Characterization of Proliferating Neural Progenitors after Spinal Cord Injury in Adult Zebrafish

Zebrafish can repair their injured brain and spinal cord after injury unlike adult mammalian central nervous system. Any injury to zebrafish spinal cord would lead to increased proliferation and neurogenesis. There are presences of proliferating progenitors from which both neuronal and glial loss can be reversed by appropriately generating new neurons and glia. We have demonstrated the presence of multiple progenitors, which are different types of proliferating populations like Sox2⁺ neural progenitor, A2B5⁺ astrocyte/ glial progenitor, NG2⁺ oligodendrocyte progenitor, radial glia and Schwann cell like progenitor. We analyzed the expression levels of two common markers of dedifferentiation like msx-b and vimentin during regeneration along with some of the pluripotency associated factors to explore the possible role of these two processes. Among the several key factors related to pluripotency, pou5f1 and sox2 are upregulated during regeneration and associated with activation of neural progenitor cells. Uncovering the molecular mechanism for endogenous regeneration of adult zebrafish spinal cord would give us more clues on important targets for future therapeutic approach in mammalian spinal cord repair and regeneration.     

Multipotent neural stem cells generate glial cells of the central complex through transit amplifying intermediate progenitors in Drosophila brain development

The neural stem cells that give rise to the neural lineages of the brain can generate their progeny directly or through transit amplifying intermediate neural progenitor cells (INPs). The INP-producing neural stem cells in Drosophila are called type II neuroblasts, and their neural progeny innervate the central complex, a prominent integrative brain center. Here we use genetic lineage tracing and clonal analysis to show that the INPs of these type II neuroblast lineages give rise to glial cells as well as neurons during postembryonic brain development. Our data indicate that two main types of INP lineages are generated, namely mixed neuronal/glial lineages and neuronal lineages. Genetic loss-of-function and gain-of-function experiments show that the gcm gene is necessary and sufficient for gliogenesis in these lineages. The INP-derived glial cells, like the INP-derived neuronal cells, make major contributions to the central complex. In postembryonic development, these INP-derived glial cells surround the entire developing central complex neuropile, and once the major compartments of the central complex are formed, they also delimit each of these compartments. During this process, the number of these glial cells in the central complex is increased markedly through local proliferation based on glial cell mitosis. Taken together, these findings uncover a novel and complex form of neurogliogenesis in Drosophila involving transit amplifying intermediate progenitors. Moreover, they indicate that type II neuroblasts are remarkably multipotent neural stem cells that can generate both the neuronal and the glial progeny that make major contributions to one and the same complex brain structure.     

Neurogenesis in the adult Drosophila brain

Neurodegenerative diseases such as Alzheimer’s and Parkinson’s currently affect ∼25 million people worldwide. The global incidence of traumatic brain injury (TBI) is estimated at ∼70 million/year. Both neurodegenerative diseases and TBI remain without effective treatments. We are utilizing adult Drosophila melanogaster to investigate the mechanisms of brain regeneration with the long-term goal of identifying targets for neural regenerative therapies. We specifically focused on neurogenesis, i.e., the generation of new cells, as opposed to the regrowth of specific subcellular structures such as axons. Like mammals, Drosophila have few proliferating cells in the adult brain. Nonetheless, within 24 hours of a penetrating traumatic brain injury (PTBI) to the central brain, there is a significant increase in the number of proliferating cells. We subsequently detect both new glia and new neurons and the formation of new axon tracts that target appropriate brain regions. Glial cells divide rapidly upon injury to give rise to new glial cells. Other cells near the injury site upregulate neural progenitor genes including asense and deadpan and later give rise to the new neurons. Locomotor abnormalities observed after PTBI are reversed within 2 weeks of injury, supporting the idea that there is functional recovery. Together, these data indicate that adult Drosophila brains are capable of neuronal repair. We anticipate that this paradigm will facilitate the dissection of the mechanisms of neural regeneration and that these processes will be relevant to human brain repair.     

Quantitative analysis reveals dominance of gliogenesis over neurogenesis in an adult brainstem oscillator

Neural stem/progenitor cells in the neurogenic niches of the adult brain are widely assumed to give rise predominantly to neurons, rather than glia. Here, we performed a quantitative analysis of the resident neural progenitors and their progeny in the adult pacemaker nucleus (Pn) of the weakly electric fish Apteronotus leptorhynchus. Approximately 15% of all cells in this brainstem nucleus are radial glia‐like neural stem/progenitor cells. They are distributed uniformly within the tissue and are characterized by the expression of Sox2 and Meis 1/2/3. Approximately 2–3% of them are mitotically active, as indicated by expression of proliferating cell nuclear antigen. Labeling of proliferating cells with a single pulse of BrdU, followed by chases of up to 100 days, revealed that new cells are generated uniformly throughout the nucleus and do not undergo substantial migration. New cells differentiate into S100+ astrocytes and Hu C/D+ small interneurons at a ratio of 4:1, reflecting the proportions of the total glia and neurons in this brain region. The continuous addition of new cells leads to a diffuse growth of the Pn, which doubles in volume and total cell number over the first 2 years following sexual maturation of the fish. However, the number of pacemaker and relay cells, which constitute the oscillatory neural network, remains constant throughout adult life. We hypothesize that the dominance of gliogenesis is an adaptation to the high‐frequency firing of the oscillatory neurons in this nucleus. © 2014 Wiley Periodicals, Inc. Develop Neurobiol 74: 934–952, 2014     

Functioning of glia and neurodegeneration in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The views on the role of glial tissue have changed greatly since the first studies in the field. The cells once regarded as “cell glue” have been shown to play important roles in development, trophic processes, production of navigation signals for axon growth, electric insulation of neurons, creation of a barrier between the brain and the hemolymph, control of extracellular homeostasis, and physiological functioning of the brain. Researchers all over the world are currently turning to Drosophila melanogaster, a well-characterized model organism in genetics, in order to investigate multiple molecular aspects of neurodegeneration processes, since the modeling of neurodegeneration mechanisms in Drosophila has a number of advantages. Fruit flies with a mutation in the swiss cheese (sws) gene show degeneration of neurons and surface glia cells of the optical lobe, and the protein product of the sws gene is essential for maintaining the functionality and integrity of the fly brain. The present review addresses the role of glial cells in Drosophila brain development and in the functioning of the adult fly brain as well as the pattern of expression of the gene sws and the distribution of the product of this gene in neurons and glia.     

The Drosophila dopamine 2-like receptor D2R (Dop2R) is required in the blood brain barrier for male courtship

The blood brain barrier (BBB) has the essential function to protect the brain from potentially hazardous molecules while also enabling controlled selective uptake. How these processes and signaling inside BBB cells control neuronal function is an intense area of interest. Signaling in the adult Drosophila BBB is required for normal male courtship behavior and relies on male-specific molecules in the BBB. Here we show that the dopamine receptor D2R is expressed in the BBB and is required in mature males for normal mating behavior. Conditional adult male knockdown of D2R in BBB cells causes courtship defects. The courtship defects observed in genetic D2R mutants can be rescued by expression of normal D2R specifically in the BBB of adult males. Drosophila BBB cells are glial cells. Our findings thus identify a specific glial function for the DR2 receptor and dopamine signaling in the regulation of a complex behavior.     

Evidence for Sexual Differentiation of Glia in Rat Brain

It is well established that gonadal steroids mediate sexual differentiation of the brain via direct effects on neurons during a restricted critical period. In addition, estrogen can influence glial morphology in the adult brain, andin vitrostudies suggest estrogen induces glial differentiation. However, there is a lack ofin vivoevidence for steroid effects on glia during the critical period. We report here a hormone-mediated sexual differentiation of arcuate glia as early as Postnatal Day 1. Using glial fibrillary acidic protein immunoreactivity (GFAP-ir), we compared the responsiveness of astroglia in the rat arcuate nucleus among five hormonally different groups. The results indicate increased GFAP-ir cell surface area 24 hr after hormonal manipulation in castrate males compared to intact males, intact females (ANOVA;P< 0.01), and females injected with testosterone propionate (50 μg; ANOVA;P< 0.05). However, astroglia in intact males extended their processes significantly greater distances from the cell body compared to all other treatment groups (ANOVA;P< 0.01). The GFAP-ir cells were categorized into four distinct classes ranging from a simple bipolar to a fully stellate morphology. The frequency distribution of classes varied between groups with more stellate cells found in intact males. Finally, these sex differences in arcuate glia persisted into adulthood. We hypothesize that during the critical period, testosterone, or its metabolite estrogen, induce sexual differentiation of glia. We further hypothesize that in females glial cells remain partially undifferentiated and this may be important to glial plasticity seen in adult female arcuate.     

Glial patterning during postembryonic development of central neuropiles in the brain of the honeybee

 Glial cells are involved in several functions during the development of the nervous system. To understand potential glial contributions to neuropile formation, we examined the cellular pattern of glia during the development of the mushroom body, antennal lobe and central complex in the brain of the honeybee. Using an antibody against the glial-specific repo-protein of Drosophila, the location of the glial somata was detected in the larval and pupal brain of the bee. In the early larva, a continuous layer of glial cell bodies defines the boundaries of all growing neuropiles. Initially, the neuropiles develop in the absence of any intrinsic glial somata. In a secondary process, glial cells migrate into defined locations in the neuropiles. The corresponding increase in the number of neuropile-associated glial cells is most likely due to massive immigrations of glial cells from the cell body rind using neuronal fibres as guidance cues. The combined data from the three brain regions suggest that glial cells can prepattern the neuropilar boundaries. Received: 3 November 1996 / Accepted: 7 February 1997     

Drosophila miR-124 regulates neuroblast proliferation through its target anachronism

MicroRNAs (miRNAs) have been implicated as regulators of central nervous system (CNS) development and function. miR-124 is an evolutionarily ancient, CNS-specific miRNA. On the basis of the evolutionary conservation of its expression in the CNS, miR-124 is expected to have an ancient conserved function. Intriguingly, investigation of miR-124 function using antisense-mediated miRNA depletion has produced divergent and in some cases contradictory findings in a variety of model systems. Here we investigated miR-124 function using a targeted knockout mutant and present evidence for a role during central brain neurogenesis in Drosophila melanogaster. miR-124 activity in the larval neuroblast lineage is required to support normal levels of neuronal progenitor proliferation. We identify anachronism (ana), which encodes a secreted inhibitor of neuroblast proliferation, as a functionally important target of miR-124 acting in the neuroblast lineage. ana has previously been thought to be glial specific in its expression and to act from the cortex glia to control the exit of neuroblasts from quiescence into the proliferative phase that generates the neurons of the adult CNS during larval development. We provide evidence that ana is expressed in miR-124-expressing neuroblast lineages and that ana activity must be limited by the action of miR-124 during neuronal progenitor proliferation. We discuss the possibility that the apparent divergence of function of miR-124 in different model systems might reflect functional divergence through target site evolution.     

Developmental regulation of glial cell phagocytic function during Drosophila embryogenesis

The proper removal of superfluous neurons through apoptosis and subsequent phagocytosis is essential for normal development of the central nervous system (CNS). During Drosophila embryogenesis, a large number of apoptotic neurons are efficiently engulfed and degraded by phagocytic glia. Here we demonstrate that glial proficiency to phagocytose relies on expression of phagocytic receptors for apoptotic cells, SIMU and DRPR. Moreover, we reveal that the phagocytic ability of embryonic glia is established as part of a developmental program responsible for glial cell fate determination and is not triggered by apoptosis per se. Explicitly, we provide evidence for a critical role of the major regulators of glial identity, gcm and repo, in controlling glial phagocytic function through regulation of SIMU and DRPR specific expression. Taken together, our study uncovers molecular mechanisms essential for establishment of embryonic glia as primary phagocytes during CNS development.