Microglia-precursor cell interactions in health and in pathology

Until recently, microglia were mainly known as the resident phagocytes of the brain, i.e. the ‘immunological warriors’ of the brain. However, extensive knowledge is being accumulated about the functions of microglia beyond immunity. Nowadays, it is well accepted that microglial cells are highly dynamic and responsive, and that they intervene in a dual manner in many developmental processes that shape the central nervous system, including neurogenesis, gliogenesis, spatial patterning, synaptic formation and elimination, and neural circuit establishment and maturation. The differentiation and the pool of precursor cells were also shown to be under microglia regulation via bidirectional communication. In this concise review, I discuss our recent work in microglia-Pax6+ cell interactions in one of the circumventricular organs, the pineal gland. An analogy with the rest of the central nervous system is also presented. In addition, I briefly examine mechanisms of interaction between microglia and non-microglial cells in both health and disease. New avenues are also introduced, which may lead us to better comprehend the impact of microglia in physiological and pathological conditions.Fil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Microglia in Circumventricular Organs: The Pineal Gland Example

The circumventricular organs (CVOs) are unique areas within the central nervous system. They serve as a portal for the rest of the body and, as such, lack a blood-brain barrier. Microglia are the primary resident immune cells of the brain parenchyma. Within the CVOs, microglial cells find themselves continuously challenged and stimulated by local and systemic stimuli, even under steady-state conditions. Therefore, CVO microglia in their typical state often resemble the activated microglial forms found elsewhere in the brain as they are responding to pathological conditions or other stressors. In this review, I focus on the dynamics of CVO microglia, using the pineal gland as a specific CVO example. Data related to microglia heterogeneity in both homeostatic and unhealthy environments are presented and discussed, including those recently generated by using advanced single-cell and single-nucleus technology. Finally, perspectives in the CVO microglia field are also included. Summary Statement Microglia in circumventricular organs (CVOs) continuously adapt to react differentially to the diverse challenges they face. Herein, I discuss microglia heterogeneity in CVOs, including pineal gland. Further studies are needed to better understand microglia dynamics in these unique brain areas.Fil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Thinking with rhythm about the benevolent side of the innate immunity

The immune system is responsible for surveillance and monitoring, and for reacting against pathogens and insults. The central nervous system (CNS) is a highly protected area that utilizes its own immune resources, due in part to anatomical barriers. These are the microglia, which are the resident phagocytes of the CNS. Microglia are highly dynamic cells in both shape and phenotype. They originate from myeloid precursors in the yolk sac that colonize the developing CNS. Beyond their defensive role, microglia participate in many processes that shape and maintain the CNS, and also implement its plasticity. One question that emerges is whether microglia adjust their functions based on their own innate temporal capacity, or whether they merely react to external temporal cues. Our group has been working with the pineal gland, as a circadian model within the CNS, in order to better understand microglia phenotypes in both ontogeny and daily cycles.Fil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaXXXVI Scientific Meeting of the Cuyo Biology SocietyMendozaArgentinaSociedad de Biología de Cuy

Cellular basis of pineal gland development: Emerging role of microglia as phenotype regulator

The adult pineal gland is composed of pinealocytes, astrocytes, microglia, and other interstitial cells that have been described in detail. However, factors that contribute to pineal development have not been fully elucidated, nor have pineal cell lineages been well characterized. We applied systematic double, triple and quadruple labeling of cell-specific markers on prenatal, postnatal and mature rat pineal gland tissue combined with confocal microscopy to provide a comprehensive view of the cellular dynamics and cell lineages that contribute to pineal gland development. The pineal gland begins as an evagination of neuroepithelium in the roof of the third ventricle. The pineal primordium initially consists of radially aligned Pax6+ precursor cells that express vimentin and divide at the ventricular lumen. After the tubular neuroepithelium fuses, the distribution of Pax6+ cells transitions to include rosette-like structures and later, dispersed cells. In the developing gland all dividing cells express Pax6, indicating that Pax6+ precursor cells generate pinealocytes and some interstitial cells. The density of Pax6+ cells decreases across pineal development as a result of cellular differentiation and microglial phagocytosis, but Pax6+ cells remain in the adult gland as a distinct population. Microglial colonization begins after pineal recess formation. Microglial phagocytosis of Pax6+ cells is not common at early stages but increases as microglia colonize the gland. In the postnatal gland microglia affiliate with Tuj1+ nerve fibers, IB4+ blood vessels, and Pax6+ cells. We demonstrate that microglia engulf Pax6+ cells, nerve fibers, and blood vessel-related elements, but not pinealocytes. We conclude that microglia play a role in pineal gland formation and homeostasis by regulating the precursor cell population, remodeling blood vessels and pruning sympathetic nerve fibers.Fil: Ibañez Rodriguez, María Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Noctor, Stephen C.. University of California; Estados UnidosFil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Transcription Regulation—Brain Development and Homeostasis—A Finely Tuned and Orchestrated Scenario in Physiology and Pathology

A finely tuned regulation of gene expression is essential for shaping the nervous system and for maintaining its homeostasis throughout life. Disruptions in gene regulation can impact brain development and physiology in ways that contribute to diverse pathologies. Classic and state-of-the art experimental models and technologies have advanced our knowledge of transcriptional regulators and the ways they interact in the healthy and diseased brain. Further in-depth characterization of the mechanisms of transcriptional regulation is needed to better understand how each element, from genes to cells, defines and maintains identities and functionalities in the nervous system. This Research Topic focuses on transcriptional regulation within the nervous system, with an emphasis on developmental and homeostatic processes, their dysregulation, and their association with neurodevelopmental disorders and neurodegenerative diseases. Eleven peer-reviewed manuscripts including six original articles, three reviews, one mini review, and one brief research report, encompass this special volume. Fifty-nine authors from research laboratories located in 10 countries: Argentina, Canada, China, Germany, Israel, Russia, Serbia, United Kingdom, United States, and Vietnam, took part in this initiative.Fil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Silva Junqueira de Souza, Flavio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Rath, Martin F.. Universidad de Copenhagen; DinamarcaFil: Martínez Cerdeño, Verónica. University of California at Davis; Estados Unido

Spatio-temporal dynamics of nuclear CREB1: what does it mean?

In the mammalian pineal gland (PG), cyclic AMP responsive element-binding protein 1 (CREB1) participates in the nocturnal melatonin synthesis that rhythmically modulates physiology and behavior. Phosphorylation of CREB1 present in pinealocyte nuclei is one of the key regulatory steps that drives pineal transcription. The spatio-temporal dynamics of CREB1 itself within PG cell types have not yet been documented. In this study we analyzed total CREB1 via Western blot, and the dynamism of CREB1 nuclear distribution in individual rat pinealocytes using fluorescence immunohistochemistry followed by confocal laser-scanning microscopy and quantitative analysis. Total CREB1 levels remained constant in the PG throughout the light:dark cycle. The distribution pattern of nuclear CREB1 did vary, however, among different PG cells. Pinealocytes emerged as having discrete CREB1 domains within their nucleoplasm that were especially distinct. The number, size, and location of CREB1 foci fluctuated among pinealocytes, within the same PG and among Zeitgeber times. A significantly larger dispersion of CREB1-immunoreactive nuclear sites was found at night. This was not accompanied by changes in the overall transcription activity, which was mostly conserved between the light and dark phases, as shown by the expression of a particular phosphorylated form of the RNA polymerase II (RNAPII-pSer5CTD). Suppression of the nocturnal norepinephrine pulse by chronic bilateral superior cervical ganglionectomy increased CREB1 dispersion in pinealocyte nuclei, as compared to sham-derived cells. In addition, differences in CREB1 distribution were found between sham-operated and non-operated rats at early night. Together, these data suggest that in mature pinealocytes nuclear CREB1 is subjected to a dynamic spatio-temporal distribution. Further studies are necessary to elucidate the underlying mechanisms, including the role of chromatin and interchromatin elements, and to understand the impact of CREB1 reorganization in the pineal transcriptome.Fil: Farías Altamirano, Luz Estefanía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Vásquez, Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Freites, Carlos Leandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Ibañez, Jorge Ernesto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Guido, Mario Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Reperfusion arrhythmias increase after superior cervical ganglionectomy due to conduction disorders and changes in repolarization

Pharmacological concentrations of melatonin reduce reperfusion arrhythmias, but less is known about the antiarrhythmic protection of the physiological circadian rhythm of melatonin. Bilateral surgical removal of the superior cervical ganglia irreversibly suppresses melatonin rhythmicity. This study aimed to analyze the cardiac electrophysiological effects of the loss of melatonin circadian oscillation and the role played by myocardial melatonin membrane receptors, SERCA2A, TNFα, nitrotyrosine, TGFβ, KATP channels, and connexin 43. Three weeks after bilateral removal of the superior cervical ganglia or sham surgery, the hearts were isolated and submitted to ten minutes of regional ischemia followed by ten minutes of reperfusion. Arrhythmias, mainly ventricular tachycardia, increased during reperfusion in the ganglionectomy group. These hearts also suffered an epicardial electrical activation delay that increased during ischemia, action potential alternants, triggered activity, and dispersion of action potential duration. Hearts from ganglionectomized rats showed a reduction of the cardioprotective MT2 receptors, the MT1 receptors, and SERCA2A. Markers of nitroxidative stress (nitrotyrosine), inflammation (TNFα), and fibrosis (TGFβ and vimentin) did not change between groups. Connexin 43 lateralization and the pore-forming subunit (Kir6.1) of KATP channels increased in the experimental group. We conclude that the loss of the circadian rhythm of melatonin predisposes the heart to suffer cardiac arrhythmias, mainly ventricular tachycardia, due to conduction disorders and changes in repolarization.Fil: Prado, Natalia Jorgelina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentina. Universidad Nacional de Cuyo; ArgentinaFil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Farías Altamirano, Luz Estefanía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Aguiar, Francisco. Universidad Nacional de Cuyo; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Ponce Zumino, Amira Zulma. Universidad Nacional de Cuyo; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ruiz Sánchez, Francisco Javier. Universidad Nacional de Cuyo; ArgentinaFil: Miatello, Roberto Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentina. Universidad Nacional de Cuyo; ArgentinaFil: Pueyo, Esther. Universidad de Zaragoza; España. Centro de Investigación Biomédica en Red en el área temática de Bioingeniería, Biomateriales y Nanomedicina; EspañaFil: Diez, Emiliano Raúl. Universidad Nacional de Cuyo; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

Dynamics of GABABR and associated proteins in the postnatal rat cerebellum

Before glutamatergic synapses are formed, GABA-mediated signaling is considered to drive cell differentiation in the developing central nervous system (CNS). GABA, a classical inhibitory neurotransmitter, can also depolarize immature cells. Although this shift is mediated by the ionotropic GABA A receptor (GABAAR), recent evidence suggests that the electrical properties of GABAARs can be modulated by the metabotropic GABA B receptor (GABABR). GABABRs are macromolecular complexes, formed by a G protein-coupled receptor and a large number of constituents that interact together and ultimately influence cell identity and behavior. The composition of these complexes exhibits wide spatiotemporal variations; however, the implications of such dynamism during development of the CNS are far from being understood. We have determined total protein expression of some constituents of GABABRs (GABABR1a; GABABR1b, GABABR2; KCTD12) in the developing cerebellum of postnatal rats at 5, 15 and 90 days after birth, by performing Western Blots. Our findings suggest that the expression levels of the core and auxiliary subunits of GABABRs vary ontogenetically. This dynamism was also observed at the mRNA levels by RT-PCR. In addition, multiple immunolabeling followed by confocal microscopy of cerebellar sections showed Purkinje cells as the most dynamic cell type in terms of subcellular localization of the different molecules studied here. Our data support a cell lineage-dependent GABABR regulation.Fil: Vásquez, Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaXXXIII Congress of the Argentine Society for Research in NeuroscienceCórdobaArgentinaSociedad Argentina de Investigación en Neurociencia

Dynamics of GABABR signaling: influence of cholesterol and aging

GABA B receptors (GABABRs) are obligatory heterodimers which belong to the superfamily of G protein-coupled receptors (GPCRs). Age-related changes in membrane cholesterol levels modulate membrane fluidity, which in-turn influences GPCRs´ localization and function. We studied the GABABR and also a transmembrane transporter structurally homologous to KCC2. To characterize transient conformational changes over time, molecular dynamics simulations were performed using a neuronal plasma membrane (PM) model. Two different membrane cholesterol levels were evaluated: 45% and 10%, which intend to resemble the composition of adult and aged neuronal PMs, respectively. For experimental verification in both young and aged cerebella, we determined protein expression and distribution, and we assessed whether the two proteins interact with each other in vivo. Techniques were: western blots (WB), co-immunoprecipitation assays, and multiple immunolabeling followed by confocal microscopy. Our results suggest that the expression and spatial distribution of both proteins change as the cerebellum grows older. Based on our in silico analyses, we infer that a G protein-independent interaction does occur. Also, we confirmed that the two proteins are part of the same complex in the cerebellum. As our simulations indicate, we propose that the underlying mechanism implies transient conformational changes, which are highly dependent on cholesterol levels and are therefore affected by the aging process.Fil: Vásquez, Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Masone, Diego Fernando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaXXXIV Annual Meeting of Argentine Society for Research in NeurosciencesVilla Carlos PazArgentinaArgentine Society for Research in Neuroscience

Surgical Techniques and Nuances for Superior Cervical Ganglionectomy and Decentralization in Rats

The sympathetic nervous system has been implicated in various physiological and pathological processes, including regulation of homeostatic functions, maintenance of the circadian rhythms, and neuronal disruption and recovery after injury. Of special interest is focus on the role of the superior cervical ganglion (SCG) in regulating the daily changes in pineal function. Removal of the superior cervical ganglion (SCGx) and decentralization have served as valuable microsurgical models to investigate the effects of surgical denervation on this gland or organ. In this chapter, we offer information about methodologies for performing SCGx along with decentralization and denervation procedures, including details about recommended equipment as well as tips that can improve these techniques.Fil: Madhani, Sarosh Irfan. University of California; Estados UnidosFil: David C. Klein. National Institutes of Health; Estados UnidosFil: Muñoz, Estela Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Savastano, Luis E.. University of California; Estados Unido