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ADME PK

eCiphr®Neuro-Human

Assessment of Neuronal Activity Using iPSC-derived Human Neurons and Microelectrode Array

  • The eCiphr®Neuro assay uses FUJIFILM Cellular Dynamics, Inc. (FCDI) human iPSC-derived glutamatergic neurons co-cultured with human iPSC-derived astrocytes.
  • Cyprotex’s neuronal assay uses high throughput microelectrode array (MEA) technology to monitor electrophysiological activity.
  • Neurons grown on microelectrode arrays recapitulate many features of neurons in vivo, including spontaneous activity (spiking and bursting), plasticity, organization and responsiveness to a wide range of neurotransmitters and pharmacological agonists/antagonists1,2.
  • This technology provides a unique in vitro system for preclinical drug discovery, neurotoxicity assessment and disease modeling.
The unique capabilities of MEAs to provide functional measurements of network activity, including spontaneous activity, evoked activity, and responses to pharmacological challenges, therefore offers an advantage over other potential screening approaches that rely on biochemical or structural endpoints.

1Robinette BL et al., (2011) Front Neuroeng 4; 1-9

Protocol

eCiphr®Neuro-Human assay protocol

Cell Type FUJIFILM Cellular Dynamics, Inc. (FCDI) Human iPSC-derived glutamatergic neurons (iCell GlutaNeurons) co-cultured with human iPSC-derived astrocytes (iCell Astrocytes)
Analysis Platform Maestro 48-well MEA system (Axion BioSystems)
Test Article Concentrations Four concentrations in triplicate (dependent on customer requirements)
Quality Controls Negative control: 0.2% DMSO (vehicle) Positive controls: picrotoxin and domoic acid (at single concentration)
Data Delivery Firing rate (spikes/second)
Burst rate (bursts/second)
Number of spikes in burst
Burst duration
Coefficient of variation (CV) of the inter-spike intervals (ISI)
Mean ISI within network burst
Skewness ISI (synchrony endpoint)

Data

Data from Cyprotex's eCiphr®Neuro-Human assay

 
Figure 1
Change in spontaneous spike activity in human iPSC-derived glutamatergic neurons co-cultured with human iPSC-derived astrocytes.

The spontaneous spike activity is recorded in human iPSC-derived glutamatergic neurons co-cultured with human iPSC-derived astrocytes using Axion Biosystems microelectrode array Maestro platform. The spike train data is extracted from baseline and post treatment measurements and quantified using custom MATLAB scripts to characterize firing and burst organization. The negative controls 0.2% DMSO (vehicle) and acetaminophen (50µM) caused no change in activity, burst characteristics or synchrony. A distinct pattern of change affecting spike activity, burst characteristics and synchrony is observed with GABAA antagonists picrotoxin and bicuculline. A different but significant pattern of activity can be seen with other proconvulsant toxins such as strychnine, a glycine receptor antagonist and SNC80, a δ-opioid receptor agonist.

Figure 2
Raster plots of spike activity before and after 1 hr treatment with 100µM bicuculline.

Raster plots of spontaneous spike activity generated by a co-culture of human iPSC-derived glutamatergic neurons with human iPSC-derived astrocytes 14 days post plating, before and after 1 hr treatment with 100µM bicuculline. The baseline data represents typical spiking characteristics observed with this co-culture after 2 weeks of maturation which include individual spikes, organized bursts and network synchrony. After a 1 hr treatment with 100µM bicuculline, changes in organization including an increase in spikes in bursts, an increase in the burst duration, an increase in ISI CV (burst organization) and an increase the network synchrony (skewness ISI) endpoint are observed. The qualitative visual differences in the dynamics of the spike train are quantified through computation of the spike train features as seen in Figure 1.

 

References

1 Robinette BL et al, (2011). In vitro assessment of developmental neurotoxicity: use of microelectrode arrays to measure functional changes in neuronal network ontogeny. Front Neuroeng 4; Article 1
2 Bradley JA et al, (2018). In vitro screening for seizure liability using microelectrode array technology. Tox Sci 163(1); 240-253

Other Relevant Articles

Novellino A et al, (2011). Development of micro-electrode array based tests for neurotoxicity: assessment of interlaboratory reproducibility with neuroactive chemicals. Front Neuroeng 4(4); 1-14
Johnstone AFM et al, (2010). Microelectrode arrays: a physiologically based neurotoxicity testing platform for the 21st century. Neuro Tox 31; 331-350
Fuentealba J et al, (2011). Potentiation and inhibition of glycine receptors by tutin. Neuropharmacology 60; 453-459
McConnell ER et al, (2012). Evaluation of multi-well microelectrode arrays for neurotoxicity screening using a chemical training set. Neuro Tox 33; 1048-1057

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