Interactions Between Astrocytes and Oligodendroglia in Myelin Development and Related Brain Diseases / 神经科学通报·英文版

Astrocytes (ASTs) and oligodendroglial lineage cells (OLGs) are major macroglial cells in the central nervous system. ASTs communicate with each other through connexin (Cx) and Cx-based network structures, both of which allow for quick transport of nutrients and signals. Moreover, ASTs interact with OLGs through connexin (Cx)-mediated networks to modulate various physiological processes in the brain. In this article, following a brief description of the infrastructural basis of the glial networks and exocrine factors by which ASTs and OLGs may crosstalk, we focus on recapitulating how the interactions between these two types of glial cells modulate myelination, and how the AST-OLG interactions are involved in protecting the integrity of the blood-brain barrier (BBB) and regulating synaptogenesis and neural activity. Recent studies further suggest that AST-OLG interactions are associated with myelin-related diseases, such as multiple sclerosis. A better understanding of the regulatory mechanisms underlying AST-OLG interactions may inspire the development of novel therapeutic strategies for related brain diseases.

Neuronal guidance genes in health and diseases

Neurons migrate from their birthplaces to the destinations, and extending axons navigate to their synaptic targets by sensing various extracellular cues in spatiotemporally controlled manners. These evolutionally conserved guidance cues and their receptors regulate multiple aspects of neural development to establish the highly complex nervous system by mediating both short- and long-range cell-cell communications. Neuronal guidance genes (encoding cues, receptors, or downstream signal transducers) are critical not only for development of the nervous system but also for synaptic maintenance, remodeling, and function in the adult brain. One emerging theme is the combinatorial and complementary functions of relatively limited classes of neuronal guidance genes in multiple processes, including neuronal migration, axonal guidance, synaptogenesis, and circuit formation. Importantly, neuronal guidance genes also regulate cell migration and cell-cell communications outside the nervous system. We are just beginning to understand how cells integrate multiple guidance and adhesion signaling inputs to determine overall cellular/subcellular behavior and how aberrant guidance signaling in various cell types contributes to diverse human diseases, ranging from developmental, neuropsychiatric, and neurodegenerative disorders to cancer metastasis. We review classic studies and recent advances in understanding signaling mechanisms of the guidance genes as well as their roles in human diseases. Furthermore, we discuss the remaining challenges and therapeutic potentials of modulating neuronal guidance pathways in neural repair.

Effects of delayed ramelteon treatment on neurological function and tissue repair after cryogenic traumatic brain injury / 中国药理学通报

:Aim To study the effects of continuous dai¬ly administration of ramelteon starting at the subacute period of cryogenic traumatic brain injury (cTBI) on neurological function and brain tissue repair in mice. Methods Thirty male C57BL/6 mice were randomly divided into sham group, vehicle group and ramelteon treatment groups. The right sensory-motor cortex was damaged by pressing a copper probe precooled by liq¬uid nitrogen onto the skull. Ramelteon ( 10 nig 'kg-1 • d"1) was administered by gavage every day starting at different time points after cTBI (1 h, 1 d,3 d) until sacrifice on day 14. Beam walking test and open field test were used to evaluate the motor function. Toluidine blue staining was used to measure the infarct volume. Immunofluorescence was used to detect the expression of GAP-43 and synaptophysin in peri-infarct area. Mi¬croglia activation was detected using Iba-1. The area and thickness of glial scars were analyzed by detecting GFAP positive areas. Results All three treatment ( 1 h - 14 d, 1 - 14 d, and 3 - 14 d) significantly im¬proved cTBI induced motor dysfunction, reduced the infarct volume, elevated the expression of GAP -43 and synaptophysin, and decreased the area and thick¬ness of glial scar and microglia activation. In addition, all ramelteon treatment groups had similar effects on the above indexes. Conclusions Delayed ramelteon treatment can improve neurological dysfunction after cTBI,and the therapeutic time window can be delayed for up to three days after cTBI. Inhibiting glial scar formation and microglia activation, and promoting ax- onal regeneration and synaptogenesis may contribute to the beneficial effects of ramelteon.

Fetal psychism: neurodynamic and psychoanalytic bases / Psiquismo fetal: bases neurodinâmicas e psicanalíticas

Backgroung: Neuroscientific research has provided great discoveries regarding the understanding of the brain functioning and its neural circuits. With advances in studies on fetal behavior, new discussions have arisen about the existence of a possible rudimentary psychic apparatus. Questioning the existence of a psychism in the fetus becomes doubly challenging. First, because of the controversy that exists in the field of neuroscience about the studies of epiphenomena. Second, because of the difficulty that psychoanalysis has in accepting the existence of a psychic structure before birth. This study was carried out considering all these controversies and scientific limitations, and for this reason it should be understood as a theoretical hypothesis and an invitation to a broad and transdisciplinary view on the complexity of human behavior. From an extensive review on the development of the nervous system and fetal synaptogenesis, and combining neurophysiological and neurophysical research, it was possible to create a link with the Freudian theory of psychic energy described in the Project for a scientific psychology. From these joints, questions were raised about fetal development, especially in the preterm phase, which would be composed of intense synaptic activities, especially in the somatosensory and thalamocortical regions that would receive exogenous and endogenous stimuli, both acting to generate an accumulation of psychic energy. Thus, it was hypothesized that this intense flow of energy would be the first sign of the development of the primitive psychic apparatus in the fetus. Thus, it was possible to assume that during the preterm period this cathected energy discharge could project directly onto the limbic and motor brain structures and leave unconscious memory traces of intrauterine life experiences. These influences of a psychic nature, together with epigenetic factors, would contribute to the appearance of certain behavioral and neurodevelopmental disorders. Therefore, suggesting an early transdisciplinary approach in at-risk infants exposed to environmental or epigenetic stressors during the gestational period, especially during the synaptic plasticity window, will provide a therapeutic opportunity through psychic reorganization and sensorimotor integration.

Introdução: As pesquisas neurocientíficas têm proporcionado grandes descobertas no que concerne ao entendimento sobre o funcionamento cerebral e seus circuitos neurais. Com os avanços nos estudos sobre o comportamento fetal novas discussões têm surgido acerca da existência de um possível aparelho psíquico rudimentar. Questionar a existência de um psiquismo no feto, torna-se duplamente desafiador. Primeiro pela controvérsia que existe no âmbito da neurociência sobre os estudos dos epifenômenos. Segundo, pela própria dificuldade que a psicanálise tem em aceitar a existência de uma estrutura psíquica antes do nascimento. Este estudo foi realizado considerando todas estas controvérsias e limitações científicas, e por este motivo deve ser entendido como uma hipótese teórica e um convite para uma ampla e transdisciplinar visão sobre a complexidade do comportamento humano. A partir de uma extensa revisão sobre o desenvolvimento do sistema nervoso e da sinaptogênese fetal, e associando as pesquisas neurofisiológicas e da neurofísica, foi possível criar uma articulação com a teoria Freudiana da energia psíquica descrita no Projeto para uma psicologia científica. A partir destas articulações, levantou-se questionamentos sobre o desenvolvimento fetal, especialmente na fase pré-termo, o qual seria composto por atividades sinápticas intensas, especialmente nas regiões somatossensoriais e talamocorticais que receberiam estímulos exógenos e endógenos, ambos atuando para gerar um acúmulo de energia psíquica. Desta forma, criou-se uma hipótese de que este intenso fluxo de energia seria o primeiro sinal do desenvolvimento do aparelho psíquico primitivo no feto. Assim, foi possível supor que durante o período pré-termo esta descarga de energia catexizada poderia se projetar diretamente sobre as estruturas cerebrais límbicas e motoras e deixar traços de memória inconscientes das experiências da vida intrauterina. Seriam estas influências de natureza psíquica em conjunto com os fatores epigenéticos, que contribuiriam para o aparecimento de certos transtornos comportamentais e do neurodesenvolvimento. Sendo assim, sugerir uma abordagem transdisciplinar precoce em bebês de risco expostos a fatores estressores ambientais ou epigenéticos durante o período gestacional, especialmente durante a janela de plasticidade sináptica, proporcionará uma oportunidade terapêutica através da reorganização psíquica e da integração sensoriomotora.

Bases neurobiológicas del autismo y modelos celulares para su estudio experimental / Neurobiological bases of autism and cellular models for its experimental study

Los trastornos del espectro autista (TEA) son una alteración funcional de la corteza cerebral, que presenta anomalías estructurales del neurodesarrollo que afectan fundamentalmente a la función sináptica y el patrón de conexiones dentro y entre columnas corticales. Desde su aspecto etiológico, el TEA tiene una importante carga genética, considerándose un desorden derivado de una combinación de mutaciones "de novo", asociadas a una predisposición derivada de variaciones comunes heredadas. Las principales anomalías genéticas asociadas a TEA implican genes que codifican proteínas de la sinapsis. Así, en pacientes con TEA se han descrito alteraciones del desarrollo inicial de las sinapsis en los circuitos de conexión entre áreas corticales de procesamiento complejo. La complejidad molecular observada en la predisposición a desarrollar un TEA, junto con la diversidad de fenotipos estructurales neuronales, ha hecho que los modelos animales reproduzcan solo parcialmente el TEA. Para avanzar en el estudio experimental se hace pues necesario desarrollar modelos más representativos, como son los modelos celulares derivados de células humanas. En las últimas décadas, el desarrollo de la biología de las células madre nos da medios para acceder a paradigmas experimentales sobre células derivadas de individuos con TEA. Actualmente, los modelos de células plutipotentes inducidas (IPs) derivadas de células humanas permiten profundizar en el estudio de las bases moleculares y celulares del TEA. Sin embargo, presentan problemas inherentes derivados de la manipulación experimental que conlleva la reprogramación de la expresión génica, por lo que otros modelos celulares se están también postulando como válidos.

Autism Spectrum Disorders (ASD) are a functional alteration of the cerebral cortex, which presents structural neurodevelopmental anomalies that affect synaptic function and the pattern of connections within and between cortical columns. From its etiological aspect, ASD has an important genetic load, considering a polygenic disorder, derived from a combination of "de novo" genetic mutations, associated to a predisposition derived from common inherited variations. The main genetic anomalies associated with ASD involve genes that encode proteins of the synapse. Thus, in patients with ASD, alterations in the initial development of the synapses have been described in the connection circuits between complex processing cortical areas. The molecular complexity observed in the predisposition to develop an ASD, together with the diversity of structural phenotypes, has made animal models reproduce only partially the ASD. To advance in the experimental study it is therefore necessary to develop representative models, such as cellular models derived from human cells. In recent decades, the advances in stem cell biology give us a way to apply experimental paradigms in cells derived from individuals with ASD. Currently, induced pluripotent cells (IPs) derived from human adult cells allow deepening the study of molecular and cellular bases of the neuronal development in humans, as well as the anomalies in this development, which give rise to disorders such as ASD. However, they present inherent problems derived from the experimental manipulation that involves the reprogramming of gene expression, therefore other models are also been explored.

Role of brain-specific angiogenesis inhibitor 1 in synapse and cognitive impairment / 中华行为医学与脑科学杂志

Adhesion G protein-coupled receptors(aGPCRs) play a significant role in cognitive impairment related diseases. As an important member of aGPCRs, brain-specific angiogenesis inhibitor 1(BAI1) has a prominent impact on anti-angiogenesis, anti-tumor and participating in immune phagocytosis. Recent research found out that BAI1 exerts a great influence on synaptogenesis and synaptic plasticity, but few studying concerning BAI1 in nervous system. Nowadays, the aging of population aggravates the occurrence of cognitive impairment. The pathogenesis of Alzheimer's disease and vascular cognitive impairment remains elusive, and identification of cognitive impairment at an early stage faces challenges. In the stage of mild cognitive impairment, synaptic damage is evident. BAI1 can regulate the function of postsynaptic membrane, synaptogenesis, synaptic signal transmission and the morphological development of dendritic spines. Therefore, it may potentially act as an early-warning index and intervention target for cognitive impairment.

Role of brain-specific angiogenesis inhibitor 1 in synapse and cognitive impairment / 中华行为医学与脑科学杂志

Adhesion G protein-coupled receptors(aGPCRs) play a significant role in cognitive impairment related diseases.As an important member of aGPCRs,brain-specific angiogenesis inhibitor 1 (BAI1) has a prominent impact on anti-angiogenesis,anti-tumor and participating in immune phagocytosis.Recent research found out that BAI1 exerts a great influence on synaptogenesis and synaptic plasticity,but few studying concerning BAI1 in nervous system.Nowadays,the aging of population aggravates the occurrence of cognitive impairment.The pathogenesis of Alzheimer's disease and vascular cognitive impairment remains elusive,and identification of cognitive impairment at an early stage faces challenges.In the stage of mild cognitive impairment,synaptic damage is evident.BAI1 can regulate the function of postsynaptic membrane,synaptogenesis,synaptic signal transmission and the morphological development of dendritic spines.Therefore,it may potentially act as an early-warning index and intervention target for cognitive impairment.

Expression of Neuroligin1 after Spinal Cord Injury in Rats / 中国康复理论与实践

Objective To observe the change of expression of neuroligin1 (NL1) in injured spinal cord in rats. Methods A total of 60 adult female Sprague-Dawley rats were randomly divided into control group (n=30) and experi-ment group (n=30), and both groups were further arranged into three days, seven days, 14 days, 21 days, and 28 days subgroups. The control group accepted T9-11 laminectomy, while the experiment group was injured at T10 spi-nal cord hit by Allen's technique (10 g×25 mm). They were assessed with Basso, Beattie & Bresnahan locomotor rating scale (BBB scale), in their time-points, while Golgi-Cox staining was used to observe the variation of den-drites and density of dendritic spine in the white matter located at upper end of spinal cord injured center, and im-munofluorescence staining was used to detect the expression of NL1. Results The score of BBB scale reduced in the experiment group compared with that in the control group in every sub-group (P<0.001). Compared to the control group, both dendrites and density of dendritic spine in the white mat-ter decreased with time after injury (P<0.001), while the level of NL1 increased three days after injury, peaked on the 14th day after injury (P<0.05). Conclusion NL1 increases spontaneously after spinal cord injury, but it is not enough to promote synaptic regeneration.

Research progress of the mTOR signaling pathway underlying the antidepressant actions of ketamine / 中国药学杂志

Depression is a common, serious psychiatric disease. Despite a wide range of antidepressants available, only one third of the patients show significant mood improvement in response lo an initial antidepressant treatment. Moreover, there is a time-lag of weeks to months with currently available medications; the suffering of patients could not be relieved quickly. Ketamine was found to produce rapid antidepressant responses in treatment resistant major depressive disorder patients. Molecular and cellular studies in rodent models demonstrated that ketamine stimulates mammalian target of rapamycin (mTOR) signaling and increases the initial translation of mTOR and downstream molecules, these effects of ketamine are accompanied by increasing levels of maturity and quantity of spine synapses in the prefrontal cortex. Ketamine was found to produce rapid antidepressant responses in treatment of depression. These studies identify the characterization of the mTOR signaling pathway in depression and its action in response to antidepressants shows great potential for the identification of new therapeutic targets for the development of antidepressant drugs.

Neurotoxicidad de los plaguicidas como agentes disruptores endocrinos: una revisión / Neurotoxicity of pesticides as endocrine disruptors agents: a review

Los plaguicidas se encuentran en el medio ambiente como contaminantes formando mezclas complejas, los estudios recientes se han centrados en la evaluación de los efectos y riesgos que pueden causar estas mezclas de plaguicidas en el ser humano y los ecosistemas.Esta revisión hace hincapié a tres plaguicidas específicos, representativos de los grupos químicos organofosforados, carbamato de tipo ditiocarbamato y triazinas, como son: Paratión® etílico, Mancozeb® y Atrazina®, respectivamente. Ha sido demostrado en animales de experimentación la neurotoxicidad de estos plaguicidas por alteración de la transmisión sináptica y de los mecanismos de la homeostasis del sistema neuroendocrino, lo cual explica la acción de estos plaguicidas como disruptores endocrinos. Varios trabajos demuestran que estos plaguicidas son potencialmente tóxicos a la salud humana, y aún en bajas concentraciones, pueden afectar al organismo causando alteraciones en el sistema neuroendocrino, especialmente en los ejes hipotálamo-hipófisis-gónada e hipotálamo-hipófisis-tiroides. Los investigadores destacan que los momentos de particular sensibilidad de la exposición a plaguicidas disruptores endocrinos son las etapas tempranas de la vida como: durante el desarrollo embrionario y la primera infancia, períodos en los cuales los procesos de crecimiento son controlados por las hormonas afectando el desarrollo del cerebro, el sistema inmune y otros órganos como el sistema reproductor y tiroideo. Los cambios neuroquímicos provocados por la exposición a estos plaguicidas durante el desarrollo cerebral constituyen un alto riesgo porque son capaces de interferir en los procesos de neurogénesis y sinaptogénesis, afectando de manera adversa las funciones cognitivas y motoras, cuyos efectos se observan durante las etapas posteriores de la vida.

Pesticides are found in the environment as complex mixtures forming pollutants, recent studies have focused on evaluating the effects and risks that may cause these mixtures of pesticides on humans and ecosystems. This review highlights three specific, representative of organophosphorus pesticide chemical groups carbamate, dithiocarbamate and triazine type, such as: ethyl Paratión®, Mancozeb® and Atrazina®, respectively.It has been demonstrated in experimental animal’s neurotoxicity of these pesticides by altering synaptic transmission and mechanisms of homeostasis of the neuroendocrine system, which explains the action of these pesticides as endocrine disruptors. Several studies show that these pesticides are potentially toxic to human health, and even in low concentrations, can affect the body causing alterations in the neuroendocrine system, especially in the hypothalamic-pituitary-gonadal and hypothalamic-pituitary-thyroid axis. The researchers point out that the moments of particular sensitivity of exposure Endocrine disruptor pesticide are the early stages of life as during embryonic development and early childhood periods in which growth processes are controlled by hormones affecting the development brain, immune system and other organs such as the thyroid and reproductive system. The neurochemical changes induced by exposure to these pesticides during brain development are at high risk because they are able to interfere with the processes of neurogenesis and synaptogenesis, adversely affecting cognitive and motor functions, whose effects are observed during the later stages of life.

Avaliação do desenvolvimento do sistema nervoso central de camundongos Balb/c expostos à fumaça do cigarro no início do período pós-natal / Evaluation of the brain development in BALB/c mice exposed to environmental tobacco smoke in the early postnatal period

Diversos estudos relatam os efeitos da exposição à nicotina nos períodos pré e pós-natal, contudo, pouco se sabe a respeito dos efeitos da fumaça do cigarro na cascata de eventos que caracteriza o desenvolvimento do sistema nervoso central (SNC). Neste contexto, o objetivo deste trabalho foi esclarecer se a exposição à fumaça do cigarro no início do período pós-natal induz prejuízo ao desenvolvimento do SNC na infância, e as possíveis consequências na adolescência e na fase adulta. Camundongos BALB/c foram expostos a uma mistura de fumaça central e lateral do cigarro referência 3R4F (Universidade de Kentucky, EUA), desde o 3° dia de vida pós-natal (P) até P14 por duas horas diárias. Nossos resultados indicam que a exposição à fumaça do cigarro no início do período pós-natal induz prejuízo ao processo de aprendizado e memória e aumento na ansiedade em todas as idades avaliadas, além de induzir diminuição da atividade locomotora na infância e na adolescência. Ainda, observamos diminuição dos níveis de BDNF e das proteínas sinápticas sinapsina e sinaptofisina no hipocampo, cerebelo, córtex pré-frontal e estriado. A fumaça do cigarro também induz diminuição na porcentagem de fibras mielinizadas no nervo óptico e aumento da proteína básica de mielina (PBM) no cerebelo na infância, além de diminuição da PBM no telencéfalo e tronco encefálico na adolescência e no cerebelo na fase adulta. Nossos resultados sugerem que a exposição à fumaça do cigarro no início do período pós-natal causa prejuízo ao desenvolvimento do SNC, sendo que não há reversão dos efeitos observados no aprendizado e memória ou mesmo nos níveis de proteína pré-sináptica na adolescência e na fase adulta

Several studies show the effects of nicotine exposure during pre- and postnatal period. However, little is known about the effects of environmental tobacco smoke (ETS) in the cascade of events that characterizes the brain development. Thus, the aim of this study was to evaluate the effects of ETS in early brain development. BALB/c mice were exposed to a mixture of mainstream and sidestream of tobacco smoke of reference cigarettes 3R4F (University of Kentucky, EUA) from the 3rd (P3) to the 14th (P14) day of life, during 2h/day. Our results showed that ETS induced impairment in learning and memory and increased anxiety in all the ages evaluated. In addition, there was a decrease in locomotor activity during childhood and adolescence. ETS also induced impairment in synaptic transmission, by a decrease in synapsin, synaptophysin and BDNF in hippocampus, cerebellum and prefrontal cortex as compared to the control group. The percentage of myelinated fibers in the optic nerve in childhood and in myelin basic protein (MBP) in the telencephalon and brainstem were lower in adolescents mice exposed to ETS when compared to the control group. In cerebellum, there was an increase in MBP in infants and a decrease in adults compared to the control group. Taken all together, our results suggest that the exposure to ETS in the early postnatal period induces impairment to the brain development. It is noteworthy that the effects on learning and memory or even in the presynaptic protein levels were not reversed in adolescence and adulthood

New Perspectives of Dyrk1A Role in Neurogenesis and Neuropathologic Features of Down Syndrome

Down syndrome (DS) is one of the most common genetic disorders accompanying with mental retardation, cognitive impairment, and deficits in learning and memory. The brains with DS also display many neuropathological features including alteration in neurogenesis and synaptogenesis and early onset of Alzheimer's disease (AD)-like symptoms. Triplication of all or a part of human chromosome 21, especially the 21q22.1~21q22.3 region called 'Down syndrome critical region (DSCR)', has been considered as the main cause of DS. One gene product of DSCR, dual-specificity tyrosine-phosphorylation-regulated kinase 1A (Dyrk1A), has been highlighted as a key contributor to the neural consequences of DS. This minireview summarizes accumulating recent reports about Dyrk1A involvement in the neuritogenesis, synaptogenesis, and AD-like neurofibrillary tangle formation, which is mainly focusing on Dyrk1A-mediated regulation of cytoskeletal proteins, such as tubulin, actin, and microtubule-associated protein tau. Understanding the molecular mechanisms of these phenomena may provide us a rational for new preventive and therapeutic treatment of DS.

Differential Expression Levels of Synaptophysin through Developmental Stages in Cerebral Cortices of Mouse Brain / 체질인류학회지

It is well known that differentiation and growth of central nervous system are accomplished through relatively early stages of development. The formation of neural synapse indicates beginning of electrical signaling between neurons, so that may be a critical step in the differentiation of neurons as well as the development and growth of central nervous system. The purpose of this study was to investigate the differential expression levels and patterns of synaptic marker (synaptophysin) between superficial and deep layers of cerebral cortex according to the developmental stages. We introduced immunofluorescence staining of synaptophysin combined with densitometric analysis for the morphological quantification. The intensities of synaptophysin immuno-reactivities in deep layers of cerebral cortices were significantly higher compared to superficial layers in cerebral cortices of embryonic and neonatal mice. The significant increase of synaptophysin expression in the deep layer of cerebral cortex was mainly confined to the embryonic stage. As the expression of synaptophysin gradually decrease thereafter, the difference of expression level between superficial and deep layers could not find in the adult mice. From this study, we could confirm indirectly through synaptophysin that synaptogenic activities in the deep layer of cerebral cortex shows unique pattern especially during the early stages of brain development. Results from this study will be helpful for understanding different patterns of synaptogenesis among the various regions of developing brain.

Differential Expression Levels of Synaptophysin through Developmental Stages in Cerebral Cortices of Mouse Brain / 체질인류학회지

It is well known that differentiation and growth of central nervous system are accomplished through relatively early stages of development. The formation of neural synapse indicates beginning of electrical signaling between neurons, so that may be a critical step in the differentiation of neurons as well as the development and growth of central nervous system. The purpose of this study was to investigate the differential expression levels and patterns of synaptic marker (synaptophysin) between superficial and deep layers of cerebral cortex according to the developmental stages. We introduced immunofluorescence staining of synaptophysin combined with densitometric analysis for the morphological quantification. The intensities of synaptophysin immuno-reactivities in deep layers of cerebral cortices were significantly higher compared to superficial layers in cerebral cortices of embryonic and neonatal mice. The significant increase of synaptophysin expression in the deep layer of cerebral cortex was mainly confined to the embryonic stage. As the expression of synaptophysin gradually decrease thereafter, the difference of expression level between superficial and deep layers could not find in the adult mice. From this study, we could confirm indirectly through synaptophysin that synaptogenic activities in the deep layer of cerebral cortex shows unique pattern especially during the early stages of brain development. Results from this study will be helpful for understanding different patterns of synaptogenesis among the various regions of developing brain.

Differential Expressions of Synaptogenic Markers between Primary Cultured Cortical and Hippocampal Neurons

Primary dissociated neuronal cultures are widely used research tools to investigate of pathological mechanisms and to treat various central and peripheral nervous system problems including trauma and degenerative neuronal diseases. We introduced a protocol that utilizes hippocampal and cortical neurons from embryonic day 17 or 18 mice. We applied appropriate markers (GAP-43 and synaptophysin) to investigate whether neurite outgrowth and synaptogenesis can be distinguished at a particular period of time. GAP-43 was found along the neural processes in a typical granular pattern, and its expression increased proportionally as neurites lengthened during the early in vitro period. Unlike GAP-43, granular immunoreactive patterns of synaptophysin along the neurites were clearly found from day 2 in vitro with relatively high immunoreactive levels. Expression of synaptic markers from cortical neurons reached peak level earlier than that of hippocampal neurons, although neurite outgrowths of hippocampal neurons were faster than those of cortical neurons. The amount of peak synaptic markers expressed was also higher in cortical neurons than that in hippocampal neurons. These results strongly suggest the usefulness of primary cultured neurons from mice embryos for synaptic function and plasticity studies, because of their clear and typical patterns of morphology that establish synapses. Results from this study also suggest the proper amount of time in vitro according to neuronal types (cortical or hippocampal) when utilized in experiments related with synaptogenesis or synaptic activities.

Effects of Erythropoietin in Hypoxia-Induced Ischemia on Differentiated Human Neuroblastoma SH-SY5Y and Rat Stroke Model / 대한정신약물학회지

OBJECTIVE: The hematopoietic cytokine, erythropoietin (EPO) is known to have neuroprotective effects including promotion of neuronal survival and regeneration after ischemic injury. This study was to investigate the effects of EPO on synaptogenesis and neural restoration in the ischemic condition on neuronal differentiated SH-SY5Y cells and on the behaviors in rat animal model induced by middle cerebral artery occlusion. METHODS: We analyzed the neurite outgrowth and the gene expression of differentiated human neuroblastoma SH-SY5Y cells after the hypoxic stress. Moreover, we performed the motor functional behavior test in EPO treatment of Sprague Dawley rats following cerebral ischemia induced by middle cerebral artery's occlusion (MCAO). RESULTS: Treatment of 2 and 10 units EPO for 1 week showed increase of neurite outgrowth SH-SY5Y cells, compared with non-treatment group (p < 0.05). The results of reverse transcriptase-polymerose chain reaction (RT-PCR) also showed that both synaptophysin (SYP) genes and Growth Associated protein 43 (GAP43) genes in EPO treated cells were significant increased compared with non-treated ischemic group, respectively. The foot fault behavior was recovered in MCAO with EPO treatment group than MCAO group, significantly. CONCLUSION: The elongation of neurite and the increased expressions of SYP and GAP43, and recovered behavioral evidence in the EPO treatment are involved in possible role in neural restoration and synaptogenesis in hypoxic injuried brain. In this study, we suggest that EPO treatment will be may supportive medication to stroke patients to improve the functional brain disturbance.

Increase of Synapsin I, Phosphosynapsin (ser-9), and GAP-43 in the Rat Hippocampus after Middle Cerebral Artery Occlusion

The loss of neurons and synaptic contacts following cerebral ischemia may lead to a synaptic plastic modification, which may contribute to the functional recovery after a brain lesion. Using synapsin I and GAP-43 as markers, we investigated the neuronal cell death and the synaptic plastic modification in the rat hippocampus of a middle cerebral artery occlusion (MCAO) model. Cresyl violet staining revealed that neuronal cell damage occurred after 2 h of MCAO, which progressed during reperfusion for 2 weeks. The immunoreactivity of synapsin I and GAP-43 was increased in the stratum lucidum in the CA3 subfield as well as in the inner and outer molecular layers of dentate gyrus in the hippocampus at reperfusion for 2 weeks. The immunoreactivity of phosphosynapsin was increased in the stratum lucidum in the CA3 subfield during reperfusion for 1 week. Our data suggest that the increase in the synapsin I and GAP-43 immunoreactivity probably mediates either the functional adaptation of the neurons through reactive synaptogenesis from the pre-existing presynaptic nerve terminals or the structural remodeling of their axonal connections in the areas with ischemic loss of target cells. Furthermore, phosphosynapsin may play some role in the synaptic plastic adaptations before or during reactive synaptogenesis after the MCAO.

Synaptic Localization of NMDA Receptor and Shank Protein in Hippocampal Neuron in Vitro / 대한해부학회지

The existence of NMDA receptor and a new organizer protein, Shank, in the postsynaptic density was studied with the cultured hippocampal neurons using by double immunofluorescence method. The hippocampi from embryonic 18 days were dissected and hippocampal neurons were obtained from dissociated hippocampi with 0.25% trypsin and 0.1% DNase in PBS. The hippocampal neurons were plated with density 3,600/cm2 on the poly-L-lysine coated coverglass and cultured 37degrees C, 5% CO2 incubator for 5 weeks. The N2 supplemented MEM was used as a culture medium. Following results are obtained from experiments: 1. The 3~5 minor processes from the cell body of hippocampal neurons were observed at 20 hr in vitro. One of the minor processes was elongated and looked like an axon, and another minor processes showed dendritic branching pattern with slender in thickness. 2. The excitatory NMDA receptor colocalized with PSD-95 which is the postsynaptic density protein. The presynaptic protein, synapsin 1, was closely apposed with PSD-95. 3. Shank which is an organizer protein colocalize with NMDA receptor/PSD-95 complex in the postsynaptic density. Shank proteins may be concerned with the cluster formation of NMDA receptor/PSD-95 in the postsynaptic membrane.

Cloning and characterization of developmentally differentially expressed novel genes from developing rat cerebral cortex / 대한해부학회지

The structural complexity and heterogeneity of cerebral cortex have been obvious since the earliest days of light microscopy. In fact, if there is one word that captures many of the key attributes of cortical structures, it is diversity. Neurodevelopmental approach is the one of the effective ways to understand complicated structures of cerebral cortex. In this experiment, as a first step to clone the genes related with development of cerebral cortex, the developmentally differentially expressed genes were cloned from developing rat brain with ordered differential display PCR(ODD-PCR). Novel genes were screened from these cloned genes by sequencing and sequence analysis with blast search program. The developmental expression patterns of novel genes in the cerebral cortex were investigated with in situ hybridization histochemistry on the developing and adult rat brain sections. Among the forty one developmentally differentially expressed cDNA bands, amplified with InEGA primer and TEAC primer by ODD PCR, twenty novel genes were screened by sequencing and sequence analysis with blast search program. Through the investigation of developmental expression pattern with in situ hybridization histochemistry, specific expression of five novel genes in the developing rat cerebral cortex was identified. 20-E14-2 was highly expressed in the cerebral cortex during the period of neurogenesis. The expression of 20-E20-1, 20-E20-6b, and 20-P0-5 was relatively well matched with neuronal cell migration in the cerebral cortex. And the strong expression of 20-P0-8b was observed in the neuronal cells of cerebral cortex during the period of syanptogenesis. From these results, it may be suspected that the five novel genes play roles in the development of cerebral cortex.