Maturation of neuronal network activity in ioGlutamatergic Neurons. The network activity of ioGlutamatergic Neurons was characterised using the MaxTwo HD-MEA System. The Axon Tracking Assay reveals the emergence of complex neural circuits through reconstructed axonal paths of travelling action potentials (left). Over the culture period, an increase in both total axon length (middle) and firing rate (right) was observed.  Together, these data show the functional maturation of neuronal networks in physiologically relevant co-culture with human iPSC-derived astrocytes.

Data courtesy of Charles River Laboratories and MaxWell Biosystems.

Electrophysiological characterisation of ioSensory Neurons using medium throughput automated patch clamp analysis. Representative recordings from ioSensory Neurons show voltage-gated sodium (Na_v) current-voltage relationships under (A) control and (B) tetrodotoxin (TTX) conditions, and (C) TTX concentration–response. (D) Example action potential firing measured in current clamp mode. Data from the “Characterising ioSensory Neurons on the Patchliner, an automated patch clamp system”  application note from Nanion Technologies.

On the Patchliner, ioSensory Neurons achieved 100% capture and >80% seal rates. Recordings confirmed NaV currents (TTX-s and TTX-r), ligand-gated responses to GABA, glutamate, and P2X, heat-activated currents, and action potential firing under current clamp.

Functional excitatory–inhibitory network interactions in a human neuronal tri-culture model. ioGABAergic Neurons form functional networks with excitatory ioGlutamatergic Neurons and inhibit their activity in a cell number dependent manner (A). MEA analysis shows that ioGABAergic Neurons inhibit the excitatory activity of ioGlutamatergic Neurons in a ratio dependent manner (B), and that the excitatory–inhibitory balance can be further modulated by drugs targeting GABergic signalling.

ioGlutamatergic Neurons, ioGABAergic Neurons, and hiPSC-derived astrocytes provide a robust in vitro model to study network hyperexcitability and support the discovery of new therapies for neurological diseases, such as epilepsy, autism and schizophrenia.

Functional network maturation of ioMotor Neurons in co-culture with rat astrocytes. Representative multielectrode array (MEA) data shows neuronal activity in ioMotor Neurons co-cultured with rat cortical astrocytes over a 42 day period. (A) Percent active area: Quantitative analysis demonstrating a progressive increase in the percentage of the array area showing neuronal activity from Day 7 through Day 42. (B) Action potential tracking for individual neurons: Clump-free morphology enables single cell assessments of ioMotor Neurons. (C) Mean firing rate heatmaps: Comparison of spontaneous activity between Day 21 (left) and Day 42 (right). The heat maps show a significant increase in mean firing rates (Hz) over time, which is consistent with the progressive maturation of the functional motor neuron network.

This protocol details how to culture ioMotor Neurons with rat cortical astrocytes for MEA assays. It includes plate preparation, media recipes, and step-by-step instructions to support stable network activity and extended recordings of motor neuron function.

1. What is an MEA assay?

An MEA assay (multi-electrode array assay) is an in vitro electrophysiology technique used to study the electrophysiology of neurons by recording extracellular action potentials across many electrodes simultaneously. 

2. What does an MEA measure?

An MEA measures changes in extracellular voltage generated by neuronal activity. These signals provide quantitative readouts such as spike rate, burst frequency, synchrony, and overall network behaviour. When combined with defined human iPSC-derived cells, MEA measurements can reveal subtle functional differences between control and disease models.

3. What is MEA analysis?

MEA analysis refers to the extraction and interpretation of electrophysiological metrics from MEA recordings. Software pipelines quantify activity patterns—such as spike trains, network bursts, connectivity, and maturation profiles—to help assess how cells respond to compounds, perturbations, or genetic backgrounds within an MEA assay workflow.

4. What is the MEA system?

A MEA system includes the multi-electrode array plate, acquisition hardware, environmental controls, and analysis software used to record neuronal activity in vitro. Systems such as Axion or MaxWell support high-throughput MEA assays and integrate with automated workflows for long-term functional recordings.

5. How do experimental conditions affect MEA assay results?

MEA assays are highly sensitive to experimental conditions—such as cell density, purity, maturation stage, and co-culture composition—which directly influence neuronal activity. Using defined human iPSC-derived cell types reduces biological variability at the source, helping researchers generate more reproducible electrophysiology data and clearer interpretation of neuronal network behaviour.