BrainPhys™ Neuronal Medium

Serum-free neurophysiological basal medium for improved neuronal function

BrainPhys™ Neuronal Medium

Serum-free neurophysiological basal medium for improved neuronal function

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Serum-free neurophysiological basal medium for improved neuronal function
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Product Advantages


  • More representative of the brain’s extracellular environment

  • Improved neuronal function and a higher proportion of synaptically active neurons

  • Perform functional assays without changing media and shocking cells

  • Supports long-term culture of ES/iPS cell- and CNS-derived neurons

  • Rigorous raw material screening and quality control ensure minimal lot-to-lot variability

What Our Scientist Says

I want to help neuroscientists like you create more physiological culture conditions, for more active and healthy neuronal cultures.

Carmen MakScientist
Carmen Mak, Scientist

Overview

Promote, rather than inhibit, neuronal activity and maturity in your cultured primary or human pluripotent stem cell (hPSC)-derived neurons. Based on the formulation by Bardy and Gage (Bardy et al. PNAS, 2015), BrainPhys™ Neuronal Medium is a serum-free basal medium that is optimized to yield a higher proportion of synaptically active neurons by mimicking the central nervous system (CNS) extracellular environment.

Use BrainPhys™ Neuronal Medium for long-term culture of hPSC- and CNS-derived neurons. To avoid shocking your cells with media changes, you can also use BrainPhys™ medium when performing functional assays, such as microelectrode array-based recordings or live-fluorescent imaging.

To ensure cell health in long-term serum-free culture, BrainPhys™ Neuronal Medium must be combined with an appropriate serum-replacement supplement, such as NeuroCult™ SM1 Neuronal Supplement and/or N2 Supplement-A. For your convenience, various BrainPhys™ kits that include the required supplement(s) for primary or hPSC-derived neurons are also available.

View our additional resources to learn more about the BrainPhys™ system.
Subtype
Basal Media, Specialized Media
Cell Type
Neural Cells, PSC-Derived, Neurons, Pluripotent Stem Cells
Species
Human, Mouse, Rat
Application
Cell Culture, Differentiation, Maintenance
Brand
BrainPhys
Area of Interest
Disease Modeling, Drug Discovery and Toxicity Testing, Neuroscience, Stem Cell Biology
Formulation Category
Serum-Free

Data Figures

Table 1. Properties of Culture Media (C Bardy et al. Proc Natl Acad Sci USA, 2015)

Check-mark denotes physiological conditions

Check-mark denotes physiological conditions and supported activities according to C Bardy et al. Proc Natl Acad Sci USA, 2015.

Rodent Neurons Matured in BrainPhys™ Neuronal Medium

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.

Rodent Neurons Matured in BrainPhys™ Neuronal Medium

Figure 2. Protocol for Culturing hPSCs with the SM1 Culture System

hPSCs were maintained in mTeSR™1 medium and then differentiated using the STEMdiff™ SMADi Neural Induction Kit. Following plating on PLO/laminin, half-medium changes were performed to transition to BrainPhys™ Neuronal Medium for maturation and long-term culture.

Primary Neuronal Cultures Matured in BrainPhys™ Neuronal Medium Have Greater Numbers of Neurons

Figure 3. 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.

Rodent Neuronal Cultures Matured in BrainPhys™ Neuronal Medium Show Improved Excitatory and Inhibitory Synaptic Activity

Figure 4. 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).

Expression of Pre-Synaptic Markers in Rodent Neurons Matured in BrainPhys™ Neuronal Medium

Figure 5. The SM1 Culture System Supports Increased Cell Survival

(A) Primary E18 rat cortical neurons were cultured in the SM1 Culture System or a Competitor Culture System for 21 days. Neurons cultured in the SM1 Culture System have a significantly higher number of viable cells compared to the competitor culture system (n = 4; mean ± 95% CI; *p < 0.05). (B) Primary E18 rat cortical neurons were cultured in Neurobasal® supplemented with NeuroCult™ SM1 Neuronal Supplement (SM1) or competitor B27-like supplements (Competitor 1,2,3) for 21 days. Cultures supplemented with NeuroCult™ SM1 Neuronal Supplement have an equal number of neurons compared to competitor-supplemented cultures. Bars represent standard error of mean.

Raster plots showing activity of neurons cultured in BrainPhys and SM1 versus commercial media

Figure 6. 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

MEA data showing mean firing rate of rodent primary neurons cultured in BrainPhys and SM1 versus commercial media

Figure 7. 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.

hPSC-Derived Neurons Generated in BrainPhys™ Neuronal Medium Express Markers of Neuronal Maturity After 14 and 44 Days of Differentiation

Figure 8. hPSC-Derived Neurons Generated in BrainPhys™ Neuronal Medium Express Markers of Neuronal Maturity After 14 and 44 Days of Differentiation

NPCs were generated from H9 cells using STEMdiff™ Neural Induction Medium in an embryoid body-based protocol. Next, NPCs were cultured in (A,C) BrainPhys™ Neuronal Medium, supplemented with 2% NeuroCult™ SM1 Supplement, 1% N2 Supplement-A, 20 ng/mL GDNF, 20 ng/mL BDNF, 1 mM db-cAMP and 200 nM ascorbic acid to initiate neuronal differentiation, or (B,D) DMEM/F12 under the same supplementation conditions. After 14 and 44 days of differentiation and maturation, neurons express the synaptic marker Synapsin 1 (green) and the mature neuronal marker MAP2 (red). In this example, neurons matured in BrainPhys™ Neuronal Medium show increased Synapsin 1 staining. Scale bar= 100 µm

hPSC-Derived Neurons Generated in BrainPhys™ Neuronal Medium and NeuroCult™ SM1 and N2 Supplements are Healthy and Morphologically Normal

Figure 9. hPSC-Derived Neurons Generated in BrainPhys™ Neuronal Medium and NeuroCult™ SM1 and N2 Supplements are Healthy and Morphologically Normal

NPCs were generated from H9 cells using STEMdiff™ Neural Induction Medium in an embryoid body-based protocol. Next, NPCs were cultured for 44 DIV in (A) BrainPhys™ Neuronal Medium, supplemented with 2% NeuroCult™ SM1 Supplement, 1% N2 Supplement-A, 20 ng/mL GDNF, 20 ng/mL BDNF, 1 mM db-cAMP and 200 nM ascorbic acid to initiate neuronal differentiation, or (B) DMEM/F12 under the same supplementation conditions. Neuronal cultures differentiated from NPCs in BrainPhys™ Neuronal Medium display extensive neurite outgrowth and reduced cellular debris compared to cultures differentiated in DMEM/F12. Scale bar= 100 µm.

hPSC-Derived Neurons Matured in BrainPhys™ Neuronal Medium Show Improved Excitatory and Inhibitory Synaptic Activity

Figure 10. hPSC-Derived Neurons Matured in BrainPhys™ Neuronal Medium Show Improved Excitatory and Inhibitory Synaptic Activity

NPCs were generated from H9 cells using STEMdiff™ Neural Induction Medium in an embryoid body-based protocol. Next, NPCs were cultured for 44 DIV in (A,C) BrainPhys™ Neuronal Medium, supplemented with 2% NeuroCult™ SM1 Supplement, 1% N2 Supplement-A, 20 ng/mL GDNF, 20 ng/mL BDNF, 1 mM db-cAMP and 200 nM ascorbic acid to initiate neuronal differentiation, or (B,D) in DMEM/F12 under the same supplementation conditions. (A,C) Neurons matured in BrainPhys™ Neuronal Medium showed spontaneous excitatory (AMPA-mediated; A) and inhibitory (GABA-mediated; C) synaptic events. The frequency and amplitude of spontaneous synaptic events is consistently greater in neuronal cultures matured in BrainPhys™ Neuronal Medium, compared to neurons plated and matured in DMEM/F12 (B,D). Traces are representative.

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
Catalog #
05790
Lot #
All
Language
English
Document Type
Safety Data Sheet
Catalog #
05790
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.

Resources and Publications

Educational Materials (21)

Publications (51)

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.
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.
One-Stop Microfluidic Assembly of Human Brain Organoids To Model Prenatal Cannabis Exposure. Z. Ao et al. Analytical chemistry 2020

Abstract

Prenatal cannabis exposure (PCE) influences human brain development, but it is challenging to model PCE using animals and current cell culture techniques. Here, we developed a one-stop microfluidic platform to assemble and culture human cerebral organoids from human embryonic stem cells (hESC) to investigate the effect of PCE on early human brain development. By incorporating perfusable culture chambers, air-liquid interface, and one-stop protocol, this microfluidic platform can simplify the fabrication procedure and produce a large number of organoids (169 organoids per 3.5 cm × 3.5 cm device area) without fusion, as compared with conventional fabrication methods. These one-stop microfluidic assembled cerebral organoids not only recapitulate early human brain structure, biology, and electrophysiology but also have minimal size variation and hypoxia. Under on-chip exposure to the psychoactive cannabinoid, $\Delta$-9-tetrahydrocannabinol (THC), cerebral organoids exhibited reduced neuronal maturation, downregulation of cannabinoid receptor type 1 (CB1) receptors, and impaired neurite outgrowth. Moreover, transient on-chip THC treatment also decreased spontaneous firing in these organoids. This one-stop microfluidic technique enables a simple, scalable, and repeatable organoid culture method that can be used not only for human brain organoids but also for many other human organoids including liver, kidney, retina, and tumor organoids. This technology could be widely used in modeling brain and other organ development, developmental disorders, developmental pharmacology and toxicology, and drug screening.