RESUMEN
The vagal ganglia, comprised of the superior (jugular) and inferior (nodose) ganglia of the vagus nerve, receive somatosensory information from the head and neck or viscerosensory information from the inner organs, respectively. Developmentally, the cranial neural crest gives rise to all vagal glial cells and to neurons of the jugular ganglia, while the epibranchial placode gives rise to neurons of the nodose ganglia. Crest-derived nodose glial progenitors can additionally generate autonomic neurons in the peripheral nervous system, but how these progenitors generate neurons is unknown. Here, we found that some Sox10+ neural crest-derived cells in, and surrounding, the nodose ganglion transiently expressed Phox2b, a master regulator of autonomic nervous system development, during early embryonic life. Our genetic lineage-tracing analysis in mice of either sex revealed that despite their common developmental origin and extreme spatial proximity, a substantial proportion of glial cells in the nodose, but not in the neighboring jugular ganglia, have a history of Phox2b expression. We used single-cell RNA-sequencing to demonstrate that these progenitors give rise to all major glial subtypes in the nodose ganglia, including Schwann cells, satellite glia, and glial precursors, and mapped their spatial distribution by in situ hybridization. Lastly, integration analysis revealed transcriptomic similarities between nodose and dorsal root ganglia glial subtypes and revealed immature nodose glial subtypes. Our work demonstrates that these crest-derived nodose glial progenitors transiently express Phox2b, give rise to the entire complement of nodose glial cells, and display a transcriptional program that may underlie their bipotent nature.
Asunto(s)
Proteínas de Homeodominio , Cresta Neural , Neuroglía , Ganglio Nudoso , Factores de Transcripción , Animales , Ganglio Nudoso/citología , Ganglio Nudoso/metabolismo , Ratones , Neuroglía/metabolismo , Neuroglía/citología , Proteínas de Homeodominio/metabolismo , Proteínas de Homeodominio/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Cresta Neural/citología , Cresta Neural/metabolismo , Femenino , Masculino , Células-Madre Neurales/metabolismo , Células-Madre Neurales/citología , Ratones Endogámicos C57BLRESUMEN
Mutations in the transcription factors encoded by PHOX2B or LBX1 correlate with congenital central hypoventilation disorders. These conditions are typically characterized by pronounced hypoventilation, central apnea, and diminished chemoreflexes, particularly to abnormally high levels of arterial PCO2. The dysfunctional neurons causing these respiratory disorders are largely unknown. Here, we show that distinct, and previously undescribed, sets of medullary neurons coexpressing both transcription factors (dB2 neurons) account for specific respiratory functions and phenotypes seen in congenital hypoventilation. By combining intersectional chemogenetics, intersectional labeling, lineage tracing, and conditional mutagenesis, we uncovered subgroups of dB2 neurons with key functions in (i) respiratory tidal volumes, (ii) the hypercarbic reflex, (iii) neonatal respiratory stability, and (iv) neonatal survival. These data provide functional evidence for the critical role of distinct medullary dB2 neurons in neonatal respiratory physiology. In summary, our work identifies distinct subgroups of dB2 neurons regulating breathing homeostasis, dysfunction of which causes respiratory phenotypes associated with congenital hypoventilation.
Asunto(s)
Proteínas de Homeodominio , Hipoventilación , Bulbo Raquídeo , Neuronas , Factores de Transcripción , Hipoventilación/congénito , Hipoventilación/genética , Animales , Neuronas/metabolismo , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Ratones , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Bulbo Raquídeo/metabolismo , Apnea Central del Sueño/genética , Fenotipo , HumanosRESUMEN
The type I transmembrane protein BT-IgSF is predominantly localized in the brain and testes. It belongs to the CAR subgroup of Ig cell adhesion proteins, that are hypothesized to regulate connexin expression or localization. Here, we studied the putative link between BT-IgSF and connexins in astrocytes, ependymal cells and neurons of the mouse. Global knockout of BT-IgSF caused an increase in the clustering of connexin43 (Gja1), but not of connexin30 (Gjb6), on astrocytes and ependymal cells. Additionally, knockout animals displayed reduced expression levels of connexin43 protein in the cortex and hippocampus. Importantly, analysis of biocytin spread in hippocampal or cortical slices from mature mice of either sex revealed a decrease in astrocytic cell-cell coupling in the absence of BT-IgSF. Blocking either protein biosynthesis or proteolysis showed that the lysosomal pathway increased connexin43 degradation in astrocytes. Localization of connexin43 in subcellular compartments was not impaired in astrocytes of BT-IgSF mutants. In contrast to connexin43 the localization and expression of connexin36 (Gjd2) on neurons was not affected by the absence of BT-IgSF. Overall, our data indicate that the IgCAM BT-IgSF is essential for correct gap junction-mediated astrocyte-to-astrocyte cell communication.Significance Statement Astrocytes regulate a variety of physiological processes in the developing and adult brain that are essential for proper brain function. Astrocytes form extensive networks in the brain and communicate via gap junctions. Disruptions of gap junction coupling are found in several diseases such as neurodegeneration or epilepsy. Here, we demonstrate that the cell adhesion protein BT-IgSF is essential for gap junction mediated coupling between astrocytes in the cortex and hippocampus.
RESUMEN
Familial hemophagocytic lymphohistiocytosis (FHL) is an inherited, often fatal immune deficiency characterized by severe systemic hyperinflammation. Although allogeneic bone marrow transplantation can be curative, more effective therapies are urgently needed. FHL is caused by inactivating mutations in proteins that regulate cellular immunity. Here, we used an adeno-associated virus-based CRISPR-Cas9 system with an inhibitor of nonhomologous end joining to repair such mutations in potentially long-lived T cells ex vivo. Repaired CD8 memory T cells efficiently cured lethal hyperinflammation in a mouse model of Epstein-Barr virus-triggered FHL2, a subtype caused by perforin-1 (Prf1) deficiency. Furthermore, repair of PRF1 and Munc13-4 (UNC13D)-whose deficiency causes the FHL subtype FHL3-in mutant memory T cells from two critically ill patients with FHL restored T cell cytotoxicity. These results provide a starting point for the treatment of genetic T cell immune dysregulation syndromes with repaired autologous T cells.
Asunto(s)
Infecciones por Virus de Epstein-Barr , Linfohistiocitosis Hemofagocítica , Animales , Ratones , Humanos , Linfohistiocitosis Hemofagocítica/genética , Linfohistiocitosis Hemofagocítica/terapia , Sistemas CRISPR-Cas , Infecciones por Virus de Epstein-Barr/genética , Infecciones por Virus de Epstein-Barr/terapia , Células T de Memoria , Herpesvirus Humano 4 , Proteínas de la Membrana/genéticaRESUMEN
Vagal sensory neurons monitor mechanical and chemical stimuli in the gastrointestinal tract. Major efforts are underway to assign physiological functions to the many distinct subtypes of vagal sensory neurons. Here, we use genetically guided anatomical tracing, optogenetics, and electrophysiology to identify and characterize vagal sensory neuron subtypes expressing Prox2 and Runx3 in mice. We show that three of these neuronal subtypes innervate the esophagus and stomach in regionalized patterns, where they form intraganglionic laminar endings. Electrophysiological analysis revealed that they are low-threshold mechanoreceptors but possess different adaptation properties. Lastly, genetic ablation of Prox2 and Runx3 neurons demonstrated their essential roles for esophageal peristalsis in freely behaving mice. Our work defines the identity and function of the vagal neurons that provide mechanosensory feedback from the esophagus to the brain and could lead to better understanding and treatment of esophageal motility disorders.
Asunto(s)
Subunidad alfa 3 del Factor de Unión al Sitio Principal , Esófago , Motilidad Gastrointestinal , Proteínas de Homeodominio , Células Receptoras Sensoriales , Nervio Vago , Animales , Ratones , Subunidad alfa 3 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 3 del Factor de Unión al Sitio Principal/metabolismo , Esófago/inervación , Esófago/metabolismo , Esófago/fisiología , Motilidad Gastrointestinal/genética , Motilidad Gastrointestinal/fisiología , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Mecanorreceptores/fisiología , Neuronas Aferentes/fisiología , Células Receptoras Sensoriales/metabolismo , Células Receptoras Sensoriales/fisiología , Estómago/inervación , Estómago/metabolismo , Estómago/fisiología , Nervio Vago/fisiologíaRESUMEN
Breathing (or respiration) is an unconscious and complex motor behavior which neuronal drive emerges from the brainstem. In simplistic terms, respiratory motor activity comprises two phases, inspiration (uptake of oxygen, O2) and expiration (release of carbon dioxide, CO2). Breathing is not rigid, but instead highly adaptable to external and internal physiological demands of the organism. The neurons that generate, monitor, and adjust breathing patterns locate to two major brainstem structures, the pons and medulla oblongata. Extensive research over the last three decades has begun to identify the developmental origins of most brainstem neurons that control different aspects of breathing. This research has also elucidated the transcriptional control that secures the specification of brainstem respiratory neurons. In this review, we aim to summarize our current knowledge on the transcriptional regulation that operates during the specification of respiratory neurons, and we will highlight the cell lineages that contribute to the central respiratory circuit. Lastly, we will discuss on genetic disturbances altering transcription factor regulation and their impact in hypoventilation disorders in humans.
RESUMEN
The mature cerebellum controls motor skill precision and participates in other sophisticated brain functions that include learning, cognition, and speech. Different types of GABAergic and glutamatergic cerebellar neurons originate in temporal order from two progenitor niches, the ventricular zone and rhombic lip, which express the transcription factors Ptf1a and Atoh1, respectively. However, the molecular machinery required to specify the distinct neuronal types emanating from these progenitor zones is still unclear. Here, we uncover the transcription factor Olig3 as a major determinant in generating the earliest neuronal derivatives emanating from both progenitor zones in mice. In the rhombic lip, Olig3 regulates progenitor cell proliferation. In the ventricular zone, Olig3 safeguards Purkinje cell specification by curtailing the expression of Pax2, a transcription factor that suppresses the Purkinje cell differentiation program. Our work thus defines Olig3 as a key factor in early cerebellar development.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Cerebelo/embriología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Diferenciación Celular , Cerebelo/citología , Cerebelo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Ratones , Ratones Noqueados , Neurogénesis , Células de Purkinje/fisiología , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Syncytial skeletal muscle cells contain hundreds of nuclei in a shared cytoplasm. We investigated nuclear heterogeneity and transcriptional dynamics in the uninjured and regenerating muscle using single-nucleus RNA-sequencing (snRNAseq) of isolated nuclei from muscle fibers. This revealed distinct nuclear subtypes unrelated to fiber type diversity, previously unknown subtypes as well as the expected ones at the neuromuscular and myotendinous junctions. In fibers of the Mdx dystrophy mouse model, distinct subtypes emerged, among them nuclei expressing a repair signature that were also abundant in the muscle of dystrophy patients, and a nuclear population associated with necrotic fibers. Finally, modifications of our approach revealed the compartmentalization in the rare and specialized muscle spindle. Our data identifies nuclear compartments of the myofiber and defines a molecular roadmap for their functional analyses; the data can be freely explored on the MyoExplorer server ( https://shiny.mdc-berlin.de/MyoExplorer/ ).
Asunto(s)
Núcleo Celular/genética , Núcleo Celular/metabolismo , Músculo Esquelético/crecimiento & desarrollo , Músculo Esquelético/metabolismo , Transcriptoma , Animales , Línea Celular , Citoplasma , Modelos Animales de Enfermedad , Regulación del Desarrollo de la Expresión Génica , Heterogeneidad Genética , Humanos , Ratones , Ratones Endogámicos mdx , Fibras Musculares Esqueléticas , Músculo Esquelético/citología , Distrofias Musculares/genética , Distrofias Musculares/metabolismo , RNA-Seq , Regeneración , TendonesRESUMEN
Exposure to prolonged, uncontrollable stress reduces reward-seeking behavior, resulting in anhedonia in neuropsychiatric disorders, such as posttraumatic stress disorder. However, it is unclear to what degree stressed subjects lose interest in rewards themselves or in reward-related cues that instigate reward-seeking behavior. In the present study, we investigated the effects of single prolonged stress (SPS) on cue-directed behavior in two different procedures: Pavlovian conditioned approach (PCA) and cue-induced reinstatement of cocaine-seeking. In Experiment 1, rats were exposed to SPS and tested for the acquisition of sign-tracking (cue-directed) and goal-tracking (reward-directed) behaviors during a PCA procedure. In Experiment 2, rats were exposed to SPS and tested for the expression of sign- and goal-tracking as well as cue-induced reinstatement of cocaine-seeking. Because dopaminergic activity in the nucleus accumbens is known to play a central role in many cue-directed behaviors, including both sign-tracking and cue-induced reinstatement, Experiment 3 used in vivo microdialysis to measure the effect of SPS on baseline and evoked dopamine levels in the nucleus accumbens. SPS decreased sign-tracking and increased goal-tracking during the acquisition of PCA behavior without affecting reward consumption. In addition, SPS decreased cue-induced reinstatement without affecting cocaine self-administration. Finally, SPS decreased evoked but not baseline levels of dopamine in the nucleus accumbens. These results suggest that SPS decreases the motivational, but not consummatory, aspects of reward-seeking behavior, which may result from long-term, SPS-induced reductions in dopamine release in the nucleus accumbens.
Asunto(s)
Cocaína/administración & dosificación , Condicionamiento Clásico/fisiología , Señales (Psicología) , Inhibidores de Captación de Dopamina/administración & dosificación , Comportamiento de Búsqueda de Drogas/efectos de los fármacos , Refuerzo en Psicología , Estrés Psicológico/fisiopatología , Animales , Condicionamiento Clásico/efectos de los fármacos , Comportamiento de Búsqueda de Drogas/fisiología , Masculino , Ratas , Ratas Sprague-Dawley , Autoadministración , Factores de TiempoRESUMEN
Although splicing is widespread and evolves rapidly among species, the mechanisms driving this evolution, as well as its functional implications, are not yet fully understood. We analyzed the evolution of splicing patterns based on transcriptome data from five tissues of humans, chimpanzees, rhesus macaques and mice. In total, 1526 exons and exon sets from 1236 genes showed significant splicing differences among primates. More than 60% of these differences represent constitutive-to-alternative exon transitions while an additional 25% represent changes in exon inclusion frequency. These two dominant evolutionary patterns have contrasting conservation, regulation and functional features. The sum of these features indicates that, despite their prevalence, constitutive-to-alternative exon transitions do not substantially contribute to long-term functional transcriptome changes. Conversely, changes in exon inclusion frequency appear to be functionally relevant, especially for changes taking place in the brain on the human evolutionary lineage.