type brain, Syap1 is found widely distributed in synaptic neuropil, notably in regions rich in glutamatergic synapses, but also in perinuclear structures associated with the Golgi apparatus of specific groups of neuronal cell bodies. In cultured motoneurons, Syap1 is located in axons and growth cones and is enriched in a perinuclear region partially overlapping with Golgi markers. We studied in detail the influence of Syap1 knockdown and knockout on structure and development of these cells. Importantly, Syap1 knockout does not affect motoneuron survival or axon growth. Unexpectedly, neither knockdown nor knockout of Syap1 in cultured motoneurons is associated with reduced Ser(473) or Thr(308) phosphorylation of Akt. Our findings demonstrate a widespread expression of Syap1 in the mouse central nervous system with regionally specific distribution patterns as illustrated in particular for olfactory bulb, hippocampus, and cerebellum.
Asunto(s)
Encéfalo/metabolismo , Neuronas Motoras/citología , Neuronas Motoras/metabolismo , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/metabolismo , Animales , Células Cultivadas , Péptidos y Proteínas de Señalización Intracelular , Ratones , Ratones Endogámicos , Ratones Noqueados , Proteínas del Tejido Nervioso/genéticaRESUMEN
BACKGROUND: Opioids are the gold standard for the treatment of acute pain despite serious side effects in the central and enteric nervous system. µ-opioid receptors (MOPs) are expressed and functional at the terminals of sensory axons, when activated by exogenous or endogenous ligands. However, the presence and function of MOP along nociceptive axons remains controversial particularly in naïve animals. Here, we characterized axonal MOPs by immunofluorescence, ultrastructural, and functional analyses. Furthermore, we evaluated hypertonic saline as a possible enhancer of opioid receptor function. RESULTS: Comparative immunolabeling showed that, among several tested antibodies, which all provided specific MOP detection in the rat central nervous system (CNS), only one monoclonal MOP-antibody yielded specificity and reproducibility for MOP detection in the rat peripheral nervous system including the sciatic nerve. Double immunolabeling documented that MOP immunoreactivity was confined to calcitonin gene-related peptide (CGRP) positive fibers and fiber bundles. Almost identical labeling and double labeling patterns were found using mcherry-immunolabeling on sciatic nerves of mice producing a MOP-mcherry fusion protein (MOP-mcherry knock-in mice). Preembedding immunogold electron microscopy on MOP-mcherry knock-in sciatic nerves indicated presence of MOP in cytoplasm and at membranes of unmyelinated axons. Application of [D-Ala(2), N-MePhe(4), Gly-ol]-enkephalin (DAMGO) or fentanyl dose-dependently inhibited depolarization-induced CGRP release from rat sciatic nerve axons ex vivo, which was blocked by naloxone. When the lipophilic opioid fentanyl was applied perisciatically in naïve Wistar rats, mechanical nociceptive thresholds increased. Subthreshold doses of fentanyl or the hydrophilic opioid DAMGO were only effective if injected together with hypertonic saline. In vitro, using ß-arrestin-2/MOP double-transfected human embryonic kidney cells, DAMGO as well as fentanyl lead to a recruitment of ß-arrestin-2 to the membrane followed by a ß-arrestin-2 reappearance in the cytosol and MOP internalization. Pretreatment with hypertonic saline prevented MOP internalization. CONCLUSION: MOPs are present and functional in the axonal membrane from naïve animals. Hypertonic saline acutely decreases ligand-induced internalization of MOP and thereby might improve MOP function. Further studies should explore potential clinical applications of opioids together with enhancers for regional analgesia.
Asunto(s)
Analgesia , Axones/metabolismo , Receptores Opioides mu/química , Receptores Opioides mu/metabolismo , Amígdala del Cerebelo/efectos de los fármacos , Amígdala del Cerebelo/metabolismo , Animales , Anticuerpos Monoclonales/metabolismo , Axones/efectos de los fármacos , Axones/ultraestructura , Conducta Animal/efectos de los fármacos , Péptido Relacionado con Gen de Calcitonina/metabolismo , Endocitosis/efectos de los fármacos , Encefalina Ala(2)-MeFe(4)-Gli(5)/farmacología , Femenino , Fentanilo/farmacología , Técnicas de Sustitución del Gen , Masculino , Potenciales de la Membrana/efectos de los fármacos , Ratones , Nocicepción/efectos de los fármacos , Potasio/farmacología , Ratas Wistar , Reproducibilidad de los Resultados , Nervio Ciático/efectos de los fármacos , Nervio Ciático/metabolismo , beta-Arrestinas/metabolismoRESUMEN
In neurons, microtubules form a dense array within axons, and the stability and function of this microtubule network is modulated by neurofilaments. Accumulation of neurofilaments has been observed in several forms of neurodegenerative diseases, but the mechanisms how elevated neurofilament levels destabilize axons are unknown so far. Here, we show that increased neurofilament expression in motor nerves of pmn mutant mice, a model of motoneuron disease, causes disturbed microtubule dynamics. The disease is caused by a point mutation in the tubulin-specific chaperone E (Tbce) gene, leading to an exchange of the most C-terminal amino acid tryptophan to glycine. As a consequence, the TBCE protein becomes instable which then results in destabilization of axonal microtubules and defects in axonal transport, in particular in motoneurons. Depletion of neurofilament increases the number and regrowth of microtubules in pmn mutant motoneurons and restores axon elongation. This effect is mediated by interaction of neurofilament with the stathmin complex. Accumulating neurofilaments associate with stathmin in axons of pmn mutant motoneurons. Depletion of neurofilament by Nefl knockout increases Stat3-stathmin interaction and stabilizes the microtubules in pmn mutant motoneurons. Consequently, counteracting enhanced neurofilament expression improves axonal maintenance and prolongs survival of pmn mutant mice. We propose that this mechanism could also be relevant for other neurodegenerative diseases in which neurofilament accumulation and loss of microtubules are prominent features.
Asunto(s)
Chaperonas Moleculares/metabolismo , Proteínas de Neurofilamentos/deficiencia , Factor de Transcripción STAT3/metabolismo , Estatmina/metabolismo , Animales , Axones/metabolismo , Axones/patología , Células Cultivadas , Estimación de Kaplan-Meier , Ratones Endogámicos C57BL , Ratones Transgénicos , Chaperonas Moleculares/genética , Actividad Motora/fisiología , Neuronas Motoras/metabolismo , Neuronas Motoras/patología , Proteínas de Neurofilamentos/genética , Fenotipo , Nervio Frénico/metabolismo , Nervio Frénico/patología , Nervio Ciático/metabolismo , Nervio Ciático/patología , Transducción de Señal , Médula Espinal/metabolismo , Médula Espinal/patologíaRESUMEN
Stiff-person syndrome is the prototype of a central nervous system disorder with autoantibodies targeting presynaptic antigens. Patients with paraneoplastic stiff-person syndrome may harbour autoantibodies to the BAR (Bin/Amphiphysin/Rvs) domain protein amphiphysin, which target its SH3 domain. These patients have neurophysiological signs of compromised central inhibition and respond to symptomatic treatment with medication enhancing GABAergic transmission. High frequency neurotransmission as observed in tonic GABAergic interneurons relies on fast exocytosis of neurotransmitters based on compensatory endocytosis. As amphiphysin is involved in clathrin-mediated endocytosis, patient autoantibodies are supposed to interfere with this function, leading to disinhibition by reduction of GABAergic neurotransmission. We here investigated the effects of human anti-amphiphysin autoantibodies on structural components of presynaptic boutons ex vivo and in vitro using electron microscopy and super-resolution direct stochastic optical reconstruction microscopy. Ultrastructural analysis of spinal cord presynaptic boutons was performed after in vivo intrathecal passive transfer of affinity-purified human anti-amphiphysin autoantibodies in rats and revealed signs of markedly disabled clathrin-mediated endocytosis. This was unmasked at high synaptic activity and characterized by a reduction of the presynaptic vesicle pool, clathrin coated intermediates, and endosome-like structures. Super-resolution microscopy of inhibitory GABAergic presynaptic boutons in primary neurons revealed that specific human anti-amphiphysin immunoglobulin G induced an increase of the essential vesicular protein synaptobrevin 2 and a reduction of synaptobrevin 7. This constellation suggests depletion of resting pool vesicles and trapping of releasable pool vesicular proteins at the plasma membrane. Similar effects were found in amphiphysin-deficient neurons from knockout mice. Application of specific patient antibodies did not show additional effects. Blocking alternative pathways of clathrin-independent endocytosis with brefeldin A reversed the autoantibody induced effects on molecular vesicle composition. Endophilin as an interaction partner of amphiphysin showed reduced clustering within presynaptic terminals. Collectively, these results point towards an autoantibody-induced structural disorganization in GABAergic synapses with profound changes in presynaptic vesicle pools, activation of alternative endocytic pathways, and potentially compensatory rearrangement of proteins involved in clathrin-mediated endocytosis. Our findings provide novel insights into synaptic pathomechanisms in a prototypic antibody-mediated central nervous system disease, which may serve as a proof-of-principle example in this evolving group of autoimmune disorders associated with autoantibodies to synaptic antigens.
Asunto(s)
Autoanticuerpos/administración & dosificación , Proteínas del Tejido Nervioso/administración & dosificación , Terminales Presinápticos/ultraestructura , Vesículas Sinápticas/ultraestructura , Animales , Autoanticuerpos/sangre , Células Cultivadas , Femenino , Humanos , Inyecciones Espinales , Ratones , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/sangre , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/ultraestructura , Embarazo , Terminales Presinápticos/efectos de los fármacos , Terminales Presinápticos/metabolismo , Ratas , Ratas Endogámicas Lew , Síndrome de la Persona Rígida/sangre , Síndrome de la Persona Rígida/diagnóstico , Vesículas Sinápticas/efectos de los fármacos , Vesículas Sinápticas/metabolismoRESUMEN
Although age-related changes in synaptic plasticity are an important focus within neuroscience, little is known about ultrastructural changes of synaptic morphology during aging. Here we report how aging affects synaptic ultrastructure by using fluorescence and electron microscopy at the adult Drosophila neuromuscular junction (NMJ) of ventral abdominal muscles. Mainly four striking morphological changes of aging NMJs were revealed. 1) Bouton size increases with proportionally rising number of active zones (AZs). 2) Synaptic vesicle density at AZs is increased in old flies. 3) Late endosomes, cisternae, and multivesicular bodies accumulate in the presynaptic terminal, and vesicles accumulate between membranes of the terminal bouton and the subsynaptic reticulum. 4) The electron-dense pre- and postsynaptic apposition is expanded in aging NMJs, which is accompanied by an expansion of the postsynaptic glutamate receptor fields. These findings suggest that aging is possibly accompanied by impaired synaptic vesicle release and recycling and a potentially compensatory expansion of AZs and postsynaptic densities.
Asunto(s)
Envejecimiento/patología , Drosophila melanogaster/ultraestructura , Unión Neuromuscular/ultraestructura , Terminales Presinápticos/ultraestructura , Animales , Tamaño de la Célula , Endosomas/ultraestructura , Femenino , Procesamiento de Imagen Asistido por Computador , Inmunohistoquímica , Microscopía Confocal , Microscopía Electrónica de Transmisión , Microscopía Fluorescente , Plasticidad Neuronal , Análisis de Supervivencia , Vesículas Sinápticas/ultraestructuraRESUMEN
In motoneuron disease and other neurodegenerative disorders, the loss of synapses and axon branches occurs early but is compensated by sprouting of neighboring axon terminals. Defective local axonal signaling for maintenance and dynamics of the axonal microtubule and actin cytoskeleton plays a central role in this context. The molecular mechanisms that lead to defective cytoskeleton architecture in two mouse models of motoneuron disease are summarized and discussed in this manuscript. In the progressive motor neuropathy (pmn) mouse model of motoneuron disease that is caused by a mutation in the tubulin-specific chaperone E gene, death of motoneuron cell bodies appears as a consequence of axonal degeneration. Treatment with bcl-2 overexpression or with glial-derived neurotrophic factor prevents loss of motoneuron cell bodies but does not influence the course of disease. In contrast, treatment with ciliary neurotrophic factor (CNTF) significantly delays disease onset and prolongs survival of pmn mice. This difference is due to the activation of Stat-3 via the CNTF receptor complex in axons of pmn mutant motoneurons. Most of the activated Stat-3 protein is not transported to the nucleus to activate transcription, but interacts locally in axons with stathmin, a protein that destabilizes microtubules. This interaction plays a major role in CNTF signaling for microtubule dynamics in axons. In Smn-deficient mice, a model of spinal muscular atrophy, defects in axonal translocation of ß-actin mRNA and possibly other mRNA species have been observed. Moreover, the regulation of local protein synthesis in response to signals from neurotrophic factors and extracellular matrix proteins is altered in motoneurons from this model of motoneuron disease. These findings indicate that local signals are important for maintenance and plasticity of axonal branches and neuromuscular endplates, and that disturbances in these signaling mechanisms could contribute to the pathophysiology of motoneuron diseases.
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Axones/fisiología , Enfermedad de la Neurona Motora/fisiopatología , Neuronas Motoras/fisiología , Plasticidad Neuronal/fisiología , Animales , Factor Neurotrófico Ciliar/fisiología , Modelos Animales de Enfermedad , Ratones , Placa Motora/fisiología , Factor de Transcripción STAT3/fisiología , Transducción de Señal/fisiologíaRESUMEN
Miz1 is a zinc finger protein that regulates the expression of cell cycle inhibitors as part of a complex with Myc. Cell cycle-independent functions of Miz1 are poorly understood. Here we use a Nestin-Cre transgene to delete an essential domain of Miz1 in the central nervous system (Miz1(ΔPOZNes)). Miz1(ΔPOZNes) mice display cerebellar neurodegeneration characterized by the progressive loss of Purkinje cells. Chromatin immunoprecipitation sequencing and biochemical analyses show that Miz1 activates transcription upon binding to a non-palindromic sequence present in core promoters. Target genes of Miz1 encode regulators of autophagy and proteins involved in vesicular transport that are required for autophagy. Miz1(ΔPOZ) neuronal progenitors and fibroblasts show reduced autophagic flux. Consistently, polyubiquitinated proteins and p62/Sqtm1 accumulate in the cerebella of Miz1(ΔPOZNes) mice, characteristic features of defective autophagy. Our data suggest that Miz1 may link cell growth and ribosome biogenesis to the transcriptional regulation of vesicular transport and autophagy.
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Autofagia/genética , Cerebelo/metabolismo , Proteínas Nucleares/genética , Proteínas Inhibidoras de STAT Activados/genética , Células de Purkinje/metabolismo , Vesículas Transportadoras/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Sitios de Unión , Cerebelo/patología , Femenino , Regulación de la Expresión Génica , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Integrasas/genética , Integrasas/metabolismo , Ratones , Ratones Noqueados , Proteínas Nucleares/metabolismo , Regiones Promotoras Genéticas , Unión Proteica , Proteínas Inhibidoras de STAT Activados/metabolismo , Estructura Terciaria de Proteína , Células de Purkinje/patología , Ribosomas/metabolismo , Análisis de Secuencia de ADN , Proteína Sequestosoma-1 , Transducción de Señal , Factor de Transcripción TFIIH , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Transcripción Genética , Ubiquitina-Proteína Ligasas , UbiquitinaciónRESUMEN
The amygdala is a core component of neural circuits that mediate processing of emotional, particularly anxiety and fear-related stimuli across species. In addition, the nuclear complex plays a key role in the central nervous system stress response, and alterations in amygdala responsivity are found in neuropsychiatric disorders, especially those precipitated or sustained by stressors. Serotonin has been shown to shape and fine-tune neural plasticity in development and adulthood, thereby allowing for network flexibility and adaptive capacity in response to environmental challenges, and is implicated in the modulation of stimulus processing and stress sensitivity in the amygdala. The fact that altered amygdala activity patterns are observed upon pharmacological manipulations of serotonergic transmission, as well as in carriers of genetic variations in serotonin pathway-associated signaling molecules representing risk factors for neuropsychiatric disorders, underlines the importance of understanding the role and mode of action of serotonergic transmission in the amygdala for human psychopathology. Here, we present a short overview over organizational principles of the amygdala in rodents, non-human primates and humans, and review findings on the origin, morphology, and targets of serotonergic innervation, the distribution patterns and cellular expression of serotonin receptors, and the consequences of stress and pharmacological manipulations of serotonergic transmission in the amygdala, focusing particularly on the extensively studied basolateral complex and central nucleus.
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Amígdala del Cerebelo/citología , Ansiedad/patología , Neuronas Serotoninérgicas/citología , Serotonina/metabolismo , Estrés Psicológico/patología , Amígdala del Cerebelo/metabolismo , Animales , Ansiedad/metabolismo , Humanos , Ratones , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/fisiología , Ratas , Neuronas Serotoninérgicas/efectos de los fármacos , Neuronas Serotoninérgicas/metabolismo , Serotoninérgicos/farmacología , Estrés Psicológico/metabolismoRESUMEN
Spinal muscular atrophy with respiratory distress type 1 (SMARD1) is a childhood motoneuron disease caused by mutations in the gene encoding for IGHMBP2, an ATPase/Helicase. Paralysis of the diaphragm is an early and prominent clinical sign resulting both from denervation and myopathy. In skeletal muscles, muscle atrophy mainly results from loss of motoneuron cell bodies and axonal degeneration. Although it is well known that loss of motoneurons at the lumbar spinal cord is an early event in the pathogenesis of the disease, it is not clear whether the corresponding proximal axons and NMJs are also early affected. In order to address this question, we have investigated the time course of the disease progression at the level of the motoneuron cell body, proximal axon (ventral root), distal axon (sciatic nerve), NMJ, and muscle fiber in Nmd(2J) mice, a mouse model for SMARD1. Our results show an early and apparently parallel loss of motoneurons, proximal axons, and NMJs. In affected muscles, however, denervated fibers coexist with NMJs with normal morphology and unaltered neurotransmission. Furthermore, unaffected axons are able to sprout and reinnervate muscle fibers, suggesting selective vulnerability of neurons to Ighmbp2 deficiency. The preservation of the NMJ morphology and neurotransmission in the Nmd(2J) mouse until motor axon loss takes place, differs from that observed in SMA mouse models in which NMJ impairment is an early and more general phenomenon in affected muscles.
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Axones/patología , Neuronas Motoras/patología , Unión Neuromuscular/patología , Atrofias Musculares Espinales de la Infancia/patología , Animales , Proteínas de Unión al ADN/genética , Modelos Animales de Enfermedad , Inmunohistoquímica , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microscopía Confocal , Músculo Esquelético/inervación , Músculo Esquelético/patología , Atrofias Musculares Espinales de la Infancia/genética , Factores de Transcripción/genéticaRESUMEN
While tryptophan hydroxylase-2 (Tph2) null mutant (Tph2(-/-)) mice are completely deficient in brain serotonin (5-HT) synthesis, the formation of serotonergic neurons and pathfinding of their projections are not impaired. However, 5-HT deficiency, during development and in the adult, might affect morphological and functional parameters of other neural systems. To assess the influence of 5-HT deficiency on γ-amino butyric acid (GABA) systems, we carried out measurements of GABA concentrations in limbic brain regions of adult male wildtype (wt), heterozygous (Tph2(+/-)) and Tph2(-/-) mice. In addition, unbiased stereological estimation of GABAergic interneuron numbers and density was performed in subregions of amygdala and hippocampus. Amygdala and prefrontal cortex displayed significantly increased and decreased GABA concentrations, respectively, exclusively in Tph2(+/-) mice while no changes were detected between Tph2(-/-) and wt mice. In contrast, in the hippocampus, increased GABA concentrations were found in Tph2(-/-) mice. While total cell density in the anterior basolateral amygdala did not differ between genotypes, the number and density of the GABAergic interneurons were significantly decreased in Tph2(-/-) mice, with the group of parvalbumin (PV)-immunoreactive (ir) interneurons contributing somewhat less to the decrease than that of non-PV-ir GABAergic interneurons. Major morphological changes were also absent in the dorsal hippocampus, and only a trend toward reduced density of PV-ir cells was observed in the CA3 region of Tph2(-/-) mice. Our findings are the first to document that life-long reduction or complete lack of brain 5-HT transmission causes differential changes of GABA systems in limbic regions which are key players in emotional learning and memory processes. The changes likely reflect a combination of developmental alterations and functional adaptations of emotion circuits to balance the lack of 5-HT, and may underlie altered emotional behavior in 5-HT-deficient mice. Taken together, our findings provide further insight into the mechanisms how life-long 5-HT deficiency impacts the pathogenesis of anxiety- and fear-related disorders.
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Neuronas GABAérgicas/metabolismo , Sistema Límbico/citología , Sistema Límbico/metabolismo , Serotonina/deficiencia , Triptófano Hidroxilasa/deficiencia , Ácido gamma-Aminobutírico/metabolismo , Animales , Neuronas GABAérgicas/química , Neuronas GABAérgicas/citología , Sistema Límbico/química , Masculino , Ratones , Ratones Noqueados , Ácido gamma-Aminobutírico/análisisRESUMEN
Fabry disease is an X-chromosomal recessive deficiency of the lysosomal hydrolase alpha-galactosidase A (alpha-Gal). This results in an accumulation of globotriaosylceramide (GL-3) in a variety of cells often with subsequent functional impairment. Here, the impact of Fabry disease on the biology of circulating angiogenic cells (CACs) and the endothelial response to transient ischemia was investigated. Untreated patients with Fabry disease (n = 26), patients after initiation of alpha-Gal enzyme replacement therapy (ERT) (n = 16) and healthy controls (n = 26) were investigated. Endothelial function was assessed by the EndoPAT2000 device. CAC numbers were assessed by flow-cytometry, CAC function by a modified Boyden chamber assay. Fabry patients showed a pathologic endothelial response, which normalized after ERT. CACs were increased in number, but functionally impaired. Immunofluorescence and electron microscopy identified an accumulation of GL-3 in Fabry CACs. ERT attenuated CAC dysfunction and improved markers of oxidative stress response in Fabry patients via a reduction in GL-3 accumulation in vitro and in vivo. Silencing of alpha-Gal in healthy CACs impaired their migratory capacity underlining a key role of this enzyme for CAC function. CAC supernatant as well as CACs from Fabry patients impaired angiogenesis and migratory capacity of HUVECs providing a mechanistic link between CAC and endothelial dysfunction. CAC adhesion to TNF-α pre-stimulated HUVECs and tube formation was impaired by alpha-Gal knockdown. Fabry patients show a dysfunction of CAC and a pathologic endothelial response. ERT improves CAC and endothelial function and thus may attenuate development of cardiovascular disease in the long term in this patient population.