the Medicine Maker, 2022
Jim Ross, PhD
How bioelectronic assays helped demonstrate the value of 3D iPSC cultures in modeling human brain development
The 2006 discovery that adult skin cells could be transformed into pluripotent stem cells and differentiated back into any cell type resolved many ethical and regulatory hurdles facing their embryonic counterparts. But scientists continue to face a number of challenges in realizing the full potential of induced pluripotent stem cells (iPSCs) – both in research and in the clinic. Whether trying to develop a successful stem cell-based therapy or model complex diseases, biologists must confirm that their differentiated cells behave like the native cells they are trying to emulate.
Soon after the development of iPSCs, researchers developed iPSC-derived cardiomyocytes and tested their genetic, protein, and electrophysiological profiles using traditional techniques (1). Similarly, researchers created and characterized iPSC-derived neurons. Today, scientists are looking at these cells as potential regenerative therapies for heart damage, spinal injury, neurodegenerative disease, and more. They could potentially improve health and reduce the need for transplants (2).
However, to be used as treatments, iPSC-derived neurons and cardiomyocytes must function like neurons and cardiomyocytes do in the body. iPSC-derived neurons must communicate with other cells; iPSC-derived cardiomyocytes must keep the heart beating in a regular rhythm. Though scientists have many simple methods to characterize gene and protein markers at their disposal, it is these cells’ electrophysiological properties that really define them.
Characterizing neurons presents an especially unique challenge. Brain function is complex and occurs at the circuit and network levels. Scientists can use patch-clamp electrophysiology to study the electrical properties of individual neurons, but the technique does not capture the rich network activity that takes place in vivo. The function of an iPSC-derived neuron is best confirmed when it is part of a network. Therefore, knowing how an iPSC-derived neuron interacts with the neurons around it is critical for understanding if it is potentially suitable for implantation.
To measure in vitro neural network activity, researchers can use a bioelectronic assay, which measures network activity among cultured neurons in real time over days or weeks. The assay records these data using multielectrode arrays (MEAs) embedded in the base of each well of a multiwell plate. Unlike other techniques, which use probes and dyes to track activity, bioelectronic assays are non-invasive and do not interfere with the natural activity of the cells under examination. Scientists use these assays to study a myriad of neurological conditions, but advances in the study of 3D cerebral organoids are enabling new insights into how these networks develop. They are allowing scientists to better understand the origins of the most complex human organ: the brain.
I’d like to draw attention to the fascinating work of Alysson Muotri of the University of California, San Diego, who has been using mini-brains to study neurological and psychiatric diseases. To characterize the mini-brain platform and demonstrate that it recapitulates human physiology, Muotri and his team used a bioelectronic assay to perform weekly extracellular recordings of spontaneous electrical activity over the course of 10 months (3). Each well in the bioelectronic assay contained 64 microelectrodes, enabling the team to generate raster plots and map neural activity across the surface of the mini-brain. The researchers measured firing rates, burst frequency, and synchrony and characterized the dynamics of network activity. This robust data allowed them to analyze how the electrical activity in the organoids evolved over time. It also let them demonstrate that the mini-brain activity increased in complexity over the course of 10 months, correlating with observations from human electroencephalograms in utero (4).
The ability of bioelectronic assays to profile complex neuronal networks over a period of weeks and months allowed Muotri’s group to demonstrate the ability of their 3D iPSC cultures to model human brain development in vitro. These models could be used to study neurodevelopmental diseases and assess potential treatments. Researchers could functionally benchmark electrically active cells developed for therapeutic purposes against their native counterparts or ensure that new disease models accurately reflect human physiology.
The ease of performing bioelectronic assays could also help therapeutic companies save considerable amounts of time and money in the long run by “failing faster” – identifying what works and what doesn’t earlier in the process. In the meantime, these assays enable researchers to study complex disease models that increasingly represent human anatomy and physiology more accurately, reducing reliance on animal models. Ultimately, combined with advances in iPSC and organoid technology, I believe bioelectronic assays will help scientists ask and answer new questions, accelerating the development of therapeutics for a range of previously intractable conditions.
- J Zhang et al., Circ Res., 27, e30-41 (2009). DOI: 10.1161/CIRCRESAHA.108.192237
- Y Kishino et al., Inflammation and Regeneration, 40 (2020). DOI: 10.1186/s41232-019-0110-4
- Axion Biosystems, “Measuring oscillatory waves in cerebral organoids,” (2021). Available at https://bit.ly/3sAxIRc
- C Trujillo et al., Cell Stem Cell, 25 (2019). DOI: 10.1016/j.stem.2019.08.002