نشكر ساندرا Bargen حصول على الدعم الفني الممتاز. وأيد هذا العمل من قبل إدارة البحوث البروتين (PRD ، TP A1.2 (RC وTS) ، ومولعا RUB البحوث وRektoratsprogramme -- wissenschaftlicher Nachwuchs (AK) وتم الحصول على الاجسام المضادة و39.4D5 F55A10 من ورم هجين دراسات الإنمائية بنك (DSHB ، مدينة أيوا ، IA).

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الإعلان عن أي تضارب في المصالح.

ميزة هذه التقنية preplating lectin المستندة هو أنه أقل تكلفة من الإجراء NTR المستندة P75 بالغسل ، وlectin هي أكثر استقرارا من الأجسام المضادة. إثراء المدرجة في الشكل. (2) وتبويب. 1 يبين أن الإجراء يسمح لتنقية عدد مماثل من الخلايا وأن غالبية هذه الخلايا تعبر عن علامة العصبون الحركي جزيرة - 1 / 2. الخطوة الأكثر أهمية هو الإجراء لعزل النخاع الشوكي القطني. إزالة السحايا وDRGs في (الشكل 2) أمر ضروري لتنقية الإجراء التالي lectin المستندة preplating. إذا كان قد نجح هذا صحيح ، وتقريبا جميع خلايا تعبر عن NTR P75 (صورة لممثل انظر الشكل 3A). سبب الفرق بين التعبير عن جزيرة - 1 / 2 وP75 NTR هو على الارجح بسبب التعبير عن حركية مماثلة القطني تفاضليا المستويات العليا والدنيا للجزيرة 1 / 2 10. في حالة وجود مستويات منخفضة من جزيرة 1 - التعبير / 2 قد هربوا هذا اهتمامنا في تلطيخ immuncytochemical والفرز اللاحقة كما كنا صارمة فيما يتعلق الخلايا إيجابية في مقابل سلبية (Tab. 1). بالإضافة إلى ذلك ، ويبين الجدول 1 بوضوح أن الخلايا المعزولة الإجراء البقاء في حالة صحية جيدة ، وهناك عدد قليل فقط التريبان الأزرق الخلايا الإيجابية التي تشير إلى أن تلف الخلايا بشكل لا رجعة فيه. هذا الإجراء بديل يسمح أيضا عزل العصبونات الحركية من أجنة واحدة ، وبالتالي من الفضلات الوراثي مختلطة من الفئران الجنينية. في الختام ، هذا البديل إلى إجراء بالغسل مع الضد NTR P75 6 ومتشابهة إن لم تكن متطابقة من حيث القدرات النطاقات الممكنة التطبيق ويوفر طريقة فعالة ورخيصة بديلة لتنقية العصبونات الحركية الفأر الجنينية.

<table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم كاشف </td><td> شركة </td><td> فهرس العدد </td><td> تعليقات </td></tr><tr><td> بولي DL - الأورنيثين هيدروبروميد </td><td> سيغما </td><td> P8638 </td><td></td></tr><tr><td> Laminin </td><td> Invitrogen </td><td> 23017-015 </td><td></td></tr><tr><td> HBSS </td><td> Gibco </td><td> 14170 </td><td></td></tr><tr><td> الزجاج coverslips </td><td> الحرارية </td><td></td><td> Ø 10 أو 14mm </td></tr><tr><td> Lectin </td><td> سيغما </td><td> L5142 </td><td></td></tr><tr><td> خلية ثقافة الطبق </td><td> نونك </td><td> 150350 </td><td> Nunclon دلتا سطح </td></tr><tr><td> حصان المصل </td><td> يناريس </td><td> SHD3250ZK </td><td> كل دفعة يجب أن يكون اختبار للثقافة MN </td></tr><tr><td> B27 الملحق </td><td> Gibco </td><td> 17504-044 </td><td></td></tr><tr><td> Glutamax </td><td> Gibco </td><td> 35050-038 </td><td></td></tr><tr><td> CNTF </td><td> سيغما </td><td> N0513 </td><td></td></tr><tr><td> BSA </td><td> Applichem </td><td> A1391 </td><td></td></tr><tr><td> Neurobasal </td><td> Gibco </td><td> 21103-041 </td><td></td></tr><tr><td> ملقط </td><td></td><td></td><td> الفولاذ المقاوم للصدأ ، وحجم 4 أو 5 </td></tr><tr><td> التربسين </td><td> ورثينجتون </td><td> LS003707 </td><td></td></tr><tr><td> التربسين - المانع </td><td> سيغما </td><td> T6522 </td><td></td></tr><tr><td> المضادة للجزيرة - 1 </td><td> DSHB </td><td> 39.4D5 </td><td> خلية ثقافة طاف </td></tr><tr><td> مكافحة Nkx6.1 </td><td> DSHB </td><td> F55A10 </td><td> خلية ثقافة طاف </td></tr><tr><td> مكافحة P75 NTR </td><td> Abcam </td><td> Ab8874 </td><td></td></tr></tbody></table> الجدول 3. الكواشف جدول محدد والمعدات.

lectin-based isolation and culture of mouse embryonic motoneurons

العصبونات الحركية في العمود الفقري نحو تطوير المراحل الجنينية المبكرة تالية للتفتل من خلال تطوير النظام العصبي ، وتنمو في وقت لاحق من التشعبات والمحاور. خلايا عصبية ظهارية من الأنبوب العصبية التي تعبر عن Nkx6.1 هي الخلايا السليفة فريدة للعصبونات حركية الشوكي 1. على الرغم من التحرك نحو حركية مماثلة تالية للتفتل موقفهم النهائي وتنظيم أنفسهم في أعمدة على طول المسالك 2،3 الشوكي. أكثر من 90 ٪ من كل هذه العصبونات الحركية متباينة ، ووضع تعبير عن عوامل النسخ جزيرة 1 / 2. انهم يعصب عضلات الاطراف وكذلك من الجسم والأجهزة الداخلية. من بين أمور أخرى ، أعرب العصبونات الحركية عادة مستقبلات عامل تقارب عالية للعصبية في الدماغ مشتقة (BDNF) وNeurotrophin 3 (NT - 3) ، والتروبوميوزين ذات الصلة كيناز باء وجيم (TrkB ، TrkC). انها لا تعبر عن كيناز التروبوميوزين المتصلة A 4 (TrkA). بجانب مستقبلات two تقارب عالية ، والعصبونات الحركية لا تعبر عن انخفاض مستقبلات neurotrophin تقارب NTR P75. يمكن ربط جميع NTR P75 neurotrophins مع تقارب مماثلة لكنها أقل لجميع neurotrophins من المستقبلات عالية تقارب وربط neurotrophins ناضجة. داخل النخاع الشوكي الجنينية ، وأعرب عن حصرا NTR P75 من العصبونات الحركية في العمود الفقري 5. وقد استخدمت هذه المعرفة لتطوير تقنيات العزل العصبون الحركي لتنقية الخلايا من الغالبية العظمى من الخلايا المحيطة 6. تحولت عزل العصبونات الحركية مع مساعدة من أضداد محددة (بالغسل) ضد من المجالات خارج الخلية P75 NTR إلى أن تكون وسيلة مكلفة وكمية الأجسام المضادة المستخدمة في تجربة واحدة هي عالية نظرا لحجم لوحة المستخدمة لبالغسل. وثمة بديل أكثر اقتصادا هو استخدام lectin. وقد تبين Lectin ربط خصيصا لP75 NTR فضلا 7. الأسلوب التالي يصف أسلوب بديل باستخدام القمح الجرثومية راصة لإجراء preplating بدلا من الأضداد NTR P75. وlectin هو بديل رخيص الثمن للغاية على الضد NTR P75 وتنقية الصفوف باستخدام lectin مماثلة لتلك التي في جسم NTR P75. يمكن عزل العصبونات الحركية من الحبل الشوكي الجنينية باستخدام هذا الأسلوب ، البقاء والنمو خارج neurites.

Watch this Scientific Journal Video about Lectin-based Isolation and Culture of Mouse Embryonic Motoneurons at JoVE.com

Lectin-based Isolation and Culture of Mouse Embryonic Motoneurons

ووصف طريقة بديلة لعزل العصبونات الحركية الفأر الجنينية من الحبل الشوكي. الأسلوب يأخذ في الاعتبار حقيقة أن lectin يمكن ربط العصب تقارب منخفضة p75NTR مستقبلات عوامل النمو. هذا preplating lectin المستندة يسمح تنقية مماثلة لتلك التي مع الأجسام المضادة المحددة ضد p75NTR.

Lectin القائم على عزل وثقافة حركية مماثلة الجنينية الماوس

Spinal motoneurons develop towards postmitotic stages through early embryonic nervous system development and subsequently grow out dendrites and axons. ...

lectin-based-isolation-and-culture-of-mouse-embryonic-motoneurons

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Neuroscience

17.8K Views.  Ruhr-University Bochum. ووصف طريقة بديلة لعزل العصبونات الحركية الفأر الجنينية من الحبل الشوكي. الأسلوب يأخذ في الاعتبار حقيقة أن lectin يمكن ربط العصب تقارب منخفضة p75NTR مستقبلات عوامل النمو. هذا preplating lectin المستندة يسمح تنقية مماثلة لتلك التي مع الأجسام المضادة المحددة ضد p75NTR.

Video: Lectin-based Isolation and Culture of Mouse Embryonic Motoneurons

Dissection and Culture of Commissural Neurons from Embryonic Spinal Cord

Commissural neurons have been widely used to investigate the mechanisms underlying axon guidance during embryonic spinal cord development. The cell bodies of these neurons are located in the dorsal spinal cord and their axons follow stereotyped trajectories during embryonic development. Commissural axons initially project ventrally towards the floorplate. After crossing the midline, these axons turn anteriorly and project towards the brain. Each of these steps is regulated by the action of several guidance cues. Cultures highly enriched in commissural neurons are ideally suited for many experiments addressing the mechanisms of axon pathfinding, including turning assays, immunochemistry and biochemistry. Here, we describe a method to dissect and culture commissural neurons from E13 rat dorsal spinal cord. First, the spinal cord is isolated and dorsal strips are dissected out. The dorsal tissue is then dissociated into a cell suspension by trypsinization and mechanical disruption. Neurons are plated onto poly-L-lysine-coated glass coverslips or tissue-culture dishes. After 30 hours in vitro, most neurons have extended an axon. The purity of the culture (Yam et al. 2009), typically over 90%, can be assessed by immunolabeling with the commissural neuron markers DCC, LH2 and TAG1 (Helms and Johnson, 1998). This neuronal preparation is a useful tool for in vitro studies of the cellular and molecular mechanisms of commissural axon growth and guidance during spinal cord development.

This video demonstrates a method to dissect and culture commissural neurons from E13 rat dorsal spinal cord. Dissociated commissural neurons are useful to study the cellular and molecular mechanisms of axon growth and guidance.

تشريح والثقافة من الخلايا العصبية من النخاع الشوكي صواري الجنينية

Commissural neurons have been widely used to investigate the mechanisms underlying axon guidance during embryonic spinal cord development. The cell bodies ...

Nucleofection and Primary Culture of Embryonic Mouse Hippocampal and Cortical Neurons

Hippocampal and cortical neurons have been used extensively to study central nervous system (CNS) neuronal polarization, axon/dendrite outgrowth, and synapse formation and function. An advantage of culturing these neurons is that they readily polarize, forming distinctive axons and dendrites, on a two dimensional substrate at very low densities. This property has made them extremely useful for determining many aspects of neuronal development. Furthermore, by providing glial conditioning for these neurons they will continue to develop, forming functional synaptic connections and surviving for several months in culture. In this protocol we outline a technique to dissect, culture and transfect embryonic mouse hippocampal and cortical neurons. Transfection is accomplished by electroporating DNA into the neurons before plating via nucleofection. This protocol has the advantage of expressing fluorescently-tagged fusion proteins early in development (~4-8hrs after plating) to study the dynamics and function of proteins during polarization, axon outgrowth and branching. We have also discovered that this single transfection before plating maintains fluorescently-tagged fusion protein expression at levels appropriate for imaging throughout the lifetime of the neuron (&gt; 2 months in culture). Thus, this methodology is useful for studying protein localization and function throughout CNS development with little or no disruption of neuronal function.

This protocol outlines the steps required to dissect, transfect via electroporation and culture mouse hippocampal and cortical neurons. Short-term cultures may be used for studies of axon outgrowth and guidance, while long-term cultures can be used for studies of synaptogenesis and dendritic spine analysis.

Nucleofection والثقافة الأولية من الخلايا العصبية الجنينية الماوس الحصين والقشرية

Hippocampal and cortical neurons have been used extensively to study central nervous system (CNS) neuronal polarization, axon/dendrite outgrowth, and ...

Establishing Embryonic Mouse Neural Stem Cell Culture Using the Neurosphere Assay

In mammalians, stem cells acts as a source of undifferentiated cells to maintain cell genesis and renewal in different tissues and organs during the life span of the animal. They can potentially replace cells that are lost in the aging process or in the process of injury and disease. The existence of neural stem cells (NSCs) was first described by Reynolds and Weiss (1992) in the adult mammalian central nervous system (CNS) using a novel serum&#8208;free culture system, the neurosphere assay (NSA). Using this assay, it is also feasible to isolate and expand NSCs from different regions of the embryonic CNS. These in vitro expanded NSCs are multipotent and can give rise to the three major cell types of the CNS. While the NSA seems relatively simple to perform, attention to the procedures demonstrated here is required in order to achieve reliable and consistent results. This video practically demonstrates NSA to generate and expand NSCs from embryonic day 14-mouse brain tissue and provides technical details so one can achieve reproducible neurosphere cultures. The procedure includes harvesting E14 mouse embryos, brain microdissection to harvest the ganglionic eminences, dissociation of the harvested tissue in NSC medium to gain a single cell suspension, and finally plating cells in NSA culture. After 5-7 days in culture, the resulting primary neurospheres are passaged to further expand the number of the NSCs for future experiments.

This video protocol demonstrates the application of the neurosphere assay for the isolation and expansion of neural stem cells from the ganglionic eminences of embryonic day 14-mouse brain.

تأسيس الخلايا الجذعية الجنينية الماوس العصبية الثقافة باستخدام الفحص Neurosphere

In mammalians, stem cells acts as a source of undifferentiated cells to maintain cell genesis and renewal in different tissues and organs during the life ...

Isolation and Culture of Rat Embryonic Neural Cells: A Quick Protocol

We are describing a quick method to dissociate and culture hippocampal or cortical neurons from E15-17 rat embryos. The procedure can be applied successfully to the isolation of mouse and human primary neurons and neural progenitors. Dissociated neurons are maintained in serum-free medium up to several weeks. These cultures can be used for nucleofection, immunocytochemistry, nucleic acids preparation, as well as electrophysiology. Older neuronal cultures can also be transfected with a good efficiency rate by lentiviral transduction and, less efficiently, with calcium phosphate or lipid-based methods such as lipofectamine.

We describe a rapid methodology to isolate and culture hippocampal and cortical neurons from rodent embryos. This protocol allows us to perform experiments in which nearly pure neuronal cultures are required.

العزلة والثقافة من خلايا الفئران العصبية الجنينية: بروتوكول سريعة

We are describing a quick method to dissociate and culture hippocampal or cortical neurons from E15-17 rat embryos. The procedure can be applied ...

Isolation and Culture of Neural Crest Cells from Embryonic Murine Neural Tube

The embryonic neural crest (NC) is a multipotent progenitor population that originates at the dorsal aspect of the neural tube, undergoes an epithelial to mesenchymal transition (EMT) and migrates throughout the embryo, giving rise to diverse cell types 1-3. NC also has the unique ability to influence the differentiation and maturation of target organs4-6. When explanted in vitro, NC progenitors undergo self-renewal, migrate and differentiate into a variety of tissue types including neurons, glia, smooth muscle cells, cartilage and bone. 
NC multipotency was first described from explants of the avian neural tube7-9. In vitro isolation of NC cells facilitates the study of NC dynamics including proliferation, migration, and multipotency. Further work in the avian and rat systems demonstrated that explanted NC cells retain their NC potential when transplanted back into the embryo10-13. Because these inherent cellular properties are preserved in explanted NC progenitors, the neural tube explant assay provides an attractive option for studying the NC in vitro. 
To attain a better understanding of the mammalian NC, many methods have been employed to isolate NC populations. NC-derived progenitors can be cultured from post-migratory locations in both the embryo and adult to study the dynamics of post-migratory NC progenitors11,14-20, however isolation of NC progenitors as they emigrate from the neural tube provides optimal preservation of NC cell potential and migratory properties13,21,22. Some protocols employ fluorescence activated cell sorting (FACS) to isolate a NC population enriched for particular progenitors11,13,14,17. However, when starting with early stage embryos, cell numbers adequate for analyses are difficult to obtain with FACS, complicating the isolation of early NC populations from individual embryos. Here, we describe an approach that does not rely on FACS and results in an approximately 96% pure NC population based on a Wnt1-Cre activated lineage reporter23. 
The method presented here is adapted from protocols optimized for the culture of rat NC11,13. The advantages of this protocol compared to previous methods are that 1) the cells are not grown on a feeder layer, 2) FACS is not required to obtain a relatively pure NC population, 3) premigratory NC cells are isolated and 4) results are easily quantified. Furthermore, this protocol can be used for isolation of NC from any mutant mouse model, facilitating the study of NC characteristics with different genetic manipulations. The limitation of this approach is that the NC is removed from the context of the embryo, which is known to influence the survival, migration and differentiation of the NC2,24-28.

Isolation of embryonic neural crest from the neural tube facilitates the use of in vitro methods for studying migration, self-renewal, and multipotency of neural crest.

العزلة والثقافة من الخلايا العصبية كريست من الأنبوبة العصبية الجنينية الفئران

The embryonic neural crest (NC) is a multipotent progenitor population that originates at the dorsal aspect of the neural tube, undergoes an epithelial to ...

Dissection, Culture, and Analysis of Xenopus laevis Embryonic Retinal Tissue

The process by which the anterior region of the neural plate gives rise to the vertebrate retina continues to be a major focus of both clinical and basic research. In addition to the obvious medical relevance for understanding and treating retinal disease, the development of the vertebrate retina continues to serve as an important and elegant model system for understanding neuronal cell type determination and differentiation1-16. The neural retina consists of six discrete cell types (ganglion, amacrine, horizontal, photoreceptors, bipolar cells, and M&uuml;ller glial cells) arranged in stereotypical layers, a pattern that is largely conserved among all vertebrates 12,14-18.
While studying the retina in the intact developing embryo is clearly required for understanding how this complex organ develops from a protrusion of the forebrain into a layered structure, there are many questions that benefit from employing approaches using primary cell culture of presumptive retinal cells 7,19-23. For example, analyzing cells from tissues removed and dissociated at different stages allows one to discern the state of specification of individual cells at different developmental stages, that is, the fate of the cells in the absence of interactions with neighboring tissues 8,19-22,24-33. Primary cell culture also allows the investigator to treat the culture with specific reagents and analyze the results on a single cell level 5,8,21,24,27-30,33-39. Xenopus laevis, a classic model system for the study of early neural development 19,27,29,31-32,40-42, serves as a particularly suitable system for retinal primary cell culture 10,38,43-45.
Presumptive retinal tissue is accessible from the earliest stages of development, immediately following neural induction 25,38,43. In addition, given that each cell in the embryo contains a supply of yolk, retinal cells can be cultured in a very simple defined media consisting of a buffered salt solution, thus removing the confounding effects of incubation or other sera-based products 10,24,44-45.
However, the isolation of the retinal tissue from surrounding tissues and the subsequent processing is challenging. Here, we present a method for the dissection and dissociation of retinal cells in Xenopus laevis that will be used to prepare primary cell cultures that will, in turn, be analyzed for calcium activity and gene expression at the resolution of single cells. While the topic presented in this paper is the analysis of spontaneous calcium transients, the technique is broadly applicable to a wide array of research questions and approaches (Figure 1).

Dissection, Culture, and Analysis of Xenopus laevis Embryonic Retinal Tissue

Xenopus laevis provides an ideal model system for studying cell fate specification and physiological function of individual retinal cells in primary cell culture. Here we present a technique for dissecting retinal tissues and generating primary cell cultures that are imaged for calcium activity and analyzed by in situ hybridization.

تشريح والثقافة، وتحليل<em&gt; القيطم المورق</em&gt; الأنسجة الجنينية الشبكية

The process by which  the anterior region of the neural plate gives rise to the vertebrate retina  continues to be a major focus of both clinical and ...

Isolation and Culture of Mouse Cortical Astrocytes

Astrocytes are an abundant cell type in the mammalian brain, yet much remains to be learned about their molecular and functional characteristics. In vitro astrocyte cell culture systems can be used to study the biological functions of these glial cells in detail. This video protocol shows how to obtain pure astrocytes by isolation and culture of mixed cortical cells of mouse pups. The method is based on the absence of viable neurons and the separation of astrocytes, oligodendrocytes and microglia, the three main glial cell populations of the central nervous system, in culture. Representative images during the first days of culture demonstrate the presence of a mixed cell population and indicate the timepoint, when astrocytes become confluent and should be separated from microglia and oligodendrocytes. Moreover, we demonstrate purity and astrocytic morphology of cultured astrocytes using immunocytochemical stainings for well established and newly described astrocyte markers. This culture system can be easily used to obtain pure mouse astrocytes and astrocyte-conditioned medium for studying various aspects of astrocyte biology.

Astrocytes have been recognized to be versatile cells participating in fundamental biological processes that are essential for normal brain development and function, and central nervous system repair. Here we present a rapid procedure to obtain pure mouse astrocyte cultures to study the biology of this major class of central nervous system cells.

العزلة وثقافة الخلايا النجمية الماوس القشرية

Astrocytes  are an abundant cell type in the mammalian brain, yet much remains to be  learned about their molecular and functional characteristics. In ...

Isolation and Culture of Endothelial Cells from the Embryonic Forebrain

Embryonic brain endothelial cells can serve as an important tool in the study of angiogenesis and neurovascular development and interactions. The two vascular networks of the embryonic forebrain, pial and periventricular, are spatially distinctive and have different origins and growth patterns. Endothelial cells from the pial and periventricular vascular networks have unique gene expression profiles and functions. Here we present a step-by-step protocol for isolation, culture, and verification of pure populations of endothelial cells from the periventricular vascular network (PVECs) of the embryonic forebrain (telencephalon). In this approach, telencephalon devoid of pial membrane obtained from embryonic day 15 mice is minced, digested with collagenase/dispase, and dispersed mechanically into a single cell suspension. PVECs are purified from cell suspension using positive selection with anti-CD-31/PECAM-1 antibody conjugated to MicroBeads using a strong magnetic separation method. Purified cells are cultured on collagen 1&#160;coated culture dishes in endothelial cell culture medium until they become confluent and further subcultured. PVECs obtained with this protocol exhibit cobblestone and spindle shaped phenotypes, as visualized by phase-contrast light microscopy and fluorescence microscopy. Purity of PVEC cultures was established with endothelial cell markers. In our hands, this method reliably and consistently yields pure populations of PVECs. This protocol will benefit studies aimed at gaining mechanistic insights into forebrain angiogenesis, understanding PVEC interactions, and cross-talks with neuronal cell types and holds tremendous potential for therapeutic angiogenesis.

This video demonstrates an easy and reliable strategy for preparation of pure cultures of endothelial cells from the embryonic forebrain within 10-12 days and will be useful for research focused on many aspects of cerebral angiogenesis.

العزلة والثقافة من الخلايا البطانية من الجنينية الدماغ المقدم

Embryonic brain endothelial cells can serve as an important tool in the study of angiogenesis and neurovascular development and interactions. The two ...

Isolation, Culture and Long-Term Maintenance of Primary Mesencephalic Dopaminergic Neurons From Embryonic Rodent Brains

Degeneration of mesencephalic dopaminergic (mesDA) neurons is the pathological hallmark of Parkinson&#8217;s diseae. Study of the biological processes involved in physiological functions and vulnerability and death of these neurons is imparative to understanding the underlying causes and unraveling the cure for this common neurodegenerative disorder. Primary cultures of mesDA neurons provide a tool for investigation of the molecular, biochemical and electrophysiological properties, in order to understand the development, long-term survival and degeneration of these neurons during the course of disease. Here we present a detailed method for the isolation, culturing and maintenance of midbrain dopaminergic neurons from E12.5 mouse (or E14.5 rat) embryos. Optimized cell culture conditions in this protocol result in presence of axonal and dendritic projections, synaptic connections and other neuronal morphological properties, which make the cultures suitable for study of the physiological, cell biological and molecular characteristics of this neuronal population.

The causes of degeneration of midbrain dopaminergic neurons during Parkinson&#8217;s disease are not fully understood. Cellular culture systems provide an essential tool for study of the neurophysiological properties of these neurons. Here we present an optimized protocol, which can be utilized for in vitro modeling of neurodegeneration.

العزلة والثقافة وطويل الأجل صيانة الابتدائية الدماغ المتوسط ​​الدوبامين العصبية من المخ الجنينية القوارض

Degeneration of mesencephalic dopaminergic (mesDA) neurons is the pathological hallmark of Parkinson&#8217;s diseae. Study of the biological processes ...

Isolation and Culture of Adult Neural Stem Cells from the Mouse Subcallosal Zone

Adult neural stem cells (aNSCs) can be used for the regeneration of damaged brain tissue. NSCs have the potential for differentiation and proliferation into three types of cells: neurons, astrocytes, and oligodendrocytes. Identifying aNSC-derived regions and characterizing the aNSC properties are critical for the potential use of aNSCs and for the elucidation of their role in neural regeneration. The subcallosal zone (SCZ), located between white matter and the hippocampus, has recently been reported to contain aNSCs and continuously give rise to neuroblasts. A low percentage of aNSCs from the SCZ is differentiated into neurons; most cells are differentiated into glial cells, such as oligodendrocytes and astrocytes. These cells are suggested to have a therapeutic potential for traumatic cortical injury. This protocol describes in detail the process to generate SCZ-aNSCs from an adult mouse brain. A brain matrix with intervals of 1 mm is used to obtain the SCZ-containing coronal slices and to precisely dissect the SCZ from the whole brain. The SCZ sections are initially subjected to a neurosphere culture. A well-developed culture system allows for the verification of their characteristics and can increase research on NSCs. A neurosphere culture system provides a useful tool for determining proliferation and collecting the genuine NSCs. A monolayer culture is also an in vitro system to assay proliferation and differentiation. Significantly, this culture system provides a more homogenous environment for NSCs than the neurosphere culture system. Thus, using a discrete brain region, these culture systems will be helpful for expanding our knowledge about aNSCs and their applications for therapeutic uses.

Establishing culture systems for the expansion of adult neural stem cells (aNSCs) allows for the examination and application of aNSCs for therapy. The subcallosal zone (SCZ) has recently been recognized as a novel neuroblast-forming region in adult mice. Here, methods for the isolation, expansion, and differentiation of SCZ-aNSCs are described.

العزلة والثقافة من الخلايا الجذعية البالغة العصبية من المنطقة تحت الثفني ماوس

Adult neural stem cells (aNSCs) can be used for the regeneration of damaged brain tissue. NSCs have the potential for differentiation and proliferation ...