Microelectrode Array With Transparent ALD TiN Electrodes

Low noise platinum black or sputtered titanium nitride (TiN) microelectrodes are typically used for recording electrical activity of neuronal or cardiac cell cultures. Opaque electrodes and tracks, however, hinder the visibility of the cells when imaged with inverted microscope, which is the standard method of imaging cells plated on microelectrode array (MEA). Even though transparent indium tin oxide (ITO) electrodes exist, they cannot compete in impedance and noise performance with above-mentioned opaque counterparts. In this work, we propose atomic layer deposition (ALD) as the method to deposit TiN electrodes and tracks which are thin enough (25–65 nm) to be transparent (transmission ∼18–45%), but still benefit from the columnar structure of TiN, which is the key element to decrease noise and impedance of the electrodes. For ALD TiN electrodes (diameter 30 μm) impedances from 510 to 590 kΩ were measured at 1 kHz, which is less than the impedance of bare ITO electrodes. Human induced pluripotent stem cell (hiPSC)-derived cortical neurons were cultured on the ALD TiN MEAs for 14 days without observing any biocompatibility issues, and spontaneous electrical activity of the neurons was recorded successfully. The results show that transparent ALD TiN film is a suitable electrode material for producing functional MEAs

Microglia-like Cells Promote Neuronal Functions in Cerebral Organoids

Human cerebral organoids, derived from induced pluripotent stem cells, offer a unique in vitro research window to the development of the cerebral cortex. However, a key player in the developing brain, the microglia, do not natively emerge in cerebral organoids. Here we show that erythromyeloid progenitors (EMPs), differentiated from induced pluripotent stem cells, migrate to cerebral organoids, and mature into microglia-like cells and interact with synaptic material. Patch-clamp electrophysiological recordings show that the microglia-like population supported the emergence of more mature and diversified neuronal phenotypes displaying repetitive firing of action potentials, low-threshold spikes and synaptic activity, while multielectrode array recordings revealed spontaneous bursting activity and increased power of gamma-band oscillations upon pharmacological challenge with NMDA. To conclude, microglia-like cells within the organoids promote neuronal and network maturation and recapitulate some aspects of microglia-neuron co-development in vivo, indicating that cerebral organoids could be a useful biorealistic human in vitro platform for studying microglia-neuron interactions

Microglia-like Cells Promote Neuronal Functions in Cerebral Organoids

Human cerebral organoids, derived from induced pluripotent stem cells, offer a unique in vitro research window to the development of the cerebral cortex. However, a key player in the developing brain, the microglia, do not natively emerge in cerebral organoids. Here we show that erythromyeloid progenitors (EMPs), differentiated from induced pluripotent stem cells, migrate to cerebral organoids, and mature into microglia-like cells and interact with synaptic material. Patch-clamp electrophysiological recordings show that the microglia-like population supported the emergence of more mature and diversified neuronal phenotypes displaying repetitive firing of action potentials, low-threshold spikes and synaptic activity, while multielectrode array recordings revealed spontaneous bursting activity and increased power of gamma-band oscillations upon pharmacological challenge with NMDA. To conclude, microglia-like cells within the organoids promote neuronal and network maturation and recapitulate some aspects of microglia-neuron co-development in vivo, indicating that cerebral organoids could be a useful biorealistic human in vitro platform for studying microglia-neuron interactions.Peer reviewe

A modular brain-on-a-chip for modelling epileptic seizures with functionally connected human neuronal networks

Epilepsies are a group of neurological disorders characterised by recurrent epileptic seizures. Seizures, defined as abnormal transient discharges of neuronal activity, can affect the entire brain circuitry or remain more focal in the specific brain regions and neuronal networks. Human pluripotent stem cell (hPSC)-derived neurons are a promising option for modelling epilepsies, but as such, they do not model groups of connected neuronal networks or focal seizures. Our solution is a Modular Platform for Epilepsy Modelling In Vitro (MEMO), a lab-on-chip device, in which three hPSC-derived networks are separated by a novel microfluidic cell culture device that allows controlled network-to-network axonal connections through microtunnels. In this study, we show that the neuronal networks formed a functional circuitry that was successfully cultured in MEMO for up to 98 days. The spontaneous neuronal network activities were monitored with an integrated custom-made microelectrode array (MEA). The networks developed spontaneous burst activity that was synchronous both within and between the axonally connected networks, i.e. mimicking both local and circuitry functionality of the brain. A convulsant, kainic acid, increased bursts only in the specifically treated networks. The activity reduction by an anticonvulsant, phenytoin, was also localised to treated networks. Therefore, modelling focal seizures in human neuronal networks is now possible with the developed chip.acceptedVersionPeer reviewe

Effect of prolonged differentiation on functional maturation of human pluripotent stem cell-derived neuronal cultures

Long-term neural differentiation of human pluripotent stem cells (hPSCs) is associated with enhanced neuronal maturation, which is a necessity for creation of representative in vitro models. It also induces neurogenic-to-gliogenic fate switch, increasing proportion of endogenous astrocytes formed from the common neural progenitors. However, the significance of prolonged differentiation on the neural cell type composition and functional development of hPSC-derived neuronal cells has not been well characterized. Here, we studied two hPSC lines, both of which initially showed good neuronal differentiation capacity. However, the propensity for endogenous astrogenesis and maturation state after extended differentiation varied. Live cell calcium imaging revealed that prolonged differentiation facilitated maturation of GABAergic signaling. According to extracellular recordings with microelectrode array (MEA), neuronal activity was limited to fewer areas of the culture, which expressed more frequent burst activity. Efficient maturation after prolonged differentiation also promoted organization of spontaneous activity by burst compaction. These results suggest that although prolonged neural differentiation can be challenging, it has beneficial effect on functional maturation, which can also improve transition to different neural in vitro models and applications. Keywords: Astrocytes, Calcium signaling, Microelectrode array, Neural development, Neurons, Pluripotent stem cell

Optimised PDMS Tunnel Devices on MEAs Increase the Probability of Detecting Electrical Activity from Human Stem Cell-Derived Neuronal Networks

Measurement of the activity of human pluripotent stem cell (hPSC)-derived neuronal networks with microelectrode arrays (MEAs) plays an important role in functional in vitro brain modelling and in neurotoxicological screening. The previously reported hPSC-derived neuronal networks do not, however, exhibit repeatable, stable functional network characteristics similar to rodent cortical cultures, making the interpretation of results difficult. In earlier studies, microtunnels have been used both to control and guide cell growth and amplify the axonal signals of rodent neurons. The aim of the current study was to develop tunnel devices that would facilitate signalling and/or signal detection in entire hPSC-derived neuronal networks containing not only axons, but also somata and dendrites. Therefore, MEA-compatible polydimethylsiloxane (PDMS) tunnel devices with 8 different dimensions were created. The hPSC-derived neurons were cultured in the tunnel devices on MEAs, and the spontaneous electrical activity of the networks was measured for 5 weeks. Although the tunnel devices improved the signal-to-noise ratio only by 1.3-fold at best, they significantly increased the percentage of electrodes detecting neuronal activity (52–100%) compared with the controls (27%). Significantly higher spike and burst counts were also obtained using the tunnel devices. Neuronal networks inside the tunnels were amenable to pharmacological manipulation. The results suggest that tunnel devices encompassing the entire neuronal network can increase the measured spontaneous activity in hPSC-derived neuronal networks on MEAs. Therefore, they can increase the efficiency of functional studies of hPSC-derived networks on MEAs

Functional Characterization of Human Pluripotent Stem Cell-Derived Models of the Brain with Microelectrode Arrays

Human pluripotent stem cell (hPSC)-derived neuron cultures have emerged as models of electrical activity in the human brain. Microelectrode arrays (MEAs) measure changes in the extracellular electric potential of cell cultures or tissues and enable the recording of neuronal network activity. MEAs have been applied to both human subjects and hPSC-derived brain models. Here, we review the literature on the functional characterization of hPSC-derived two- and three-dimensional brain models with MEAs and examine their network function in physiological and pathological contexts. We also summarize MEA results from the human brain and compare them to the literature on MEA recordings of hPSC-derived brain models. MEA recordings have shown network activity in two-dimensional hPSC-derived brain models that is comparable to the human brain and revealed pathology-associated changes in disease models. Three-dimensional hPSC-derived models such as brain organoids possess a more relevant microenvironment, tissue architecture and potential for modeling the network activity with more complexity than two-dimensional models. hPSC-derived brain models recapitulate many aspects of network function in the human brain and provide valid disease models, but certain advancements in differentiation methods, bioengineering and available MEA technology are needed for these approaches to reach their full potential