Role of Glia in Sculpting Synaptic Connections at the Drosophila Neuromuscular Junction: A Dissertation

Emerging evidence in both vertebrates and invertebrates is redefining glia as active players in the development and integrity of the nervous system. The formation of functional neuronal circuits requires the precise addition of new synapses. Mounting evidence implicates glial function in synapse remodeling and formation. However, the precise molecular mechanisms governing these functions are poorly understood. My thesis work begins to define the molecular mechanisms by which glia communicate with neurons at the Drosophila neuromuscular junction (NMJ). During development glia play a critical role in remodeling neuronal circuits in the CNS. In order to understand how glia remodel synapses, I manipulated a key component of the glial engulfment machinery, Draper. I found that during normal NMJ growth presynaptic boutons constantly shed membranes or debris. However, a loss of Draper resulted in an accumulation of debris and ghost boutons, which inhibited synaptic growth. I found that glia use the Draper pathway to engulf these excess membranes to sculpt synapses. Surprisingly, I found that muscle cells function as phagocytic cells as well by eliminating immature synaptic ghost boutons. This demonstrates that the combined efforts of glia and muscle are required for the addition of synapses and proper growth. My work establishes that glia play a crucial role in synapse development at the NMJ and suggests that there are other glial-derived molecules that regulate synapse function. I identified one glial derived molecule critical for the development of the NMJ, a TGF-β ligand called Maverick. Presynaptically, Maverick regulates the activation of BMP pathway confirmed by reducing the transcription of the known target gene Trio. Postsynaptically, it regulates the transcription of Glass bottom boat (Gbb) in the muscle suggesting that glia modulate the function of Gbb and consequently the activation of the BMP retrograde pathway at NMJ. Surprisingly, I also found that glial Maverick regulates the transcription of Shaker potassium channel, suggesting that glia potentially could regulate muscle excitability and consequently modulate synaptic transmission. Future work will elucidate such hypothesis. My work has demonstrated two novel roles for glia at the NMJ. First is that glia engulfing activity is important for proper synaptic growth. Second is that the secretion of glial-derived molecules are required to orchestrate synaptic development. This further supports that glia are critical active players in maintaining a functional nervous system

Integration of a Retrograde Signal during Synapse Formation by Glia-Secreted TGF-β Ligand

SummaryGlial cells are crucial regulators of synapse formation, elimination, and plasticity [1, 2]. In vitro studies have begun to identify glial-derived synaptogenic factors [1], but neuron-glia signaling events during synapse formation in vivo remain poorly defined. The coordinated development of pre- and postsynaptic compartments at the Drosophila neuromuscular junction (NMJ) depends on a muscle-secreted retrograde signal, the TGF-β/BMP Glass bottom boat (Gbb) [3, 4]. Muscle-derived Gbb activates the TGF-β receptors Wishful thinking (Wit) and either Saxophone (Sax) or Thick veins (Tkv) in motor neurons [3, 4]. This induces phosphorylation of Mad (P-Mad) in motor neurons, its translocation into the nucleus with a co-Smad, and activation of transcriptional programs controlling presynaptic bouton growth [5]. Here we show that NMJ glia release the TGF-β ligand Maverick (Mav), which likely activates the muscle activin-type receptor Punt to potently modulate Gbb-dependent retrograde signaling and synaptic growth. Loss of glial Mav results in strikingly reduced P-Mad at NMJs, decreased Gbb transcription in muscle, and in turn reduced muscle-to-motor neuron retrograde TGF-β/BMP signaling. We propose that by controlling Gbb release from muscle, glial cells fine tune the ability of motor neurons to extend new synaptic boutons in correlation to muscle growth. Our work identifies a novel glia-derived synaptogenic factor by which glia modulate synapse formation in vivo

Glia and Muscle Sculpt Neuromuscular Arbors by Engulfing Destabilized Synaptic Boutons and Shed Presynaptic Debris

As synapses grow at the Drosophila neuromuscular junction, they shed membrane material in an activity-dependent manner. Glia and postsynaptic muscle cells are required to engulf this debris to ensure new synaptic growth

Composición, propiedades, estabilidad y comportamiento térmico del aceite de semilla de tamarindo (Tamarindus indica)

The composition, thermal stability and phase behavior of tamarind (Tamarindus indica) seed oil were analyzed to contribute to the exploration of their potential uses. The oil was extracted from the kernel of the tamarind seed with hexane, and its main physical, chemical and thermal properties were analyzed by infrared spectrometry, gas chromatography-mass spectrometry, DSC, and TGA. The results showed that the tamarind seed had a 3.76 ± 0.20% oil with a saponification index of 174.80 ± 9.87 mg KOH/g and the major fatty acids were lignoceric (20.15%), oleic (18.99%) and palmitic (11.99%). Stearic, behenic, linoleic, arachidic, and other fatty acids were also present. TGA and DSC showed that in an inert atmosphere, the triacylglycerols of tamarind seed oil (TSO) are decomposed in a single stage that starts at 224.1 °C and in an air atmosphere in three stages, initiating its decomposition at 218 °C. The TSO showed crystallization and fusion curves with a single maximum peak with Tonset and Toffset of 20.16 and ?38.8 °C and ?22.2 and 28.6 °C, respectively. The solid fat profile of the oil showed a semi-solid and liquid consistency in the ambient temperature range. The composition, thermal and phase behavior showed that TSO is potentially useful for alimentary, pharmacological, and cosmetological purposes.La composición, estabilidad y comportamiento térmico del aceite de semilla de tamarindo (Tamarindus indica) fueron analizadas con el fin de contribuir al conocimiento de sus potenciales usos. El aceite fue extraído del núcleo de la semilla con hexano y analizado mediante sus principales propiedades físicas, químicas y térmicas mediante espectrometría infrarroja, cromatografía de gases, espectroscopia de masas, calorimetría (DSC) y termogravimetría (TGA). Los resultados mostraron que las semillas del tamarindo tuvieron un contenido de aceite de 3,76 ± 0,20%, con un índice de saponificación de 174,80 ± 9,87mg KOH/g y ácidos grasos mayoritarios: Lignocérico (20,15%), oleico (18,99%), palmítico (11,99%) y en cantidades menores los ácidos esteárico, behénico, linoleico y araquídico, entre otros. El análisis mediante TGA y DSC mostró que la temperatura inicial de descomposición del aceite fue de 224,1 °C en una sola etapa en atmósfera inerte y en atmósfera de aire fue a 218 °C en tres etapas. El aceite mostró curvas de cristalización y fusión con un solo máximo, iniciándose y finalizando estos cambios de fase a 20,16 y -38,8 °C, and -22,2 y 28,6 °C, respectivamente. Estas propiedades mostraron que el aceite de la semilla de tamarindo tiene potenciales aplicaciones en alimentos y productos farmacológicos y cosméticos

Caracterización fisicoquímica y comportamiento térmico del aceite de “almendra” de guanábana (Annona muricata, L)

In this work some physicochemical properties and the thermal behavior and stability of sour sop or guanabana (Annona muricata) seed “almond” oil were studied by means of chemical, DSC and TG analysis. The results showed that the almond has 2.5% ash, 17.9% crude fiber, 15.7% protein, 26.0% de carbohydrates and 37.7% oil (dry base). The composition of almond oil showed 68.5% unsaturated fatty acids, mainly oleic and linoleic, and some palmitoleic acids, and 31.5% saturated, principally palmitic and stearic fatty acids; refraction index was 1.468 and saponification and iodine value were 168.2 and 87.0, respectively. DSC thermal analysis showed that oil crystallization initiates at -4.5 °C and ends at -79.0 °C with a crystallization enthalpy of 48.2 J/g; the oil melts in a temperature range from -42.4 to +16.9 °C, with a maximum peak at -15 °C and a fusion enthalpy of 80.5 J/g. The oil remained liquid at refrigeration temperatures with minimal SFC and free of crystals at temperatures over 10 °C. TG analysis showed that the thermal decomposition of the oil in a N2 atmosphere starts at 380 °C and ends at 442 °C, with a maximum decomposition rate at 412 °C. Under oxidizing conditions its decomposition begins at 206 °C and concludes at 567 °C. In accordance with this study, sour sop almond seed contains large amounts of an oil that possesses similar characteristics to those of salad and cooking oils.En esta investigación se estudiaron las propiedades físicoquímicas y el comportamiento térmico, mediante calorimetría diferencial de barrido y termogravimetría, del aceite extraído de las “almendras” de las semillas de guanábana (Annona muricata, L). Los resultados mostraron que las almendras de las semillas de guanábana contienen 2.5% de cenizas, 17.9% de fibra cruda, 15.7% de proteínas, 26.0% de carbohidratos y 37.7% de aceite (base seca). El aceite de las almendras de guanábana mostró una composición con predominio de ácidos grasos insaturados (68.5%) mayoritariamente oleico y linoleico y menores cantidades de palmitoleico y linoleico, principalmente; los ácidos grasos saturados fueron principalmente palmítico y esteárico (31.5%), el índice de refracción fue de 1.468, el valor de saponificación y de yodo fueron de 168.2 y 87.0 respectivamente. El análisis térmico mostró que este aceite inicia su cristalización a -4.5 °C y termina a los -79.0 °C con una entalpía de cristalización de 48.2 J/g y funde en un intervalo que va de -42.4 a 16.9 °C con un máximo de fusión a los -15.4 °C y una entalpía de fusión de 80.5 J/g. El contenido de grasa sólida (SFC) fue mínimo a temperaturas de refrigeración, manteniéndose líquido y libre de cristales a temperaturas superiores a los 10 °C. El análisis termogravimétrico mostró que la descomposición térmica del aceite en atmósfera inerte se inicia a los 380 °C y termina a los 442 °C con un valor máximo en la velocidad de descomposición a los 412 °C. En atmósfera oxidante el aceite inicia su descomposición a los 206 °C y concluye a 567 °C. De acuerdo con las características estudiadas las almendras de las semillas de guanábana tiene un alto contenido de aceite y éste posee características propias de los aceites de mesa

Glial wingless/Wnt regulates glutamate receptor clustering and synaptic physiology at the Drosophila neuromuscular junction

Glial cells are emerging as important regulators of synapse formation, maturation, and plasticity through the release of secreted signaling molecules. Here we use chromatin immunoprecipitation along with Drosophila genomic tiling arrays to define potential targets of the glial transcription factor Reversed polarity (Repo). Unexpectedly, we identified wingless (wg), a secreted morphogen that regulates synaptic growth at the Drosophila larval neuromuscular junction (NMJ), as a potential Repo target gene. We demonstrate that Repo regulates wg expression in vivo and that local glial cells secrete Wg at the NMJ to regulate glutamate receptor clustering and synaptic function. This work identifies Wg as a novel in vivo glial-secreted factor that specifically modulates assembly of the postsynaptic signaling machinery at the Drosophila NMJ

Selective Remodeling: Refining Neural Connectivity at the Neuromuscular Junction

A primer on new research by Fuentes-Medel and colleagues explains the important role of non-neural cells in clearing neural debris, which is continuously produced during the normal remodeling processes that establish and maintain neural connectivity

A Conserved Role for SNX9-Family Members in the Regulation of Phagosome Maturation during Engulfment of Apoptotic Cells

Clearance of apoptotic cells is of key importance during development, tissue homeostasis and wound healing in multi-cellular animals. Genetic studies in the nematode Caenorhabditis elegans have identified a set of genes involved in the early steps of cell clearance, in particular the recognition and internalization of apoptotic cells. A pathway that orchestrates the maturation of phagosomes containing ingested apoptotic cells in the worm has recently been described. However, many steps in this pathway remain elusive. Here we show that the C. elegans SNX9-family member LST-4 (lateral signaling target) and its closest mammalian orthologue SNX33 play an evolutionary conserved role during apoptotic cell corpse clearance. In lst-4 deficient worms, internalized apoptotic cells accumulated within non-acidified, DYN-1-positive but RAB-5-negative phagosomes. Genetically, we show that LST-4 functions at the same step as DYN-1 during corpse removal, upstream of the GTPase RAB-5. We further show that mammalian SNX33 rescue C. elegans lst-4 mutants and that overexpression of truncated SNX33 fragments interfered with phagosome maturation in a mammalian cell system. Taken together, our genetic and cell biological analyses suggest that LST-4 is recruited through a combined activity of DYN-1 and VPS-34 to the early phagosome membrane, where it cooperates with DYN-1 to promote recruitment/retention of RAB-5 on the early phagosomal membrane during cell corpse clearance. The functional conservation between LST-4 and SNX33 indicate that these early steps of apoptotic phagosome maturation are likely conserved through evolution

Atención de la urgencia quirúrgica durante la pandemia COVID-19. Recomendaciones de la Asociación Española de Cirujanos

La infección por el nuevo coronavirus SARS-CoV-2 (enfermedad por coronavirus 2019 [COVID-19]) ha determinado la necesidad de la reorganización de muchos centros hospitalarios en el mundo. España, como uno de los epicentros de la enfermedad, ha debido asumir cambios en la práctica totalidad de su territorio. Sin embargo, y desde el inicio de la pandemia, en todos los centros que atienden urgencias quirúrgicas ha sido necesario el mantenimiento de su cobertura, aunque igualmente ha sido inevitable introducir directrices especiales de ajuste al nuevo escenario que permitan el mantenimiento de la excelencia en la calidad asistencial. Este documento desarrolla una serie de indicaciones generales para la cirugía de urgencias y la atención al politraumatizado desarrolladas desde la literatura disponible y consensuadas por un subgrupo de profesionales desde el grupo general Cirugía-AEC-COVID-19. Estas medidas van encaminadas a contemplar un riguroso control de la exposición en pacientes y profesionales, a tener en cuenta las implicaciones de la pandemia sobre diferentes escenarios perioperatorios relacionados con la urgencia y a una adaptación ajustada a la situación del centro en relación con la atención a pacientes infectados. New coronavirus SARS-CoV-2 infection (coronavirus disease 2019 [COVID-19]) has determined the necessity of reorganization in many centers all over the world. Spain, as an epicenter of the disease, has been forced to assume health policy changes in all the territory. However, and from the beginning of the pandemic, every center attending surgical urgencies had to guarantee the continuous coverage adopting correct measures to maintain the excellence of quality of care. This document resumes general guidelines for emergency surgery and trauma care, obtained from the available bibliography and evaluated by a subgroup of professionals designated from the general group of investigators Cirugía-AEC-COVID-19 from the Spanish Association of Surgeons, directed to minimize professional exposure, to contemplate pandemic implications over different urgent perioperative scenarios and to adjust decision making to the occupational pressure caused by COVID-19 patients

Glial Processes at the Drosophila Larval Neuromuscular Junction Match Synaptic Growth

Glia are integral participants in synaptic physiology, remodeling and maturation from blowflies to humans, yet how glial structure is coordinated with synaptic growth is unknown. To investigate the dynamics of glial development at the Drosophila larval neuromuscular junction (NMJ), we developed a live imaging system to establish the relationship between glia, neuronal boutons, and the muscle subsynaptic reticulum. Using this system we observed processes from two classes of peripheral glia present at the NMJ. Processes from the subperineurial glia formed a blood-nerve barrier around the axon proximal to the first bouton. Processes from the perineurial glial extended beyond the end of the blood-nerve barrier into the NMJ where they contacted synapses and extended across non-synaptic muscle. Growth of the glial processes was coordinated with NMJ growth and synaptic activity. Increasing synaptic size through elevated temperature or the highwire mutation increased the extent of glial processes at the NMJ and conversely blocking synaptic activity and size decreased the presence and size of glial processes. We found that elevated temperature was required during embryogenesis in order to increase glial expansion at the nmj. Therefore, in our live imaging system, glial processes at the NMJ are likely indirectly regulated by synaptic changes to ensure the coordinated growth of all components of the tripartite larval NMJ