BrainPhys™

For Improved Neuronal Function

BrainPhys™ Neuronal Medium is a new, defined and serum-free neuronal basal medium for the culture of primary neurons and the generation and maturation of human pluripotent stem cell (hPSC)-derived neurons. Based on the formulation invented by Dr. Cedric Bardy and Dr. Fred H. Gage (C Bardy et al. Proc Natl Acad Sci USA, 20151), BrainPhys™ Neuronal Medium is more representative of the brain’s extracellular environment and increases the proportion of synaptically active neurons. Using BrainPhys™ Neuronal Medium increases the physiological relevance of in vitro neuroscience research and improves the probability of successful translation of scientific discoveries from the laboratory to the clinic.

Our goal was to develop a culture condition that would support both healthy neuronal circuits and let us monitor the functional properties of mature neurons, without having to switch to different media. Specifically, we reasoned that such cell media should allow for optimal physiological neuronal activity. After sharing our work with our colleagues, we wanted to make this neuronal medium easily available to the scientific community.

Dr. Cedric Bardy, Salk Institute for Biological Studies

What is BrainPhys™?

On a journey through the brain, fly amongst the neurons and discover why scientists are so excited about BrainPhys™.

Why Use BrainPhys™ Neuronal Medium?

  • More representative of the brain’s extracellular environment.
  • Improved neuronal function and a higher proportion of synaptically active neurons.
  • Perform functional assays without replacing media.
  • Supports long-term culture of primary and ES/iPS cell-derived neurons.
  • Rigorous raw material screening and quality control ensure minimal lot-to-lot variability.

BrainPhys™ Neuronal Medium

Cell Type:

  • Primary mouse and rat neurons
  • hPSC-derived neural cells

Formulation:

Applications:

  • Culture of primary mouse or rat neurons
  • Differentiation and maturation of human ES/iPS cell-derived neurons
  • Microelectrode array-based recording of neuronal activity
  • Fluorescence-based imaging, including calcium imaging and optogenetic stimulation and recording
  • Transdifferentiation (lineage conversion) of somatic cells to neurons

Brand History

Realizing the shortcomings of traditional culture media, Cedric Bardy, in the laboratory of Fred H. Gage, set out to identify which properties could be altered to better support in vitro neuronal activity. They found many crucial neurophysiological properties to be impaired, such as action potential generation and synaptic activity. Their solution was the invention of a new tissue culture medium, BrainPhys™, published in 2015 (C Bardy et al. Proc Natl Acad Sci USA, 2015). In BrainPhys™, cultures experience brain-like physiological conditions and have a higher proportion of synaptically active neurons. We developed BrainPhys™ Neuronal Medium based on this formulation, applying more than 20 years of experience in media manufacturing, together with industry-leading raw material screening and performance testing programs, to create a product with unfailing performance and minimal lot-to-lot variability.

Data

Table 1. Properties of Culture Media According to Bardy et al.1

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. Rodent Neurons Matured in BrainPhys™ Neuronal Medium are Healthy and Morphologically Mature

(A,C) Primary rat E18 cortical neurons were plated in NeuroCult™ Neuronal Basal Medium, supplemented with NeuroCult™ SM1 Neuronal Supplement. After 5 DIV, the cultures were transitioned to BrainPhys™ Neuronal Medium, supplemented with NeuroCult™ SM1, by performing half-medium changes every 3-4 days. Neurons were cultured for 14 (A) or 21 (C) DIV. (B,D) Primary rat E18 cortical neurons were plated and matured in a traditional neuronal medium (Neurobasal Medium), supplemented with NeuroCult™ SM1 Neuronal Supplement for 14 (B) or 21 (D) DIV. Neuronal morphology of BrainPhys™ Neuronal Medium-matured neurons is consistent with neurons plated and matured in a traditional neuronal medium.

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

Figure 2. Primary Neuronal Cultures Matured in BrainPhys™ Neuronal Medium Have Greater Numbers of Neurons

Primary rat E18 cortical neurons were plated in either NeuroCult™ Neuronal Basal Medium (NCSM1) or Neurobasal Medium (NBSM1), supplemented with NeuroCult™ SM1. After 5 DIV, half of the cultures were transitioned to BrainPhys™ Neuronal Medium, supplemented with NeuroCult™ SM1, by performing half‑medium changes every 3-4 days. The other half of the cultures were maintained in the same medium as used for plating. After 21 DIV, more neurons were evident in the cultures matured in BrainPhys™ Neuronal Medium, regardless of whether NeuroCult™ Neuronal Basal Medium or Neurobasal Medium was used as the plating medium. (n = 2, mean ± SEM [triplicate wells were set up for each experiment]).


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

Figure 3. Rodent Neuronal Cultures Matured in BrainPhys™ Neuronal Medium Show Improved Excitatory and Inhibitory Synaptic Activity

(A,C) Primary rat E18 cortical neurons were plated in NeuroCult™ Neuronal Basal Medium, supplemented with NeuroCult™ SM1 Neuronal Supplement. After 5 DIV, the cultures were transitioned to BrainPhys™ Neuronal Medium, supplemented with NeuroCult™ SM1 Neuronal Supplement, by performing half-medium changes every 3 - 4 days. Neurons were cultured for 21 DIV. (B,D) Primary rat E18 cortical neurons were plated and matured in a traditional neuronal medium (Neurobasal Medium), supplemented with NeuroCult™ SM1 Neuronal Supplement for 21 DIV. (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 a traditional neuronal medium (B,D). Traces are representative.


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

Figure 4. Expression of Pre-Synaptic Markers in Rodent Neurons Matured in BrainPhys™ Neuronal Medium

Primary rat E18 cortical neurons were plated in NeuroCult™ Neuronal Basal Medium, supplemented with NeuroCult™ SM1 Neuronal Supplement. After 5 DIV, the cultures were transitioned to BrainPhys™ Neuronal Medium, supplemented with NeuroCult™ SM1 Neuronal Supplement, by performing half-medium changes every 3 - 4 days. Neurons cultured for 21 DIV are phenotypically mature, as indicated by the presence of an extensive dendritic arbor. The pre-synaptic marker synapsin (A,B; green) is concentrated in discrete puncta distributed along the somata and dendritic processes, as defined by the dendritic marker MAP2 (A,C; red). Scale bar= 50 µm.


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

Figure 5. 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 Generated in BrainPhys™ Neuronal Medium Express Markers of Neuronal Maturity After 14 and 44 Days of Differentiation

Figure 6. 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 Matured in BrainPhys™ Neuronal Medium Show Improved Excitatory and Inhibitory Synaptic Activity

Figure 7. 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.

References

  1. Bardy C et al. (2015) Neuronal medium that supports basic synaptic functions and activity of human neurons in vitro. Proc Natl Acad Sci 112 (20) E2725-E2734.
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