Human Induced Pluripotent Stem Cell-derived Glutamatergic Neurons - Evaluating Maturation and Neurotoxic Predictability Using a Microelectrode Array Platform

Sep 11, 2026

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Microelectrode array (MEA) electrophysiology is used to assess seizurogenic and neurotoxic compounds with hiPSC-derived neuronal models, addressing prior limitations in burst organization. Glutamatergic neurons are cultured on 48-well MEA plates for up to three weeks at multiple densities, with or without defined ratios of GABAergic neurons and astrocytes. Co-culture accelerates maturation, increasing firing rate, burst spiking, and multiple network endpoints versus ~5 to ~15 days in vitro. Responsiveness is evaluated using domoic acid, DMSO, and seizurogenic agents, revealing distinct dose-response and burst/network disruption patterns.

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Abstract

Cellular Models and Experimental Setup

iCell neuronal populations

Maestro microelectrode array platform

Methods

Plate preparation and cell seeding

Maintenance and recording schedule

Data acquisition and analysis endpoints

Results: Maturation

Human Induced Pluripotent Stem Cell-derived Glutamatergic Neurons: Evaluating Maturation and Neurotoxic Predictability in the Presence or Absence of GABAergic Neurons and Astrocytes Using a Microelectrode Array Platform Jenifer A. Bradley1 , Kile Magnin2 , Christian Kannemeier2 , Brad Swanson2 , Christopher J. Strock 1 1. Cyprotex US, LLC. Watertown, MA. 2. Cellular Dynamics International, A FujiFilm Company, Madison, WI | Abstract
The prediction of seizurogenic and neurotoxic compounds using microelectrode array (MEA) technology and rodent neuronal models has proven to be a very powerful tool. Early hiPSC derived neuronal models lacked complex burst organization, making electrophysiological neurotoxic prediction challenging when utilizing an MEA platform. Advancements in hiPSC neuronal models have addressed these disadvantages. Glutamatergic neurons were plated on 48-well MEA plates and maintained for up to three weeks at different densities with and without various ratios of GABAergic neurons and astrocytes. When cultured alone, the glutamatergic neurons demonstrate robust firing rates, burst organization and network characteristics within 2 weeks of plating. Spike train analyses indicate a significant maturation rate with a 100% increase in firing rate when plated with astrocytes and GABAergic neurons, 1200% increase in spikes that occur within bursts, and 200-400% increase in other burst and network organization endpoints (i.e. ISI CV, ISI Skewness) when comparing ~5 days post plating to ~ 15 days. We also evaluated these combinations for their response to the neurotoxin domoic acid, negative control DMSO (0 .2 %) and seizurogenic agents, picrotoxin, gabazine, 4-aminopyridine, SCN80 and strychnine. As expected, 10µM domoic acid caused a complete loss in spike activity while 0.2 % DMSO caused insignificant changes in activity. 4-aminopyridine decreased overall activity while increasing the burst length by 100% with an overall increase in burst organization. Both strychnine and SNC80 caused a unique dose response pattern with changes in spike/burst rates and a deleterious effect on burst and network organization. The GABAA antagonists, picrotoxin and gabazine, did not have robust responses in this study and require further evaluation. In conclusion, hiPS cell-derived glutamatergic neurons, alone, and in combination with hiPSC derived GABAergic neurons and astrocytes create a robust population of cells that are ideal for evaluating developmental patterns as well as responses to some neurotoxic/seizurogenic compounds when tested on a multi-well MEA platform. | | --- | | Cellular Dynamics International’s (CDI) GlutaNeurons, iCell Neurons, iCell Astrocytes

􀁺 iCell GlutaNeurons: iPS cell-derived human cortical neurons consisting primarily of 90% glutamatergic (excitatory) neurons.
􀁺 iCell Neurons: iPS cell-derived mixed populations of human cortical neurons consisting of 95% GABAergic (inhibitory) neurons.
􀁺 iCell Astrocytes: iPS cell-derived human astrocytes (>95% pure astrocytes) . | Axion BioSystems’Maestro Microelectrode Array (MEA) Platform Figure 1A.The Maestro, Axion BioSystems. 768 recording channels with fully integrated heater and software controls. Accommodates 12, 48 and 96 well MEA plates. Figure 1B. 48 well configured MEA plate, Axion BioSystems. 16 microelectrodes per well, ANSI compliant, nano-textured gold electrodes with evaporation reducing lid. All recordings were acquired on the Axion Maestro platform using 48-well configured MEA plates. The Axion ECmini was used to deliver pre-mixed CO2 throughout the recordings. A Constant temperature of 37 °C was maintained through the software controller. Methods 􀁺 48-well MEA plates were pre-coated with a 0.1 % PEI solution and allowed to dry overnight. One hour before plating cells, the plates were treated with a laminin solution by dispensing a 10µL dot directly over the electrode grid and incubating at 37 °C. 􀁺 iCell GlutaNeurons, iC

FAQs

How are glutamatergic neurons cultured for maturation and testing?

Glutamatergic neurons are plated on 48-well MEA plates and maintained for up to three weeks, with different densities and with or without GABAergic neurons and astrocytes.

What does co-culture with astrocytes and GABAergic neurons change in maturation?

Co-culture increases maturation, including higher firing rates, more spikes occurring within bursts, and elevated burst and network organization endpoints when comparing earlier and later post-plating timepoints.

Which compounds are used to evaluate neurotoxic or seizurogenic responsiveness?

The study tests domoic acid, DMSO negative control, and seizurogenic agents including picrotoxin, gabazine, 4-aminopyridine, SCN80, and strychnine.

Sep 11, 2026

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