NeuroCult™ SM1 Neuronal Culture Supplement

概述

使用Brewer 's b27为基础的补充物避免培养失败，该补充物为中枢神经系统来源或人类多能干细胞（hPSC）来源的细胞提供一致的神经元培养体验。

NeuroCult™SM1 （STEMCELL Modified-1）神经元补充剂是基于已发表的配方（Brewer等人）。J Neurosci Res., 1993)和标准化，以更可重复地支持功能性原代和人类多能干细胞（hPSC）来源的神经元的存活和成熟。这种无血清补充剂可以与基础培养基和各种不同的诱导因子或细胞因子一起使用，以支持外胚层、中胚层和内胚层谱系的分化。NeuroCult™SM1也可用作各种可定制应用的血清替代补充剂，如神经毒性测定和钙成像。

为了您的方便，NeuroCult™SM1包含在多个组件中BrainPhys™神经元培养基培养试剂盒用于初级和hpsc衍生的神经元（目录#05792,05793,05794和05795）。欲了解更多详细信息，请参阅BrainPhys™的性能数据下面.

Contains
• Antioxidants
• Vitamin A
• Insulin
• Other ingredients

Subtype
Supplements

Cell Type
Neural Cells, PSC-Derived, Neurons, Pluripotent Stem Cells

Species
Human, Mouse, Rat

Application
Cell Culture, Differentiation, Maintenance

Brand
NeuroCult

Area of Interest
Neuroscience, Stem Cell Biology

Formulation Category
Serum-Free

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Data Figures

Morphology of Neurons in Representative NeuroCult™ SM1 Cultures at 7 and 21 Days in Vitro

Figure 1. Protocol for Plating and Culturing Primary Neurons with the SM1 Culture System

Primary rodent tissue dissociated in papain was plated in NeuroCult™ Neuronal Plating Medium, supplemented with NeuroCult™ SM1 Neuronal Supplement, L-Glutamine, and L-Glutamic Acid. On day 5, primary neurons were transitioned to BrainPhys™ Neuronal Medium, supplemented with NeuroCult™ SM1 Neuronal Supplement, by performing half-medium changes every 3 - 4 days.

Number of Neurons in NeuroCult™ SM1 and TSFM Cultures After 7 and 21 Days in Vitro

Figure 2. The SM1 Culture System Supports Long-Term Culture of Rodent Neurons

Primary E18 rat cortical neurons were cultured in the SM1 Culture System. A large number of viable neurons are visible after (A) 21 and (B) 35 days, as demonstrated by their bright neuronal cell bodies, and extensive neurite outgrowth and branching. Neurons are evenly distributed over the culture surface with minimal cell clumping.

Neurite Outgrowth of Primary Neurons Cultured in NeuroCult™ SM1 and TSFM for 7 and 21 Days

Figure 3. Pre- and Post-Synaptic Markers are Expressed in Rodent Neurons Cultured in the SM1 Culture System

Primary E18 rat cortical neurons were cultured in the SM1 Culture System. At 21 DIV, neurons are phenotypically mature, as indicated by the presence of an extensive dendritic arbor, and appropriate expression and localization of pre-synaptic synapsin (A,C; green) and post-synaptic PSD-95 (A,B; red) markers. Synapsin is concentrated in discrete puncta distributed along the somata and dendritic processes, as defined by the dendritic marker MAP2 (A,D; blue).

MEA data showing mean firing rate of rodent primary neurons cultured in BrainPhys™ and other commercial media

Figure 4. Glucose Supplementation in BrainPhys™ Maintains Neuronal Activity Over 8 Weeks in Culture

Primary E18 rat cortical neurons were cultured with BrainPhys™ and SM1 or other commercially available culture systems for 8 weeks. Neuronal activity can be detected at Day 9 with BrainPhys™, whereas activity is not detected until Day 14 in cultures maintained in either of the Commercial Media with Commercial Supplements. For Commercial Medium and Supplement-cultured neurons, mean firing rate remains low throughout culture. In contrast, a “peak-drop” activity pattern is observed in the Commercial Medium Plus condition, where mean firing rate increases rapidly within 2 days, followed by a drop in activity in the next 2 - 4 days. BrainPhys™and SM1 Kit with 15 mM glucose maintains the highest level of activity throughout the 8-week culture period.

Raster plots showing activity of neurons cultured in BrainPhys™ and other commercial media

Figure 5. BrainPhys™ Supports Improved Neuronal Activity and More Consistent Network Bursting in Long-Term Culture

Raster plots from MEA recordings show the firing patterns of neurons across 8 electrodes at Weeks 2, 4, 6 and 8. Neurons were either cultured with a Commercial Medium with Supplements, Commercial Medium Plus with Supplements, BrainPhys™ and SM1, or BrainPhys™ and SM1 with 15 mM glucose. Detected spikes (black lines), single channel bursts (blue lines; a collection of at least 5 spikes, each separated by an ISI of no more than 100 ms), and network bursts (magenta boxes; a collection of at least 50 spikes from a minimum of 35% of participating electrodes across each well, each separated by an ISI of no more than 100 ms) were recorded for each medium. (A-D) Neurons cultured with Commercial Medium exhibited network bursting in Week 2 but no spiking activity was detected in subsequent timepoints. (E-H) In Commercial Medium Plus-cultured neurons, a high number of spikes and regular network bursting were detected at Week 2. A decreased number of spikes and inconsistent network bursting were observed in later time points, corresponding to the drop in MFR seen in Figure 4. (I-L) Without glucose, individual spiking was observed at Weeks 2 and 4 with BrainPhys™ and SM1 but network bursting was not detected until Weeks 6 and 8. (M-T) In contrast, neurons cultured with BrainPhys™ and SM1 with 15 mM glucose demonstrated strong spiking activity and consistent network bursting at all timepoints. MEA = microelectrode array; ISI = inter-spike interval; MFR = mean firing rate

Protocols and Documentation

Find supporting information and directions for use in the Product Information Sheet or explore additional protocols below.

Document Type

Product Name

Catalog #

Lot #

Language

Document Type

Product Information Sheet

Product Name

NeuroCult™ SM1 Neuronal Supplement

Catalog #

100-1281

Lot #

All

Language

English

Document Type

Product Information Sheet

Product Name

NeuroCult™ SM1 Neuronal Supplement

Catalog #

05711

Lot #

All

Language

English

Document Type

Safety Data Sheet

Product Name

NeuroCult™ SM1 Neuronal Supplement

Catalog #

100-1281

Lot #

All

Language

English

Document Type

Safety Data Sheet

Product Name

NeuroCult™ SM1 Neuronal Supplement

Catalog #

05711

Lot #

All

Language

English

Applications

This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.

Research Area

Workflow Stages

Workflow Stages for Human Pluripotent Stem Cell Research

Reprogramming

Maintenance

Differentiation

Workflow Stages for hPSC-Derived Endoderm Cell Research

Endoderm Induction

Characterization and Isolation

Downstream Differentiation

Workflow Stages for hPSC-Derived Neural Cell Research

Cell Sourcing

Neural Induction

Expansion

Differentiation and Maturation

Characterization

Workflow Stages for Neural Stem and Progenitor Cell Research

Workflow Stages for Neuronal Research

Tissue Dissociation

Cell Culture

Characterization

Resources and Publications

Modelling Lyssavirus Infections in Human Stem Cell-Derived Neural Cultures. V. Sundaramoorthy et al. Viruses 2020 mar

Abstract

Rabies is a zoonotic neurological infection caused by lyssavirus that continues to result in devastating loss of human life. Many aspects of rabies pathogenesis in human neurons are not well understood. Lack of appropriate ex-vivo models for studying rabies infection in human neurons has contributed to this knowledge gap. In this study, we utilize advances in stem cell technology to characterize rabies infection in human stem cell-derived neurons. We show key cellular features of rabies infection in our human neural cultures, including upregulation of inflammatory chemokines, lack of neuronal apoptosis, and axonal transmission of viruses in neuronal networks. In addition, we highlight specific differences in cellular pathogenesis between laboratory-adapted and field strain lyssavirus. This study therefore defines the first stem cell-derived ex-vivo model system to study rabies pathogenesis in human neurons. This new model system demonstrates the potential for enabling an increased understanding of molecular mechanisms in human rabies, which could lead to improved control methods.

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Microtubule-associated protein 2 mediates induction of long-term potentiation in hippocampal neurons Y. Kim et al. FASEB Journal 2020 5

Abstract

Microtubule-associated protein (MAP) 2 has been perceived as a static cytoskeletal protein enriched in neuronal dendritic shafts. Emerging evidence indicates dynamic functions for various MAPs in activity-dependent synaptic plasticity. However, it is unclear how MAP2 is associated with synaptic plasticity mechanisms. Here, we demonstrate that specific silencing of high-molecular-weight MAP2 in vivo abolished induction of long-term potentiation (LTP) in the Schaffer collateral pathway of CA1 pyramidal neurons and in vitro blocked LTP-induced surface delivery of AMPA receptors and spine enlargement. In mature hippocampal neurons, we observed rapid translocation of a subpopulation of MAP2, present in dendritic shafts, to spines following LTP stimulation. Time-lapse confocal imaging showed that spine translocation of MAP2 was coupled with LTP-induced spine enlargement. Consistently, immunogold electron microscopy revealed that LTP stimulation of the Schaffer collateral pathway promoted MAP2 labeling in spine heads of CA1 neurons. This translocation depended on NMDA receptor activation and Ras-MAPK signaling. Furthermore, LTP stimulation led to an increase in surface-expressed AMPA receptors specifically in the neurons with MAP2 spine translocation. Altogether, this study indicates a novel role for MAP2 in LTP mechanisms and suggests that MAP2 participates in activity-dependent synaptic plasticity in mature hippocampal networks.

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Maturation of Human Pluripotent Stem Cell-Derived Cerebellar Neurons in the Absence of Co-culture. T. P. Silva et al. Frontiers in bioengineering and biotechnology 2020

Abstract

The cerebellum plays a critical role in all vertebrates, and many neurological disorders are associated with cerebellum dysfunction. A major limitation in cerebellar research has been the lack of adequate disease models. As an alternative to animal models, cerebellar neurons differentiated from pluripotent stem cells have been used. However, previous studies only produced limited amounts of Purkinje cells. Moreover, in vitro generation of Purkinje cells required co-culture systems, which may introduce unknown components to the system. Here we describe a novel differentiation strategy that uses defined medium to generate Purkinje cells, granule cells, interneurons, and deep cerebellar nuclei projection neurons, that self-formed and differentiated into electrically active cells. Using a defined basal medium optimized for neuronal cell culture, we successfully promoted the differentiation of cerebellar precursors without the need for co-culturing. We anticipate that our findings may help developing better models for the study of cerebellar dysfunctions, while providing an advance toward the development of autologous replacement strategies for treating cerebellar degenerative diseases.

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