Publications

Selected Publications

High-resolution CMOS MEA platform to study neurons at subcellular, cellular, and network levels

Presenting measurements of neuronal preparations with a novel CMOS-based microelectrode array at high-spatiotemporal-resolution on subcellular, cellular, and network level.

J. Müller, M. Ballini, P. Livi, Y. Chen, M. Radivojevic, A. Shadmani, V. Viswam, I. L. Jones, M. Fiscella, R. Diggelmann, A. Stettler, U. Frey, D. J. Bakkum, and A. Hierlemann, “High-resolution CMOS MEA platform to study neurons at subcellular, cellular, and network levels,” Lab Chip, vol. 15, no. 13, pp. 2767–2780, May 2015.

Revealing Neuronal Function through Microelectrode Array Recordings

Reviewing the current understanding of microelectrode signals and the techniques for analyzing them, with focus on the ongoing advancements in microelectrode technology (in vivo and in vitro) and recent advanced microelectrode array measurement methods that facilitate the understanding of single neurons and network function.

M. E. J. Obien, K. Deligkaris, T. Bullmann, D. J. Bakkum, and U. Frey, “Revealing Neuronal Function through Microelectrode Array Recordings,” Front. Neurosci., 8:423, Jan 2015.

A 1024-Channel CMOS Microelectrode Array With 26,400 Electrodes for Recording and Stimulation of Electrogenic Cells In Vitro

A high-resolution CMOS-based microelectrode array featuring 1,024 low-noise readout channels, 26,400 electrodes at a density of 3,265 electrodes per mm², including on-chip 10bit ADCs and consuming only 75 mW.

M. Ballini, J. Muller, P. Livi, Y. Chen, U. Frey, A. Stettler, A. Shadmani, V. Viswam, I. L. Jones, D. Jackel, M. Radivojevic, M. K. Lewandowska, W. Gong, M. Fiscella, D. J. Bakkum, F. Heer, and A. Hierlemann, “A 1024-Channel CMOS Microelectrode Array With 26,400 Electrodes for Recording and Stimulation of Electrogenic Cells In Vitro,” IEEE Journal of Solid-State Circuits, vol. 49, no. 11, pp. 2705-2719, 2014.

Tracking axonal action potential propagation on a high-density microelectrode array across hundreds of sites

Demonstrating a method to electrically visualize action potential propagation on axons and revealing
large variations in velocity.

D. J. Bakkum, U. Frey, M. Radivojevic, T. L. Russell, J. Muller, M. Fiscella, H. Takahashi, and A. Hierlemann, “Tracking axonal action potential propagation on a high-density microelectrode array across hundreds of sites,” Nature Communications, 4:2181, Jul 2013.

Microelectronic System for High-Resolution Mapping of Extracellular Electric Fields Applied to Brain Slices

Recording and modeling extracellular action potentials of Purkinje cells at subcellular resolution.

U. Frey, U. Egert, F. Heer, S. Hafizovic, and A. Hierlemann, “Microelectronic System for High-Resolution Mapping of Extracellular Electric Fields Applied to Brain Slices,” Biosensors and Bioelectronics, vol. 24, no. 7, pp. 2191-2198, 2009.

Modulation of Cardiomyocyte Electrical Properties Using Regulated Bone Morphogenetic Protein-2 Expression

Controlling BMP-2 expression to modulate the electrophysiological properties of cardiomyocytes using an HD-MEA for detailed monitoring.

C. D. Sanchez-Bustamante, U. Frey, J. M. Kelm, A. Hierlemann, and M. Fussenegger,
“Modulation of Cardiomyocyte Electrical Properties Using Regulated Bone Morphogenetic Protein-2 Expression,” Tissue Engineering Part A, vol. 14, no. 12, pp. 1969-1988, 2008.

All Publications

153 entries « ‹ 1 of 8 › »

1.	Habibollahi, Forough; Kagan, Brett J; Burkitt, Anthony N; French, Chris: Critical dynamics arise during structured information presentation within embodied in vitro neuronal networks. In: Nature Communications, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Habibollahi2023, title = {Critical dynamics arise during structured information presentation within embodied in vitro neuronal networks}, author = {Forough Habibollahi and Brett J. Kagan and Anthony N. Burkitt and Chris French }, url = {https://www.nature.com/articles/s41467-023-41020-3}, doi = {https://doi.org/10.1038/s41467-023-41020-3}, year = {2023}, date = {2023-08-30}, journal = {Nature Communications}, abstract = {Understanding how brains process information is an incredibly difficult task. Amongst the metrics characterising information processing in the brain, observations of dynamic near-critical states have generated significant interest. However, theoretical and experimental limitations associated with human and animal models have precluded a definite answer about when and why neural criticality arises with links from attention, to cognition, and even to consciousness. To explore this topic, we used an in vitro neural network of cortical neurons that was trained to play a simplified game of ‘Pong’ to demonstrate Synthetic Biological Intelligence (SBI). We demonstrate that critical dynamics emerge when neural networks receive task-related structured sensory input, reorganizing the system to a near-critical state. Additionally, better task performance correlated with proximity to critical dynamics. However, criticality alone is insufficient for a neuronal network to demonstrate learning in the absence of additional information regarding the consequences of previous actions. These findings offer compelling support that neural criticality arises as a base feature of incoming structured information processing without the need for higher order cognition.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Understanding how brains process information is an incredibly difficult task. Amongst the metrics characterising information processing in the brain, observations of dynamic near-critical states have generated significant interest. However, theoretical and experimental limitations associated with human and animal models have precluded a definite answer about when and why neural criticality arises with links from attention, to cognition, and even to consciousness. To explore this topic, we used an in vitro neural network of cortical neurons that was trained to play a simplified game of ‘Pong’ to demonstrate Synthetic Biological Intelligence (SBI). We demonstrate that critical dynamics emerge when neural networks receive task-related structured sensory input, reorganizing the system to a near-critical state. Additionally, better task performance correlated with proximity to critical dynamics. However, criticality alone is insufficient for a neuronal network to demonstrate learning in the absence of additional information regarding the consequences of previous actions. These findings offer compelling support that neural criticality arises as a base feature of incoming structured information processing without the need for higher order cognition. Close https://www.nature.com/articles/s41467-023-41020-3 doi:https://doi.org/10.1038/s41467-023-41020-3 Close
2.	Radivojevic, Milos; Punga, Anna Rostedt: Functional imaging of conduction dynamics in cortical and spinal axons. In: eLife, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Radivojevic2023_2, title = {Functional imaging of conduction dynamics in cortical and spinal axons}, author = {Milos Radivojevic and Anna Rostedt Punga}, url = {https://elifesciences.org/articles/86512}, doi = {https://doi.org/10.7554/eLife.86512}, year = {2023}, date = {2023-08-22}, journal = {eLife}, abstract = {Mammalian axons are specialized for transmitting action potentials to targets within the central and peripheral nervous system. A growing body of evidence suggests that, besides signal conduction, axons play essential roles in neural information processing, and their malfunctions are common hallmarks of neurodegenerative diseases. The technologies available to study axonal function and structure integrally limit the comprehension of axon neurobiology. High-density microelectrode arrays (HD-MEAs) allow for accessing axonal action potentials at high spatiotemporal resolution, but provide no insights on axonal morphology. Here, we demonstrate a method for electrical visualization of axonal morphologies based on extracellular action potentials recorded from cortical and motor neurons using HD-MEAs. The method enabled us to reconstruct up to 5-cm-long axonal arbors and directly monitor axonal conduction across thousands of recording sites. We reconstructed 1.86 m of cortical and spinal axons in total and found specific features in their structure and function.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Mammalian axons are specialized for transmitting action potentials to targets within the central and peripheral nervous system. A growing body of evidence suggests that, besides signal conduction, axons play essential roles in neural information processing, and their malfunctions are common hallmarks of neurodegenerative diseases. The technologies available to study axonal function and structure integrally limit the comprehension of axon neurobiology. High-density microelectrode arrays (HD-MEAs) allow for accessing axonal action potentials at high spatiotemporal resolution, but provide no insights on axonal morphology. Here, we demonstrate a method for electrical visualization of axonal morphologies based on extracellular action potentials recorded from cortical and motor neurons using HD-MEAs. The method enabled us to reconstruct up to 5-cm-long axonal arbors and directly monitor axonal conduction across thousands of recording sites. We reconstructed 1.86 m of cortical and spinal axons in total and found specific features in their structure and function. Close https://elifesciences.org/articles/86512 doi:https://doi.org/10.7554/eLife.86512 Close
3.	Duru, Jens; Maurer, Benedikt; Doran, Ciara Giles; Jelitto, Robert; Küchler, Joël; Ihle, Stephan J; Ruff, Tobias; John, Robert; Genocchi, Barbara; Vörös, János: Investigation of the input-output relationship of engineered neural networks using high-density microelectrode arrays. In: Biosensors and Bioelectronics, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Duru2023b, title = {Investigation of the input-output relationship of engineered neural networks using high-density microelectrode arrays}, author = {Jens Duru and Benedikt Maurer and Ciara Giles Doran and Robert Jelitto and Joël Küchler and Stephan J. Ihle and Tobias Ruff and Robert John and Barbara Genocchi and János Vörös }, url = {https://www.sciencedirect.com/science/article/pii/S095656632300533X?via%3Dihub}, doi = {https://doi.org/10.1016/j.bios.2023.115591}, year = {2023}, date = {2023-08-18}, journal = {Biosensors and Bioelectronics}, abstract = {Bottom-up neuroscience utilizes small, engineered biological neural networks to study neuronal activity in systems of reduced complexity. We present a platform that establishes up to six independent networks formed by primary rat neurons on planar complementary metal–oxide–semiconductor (CMOS) microelectrode arrays (MEAs). We introduce an approach that allows repetitive stimulation and recording of network activity at any of the over 700 electrodes underlying a network. We demonstrate that the continuous application of a repetitive super-threshold stimulus yields a reproducible network answer within a 15 ms post-stimulus window. This response can be tracked with high spatiotemporal resolution across the whole extent of the network. Moreover, we show that the location of the stimulation plays a significant role in the networks' early response to the stimulus. By applying a stimulation pattern to all network-underlying electrodes in sequence, the sensitivity of the whole network to the stimulus can be visualized. We demonstrate that microchannels reduce the voltage stimulation threshold and induce the strongest network response. By varying the stimulation amplitude and frequency we reveal discrete network transition points. Finally, we introduce vector fields to follow stimulation-induced spike propagation pathways within the network. Overall we show that our defined neural networks on CMOS MEAs enable us to elicit highly reproducible activity patterns that can be precisely modulated by stimulation amplitude, stimulation frequency and the site of stimulation.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Bottom-up neuroscience utilizes small, engineered biological neural networks to study neuronal activity in systems of reduced complexity. We present a platform that establishes up to six independent networks formed by primary rat neurons on planar complementary metal–oxide–semiconductor (CMOS) microelectrode arrays (MEAs). We introduce an approach that allows repetitive stimulation and recording of network activity at any of the over 700 electrodes underlying a network. We demonstrate that the continuous application of a repetitive super-threshold stimulus yields a reproducible network answer within a 15 ms post-stimulus window. This response can be tracked with high spatiotemporal resolution across the whole extent of the network. Moreover, we show that the location of the stimulation plays a significant role in the networks' early response to the stimulus. By applying a stimulation pattern to all network-underlying electrodes in sequence, the sensitivity of the whole network to the stimulus can be visualized. We demonstrate that microchannels reduce the voltage stimulation threshold and induce the strongest network response. By varying the stimulation amplitude and frequency we reveal discrete network transition points. Finally, we introduce vector fields to follow stimulation-induced spike propagation pathways within the network. Overall we show that our defined neural networks on CMOS MEAs enable us to elicit highly reproducible activity patterns that can be precisely modulated by stimulation amplitude, stimulation frequency and the site of stimulation. Close https://www.sciencedirect.com/science/article/pii/S095656632300533X?via%3Dihub doi:https://doi.org/10.1016/j.bios.2023.115591 Close
4.	Zhao, Eric T; Hull, Jacob M; Hemed, Nofar Mintz; Ulusan, Hasan; Bartram, Julian; Zhang, Anqi; Wang, Pingyu; Pham, Albert; Silvia Ronchi, John Huguenard R; Hierlemann, Andreas; Melosh, Nicholas A: A CMOS-based highly scalable flexible neural electrode interface. In: Science Advances, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Zhao2023, title = {A CMOS-based highly scalable flexible neural electrode interface}, author = {Eric T. Zhao and Jacob M. Hull and Nofar Mintz Hemed and Hasan Ulusan and Julian Bartram and Anqi Zhang and Pingyu Wang and Albert Pham and Silvia Ronchi, John R. Huguenard and Andreas Hierlemann and Nicholas A. Melosh}, url = {https://www.science.org/doi/10.1126/sciadv.adf9524}, doi = {DOI: 10.1126/sciadv.adf9524}, year = {2023}, date = {2023-06-07}, journal = {Science Advances}, abstract = {Perception, thoughts, and actions are encoded by the coordinated activity of large neuronal populations spread over large areas. However, existing electrophysiological devices are limited by their scalability in capturing this cortex-wide activity. Here, we developed an electrode connector based on an ultra-conformable thin-film electrode array that self-assembles onto silicon microelectrode arrays enabling multithousand channel counts at a millimeter scale. The interconnects are formed using microfabricated electrode pads suspended by thin support arms, termed Flex2Chip. Capillary-assisted assembly drives the pads to deform toward the chip surface, and van der Waals forces maintain this deformation, establishing Ohmic contact. Flex2Chip arrays successfully measured extracellular action potentials ex vivo and resolved micrometer scale seizure propagation trajectories in epileptic mice. We find that seizure dynamics in absence epilepsy in the Scn8a+/− model do not have constant propagation trajectories.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Perception, thoughts, and actions are encoded by the coordinated activity of large neuronal populations spread over large areas. However, existing electrophysiological devices are limited by their scalability in capturing this cortex-wide activity. Here, we developed an electrode connector based on an ultra-conformable thin-film electrode array that self-assembles onto silicon microelectrode arrays enabling multithousand channel counts at a millimeter scale. The interconnects are formed using microfabricated electrode pads suspended by thin support arms, termed Flex2Chip. Capillary-assisted assembly drives the pads to deform toward the chip surface, and van der Waals forces maintain this deformation, establishing Ohmic contact. Flex2Chip arrays successfully measured extracellular action potentials ex vivo and resolved micrometer scale seizure propagation trajectories in epileptic mice. We find that seizure dynamics in absence epilepsy in the Scn8a+/− model do not have constant propagation trajectories. Close https://www.science.org/doi/10.1126/sciadv.adf9524 doi:DOI: 10.1126/sciadv.adf9524 Close
5.	Girardi, Gregory; Zumpano, Danielle; Goshi, Noah; Raybould, Helen; Seker, Erkin: Cultured Vagal Afferent Neurons as Sensors for Intestinal Effector Molecules. In: biosensors, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Girardi2023, title = {Cultured Vagal Afferent Neurons as Sensors for Intestinal Effector Molecules}, author = {Gregory Girardi and Danielle Zumpano and Noah Goshi and Helen Raybould and Erkin Seker}, url = {https://www.mdpi.com/2079-6374/13/6/601}, doi = {10.3390/bios13060601}, year = {2023}, date = {2023-05-31}, journal = {biosensors}, abstract = {The gut–brain axis embodies the bi-directional communication between the gastrointestinal tract and the central nervous system (CNS), where vagal afferent neurons (VANs) serve as sensors for a variety of gut-derived signals. The gut is colonized by a large and diverse population of microorganisms that communicate via small (effector) molecules, which also act on the VAN terminals situated in the gut viscera and consequently influence many CNS processes. However, the convoluted in vivo environment makes it difficult to study the causative impact of the effector molecules on VAN activation or desensitization. Here, we report on a VAN culture and its proof-of-principle demonstration as a cell-based sensor to monitor the influence of gastrointestinal effector molecules on neuronal behavior. We initially compared the effect of surface coatings (poly-L-lysine vs. Matrigel) and culture media composition (serum vs. growth factor supplement) on neurite growth as a surrogate of VAN regeneration following tissue harvesting, where the Matrigel coating, but not the media composition, played a significant role in the increased neurite growth. We then used both live-cell calcium imaging and extracellular electrophysiological recordings to show that the VANs responded to classical effector molecules of endogenous and exogenous origin (cholecystokinin serotonin and capsaicin) in a complex fashion. We expect this study to enable platforms for screening various effector molecules and their influence on VAN activity, assessed by their information-rich electrophysiological fingerprints.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close The gut–brain axis embodies the bi-directional communication between the gastrointestinal tract and the central nervous system (CNS), where vagal afferent neurons (VANs) serve as sensors for a variety of gut-derived signals. The gut is colonized by a large and diverse population of microorganisms that communicate via small (effector) molecules, which also act on the VAN terminals situated in the gut viscera and consequently influence many CNS processes. However, the convoluted in vivo environment makes it difficult to study the causative impact of the effector molecules on VAN activation or desensitization. Here, we report on a VAN culture and its proof-of-principle demonstration as a cell-based sensor to monitor the influence of gastrointestinal effector molecules on neuronal behavior. We initially compared the effect of surface coatings (poly-L-lysine vs. Matrigel) and culture media composition (serum vs. growth factor supplement) on neurite growth as a surrogate of VAN regeneration following tissue harvesting, where the Matrigel coating, but not the media composition, played a significant role in the increased neurite growth. We then used both live-cell calcium imaging and extracellular electrophysiological recordings to show that the VANs responded to classical effector molecules of endogenous and exogenous origin (cholecystokinin serotonin and capsaicin) in a complex fashion. We expect this study to enable platforms for screening various effector molecules and their influence on VAN activity, assessed by their information-rich electrophysiological fingerprints. Close https://www.mdpi.com/2079-6374/13/6/601 doi:10.3390/bios13060601 Close
6.	Bartram, Julian; Franke, Felix; Kumar, Sreedhar Saseendran; Buccino, Alessio Paolo; Xue, Xiaohan; Gänswein, Tobias; Schröter, Manuel; Kim, Taehoon; Kasuba, Krishna Chaitanya; Hierlemann, Andreas: Parallel reconstruction of the excitatory and inhibitory inputs received by single neurons reveals the synaptic basis of recurrent spiking. In: eLife, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Bartram2023b, title = {Parallel reconstruction of the excitatory and inhibitory inputs received by single neurons reveals the synaptic basis of recurrent spiking}, author = {Julian Bartram and Felix Franke and Sreedhar Saseendran Kumar and Alessio Paolo Buccino and Xiaohan Xue and Tobias Gänswein and Manuel Schröter and Taehoon Kim and Krishna Chaitanya Kasuba and Andreas Hierlemann}, url = {https://elifesciences.org/reviewed-preprints/86820}, doi = {10.7554/eLife.86820}, year = {2023}, date = {2023-05-17}, journal = {eLife}, abstract = {Self-sustained recurrent activity in cortical networks is thought to be important for multiple crucial processes, including circuit development and homeostasis. Yet, the precise relationship between the synaptic input patterns and the spiking output of individual neurons remains largely unresolved. Here, we developed, validated and applied a novel in vitro experimental platform and analytical procedures that provide – for individual neurons – simultaneous excitatory and inhibitory synaptic activity estimates during recurrent network activity. Our approach combines whole-network high-density microelectrode array (HD-MEA) recordings from rat neuronal cultures with patch clamping and enables a comprehensive mapping and characterization of active incoming connections to single postsynaptic neurons. We found that, during network states with excitation(E)-inhibition(I) balance, postsynaptic spiking coincided precisely with the maxima of fast fluctuations in the input E/I ratio. These spike-associated E/I ratio escalations were largely due to a rapid bidirectional change in synaptic inhibition that was modulated by the network-activity level. Our approach also uncovered the underlying circuit architecture and we show that individual neurons received a few key inhibitory connections – often from special hub neurons – that were instrumental in controlling postsynaptic spiking. Balanced network theory predicts dynamical regimes governed by small and rapid input fluctuation and featuring a fast neuronal responsiveness. Our findings – obtained in self-organized neuronal cultures – suggest that the emergence of these favorable regimes and associated network architectures is an inherent property of cortical networks in general.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Self-sustained recurrent activity in cortical networks is thought to be important for multiple crucial processes, including circuit development and homeostasis. Yet, the precise relationship between the synaptic input patterns and the spiking output of individual neurons remains largely unresolved. Here, we developed, validated and applied a novel in vitro experimental platform and analytical procedures that provide – for individual neurons – simultaneous excitatory and inhibitory synaptic activity estimates during recurrent network activity. Our approach combines whole-network high-density microelectrode array (HD-MEA) recordings from rat neuronal cultures with patch clamping and enables a comprehensive mapping and characterization of active incoming connections to single postsynaptic neurons. We found that, during network states with excitation(E)-inhibition(I) balance, postsynaptic spiking coincided precisely with the maxima of fast fluctuations in the input E/I ratio. These spike-associated E/I ratio escalations were largely due to a rapid bidirectional change in synaptic inhibition that was modulated by the network-activity level. Our approach also uncovered the underlying circuit architecture and we show that individual neurons received a few key inhibitory connections – often from special hub neurons – that were instrumental in controlling postsynaptic spiking. Balanced network theory predicts dynamical regimes governed by small and rapid input fluctuation and featuring a fast neuronal responsiveness. Our findings – obtained in self-organized neuronal cultures – suggest that the emergence of these favorable regimes and associated network architectures is an inherent property of cortical networks in general. Close https://elifesciences.org/reviewed-preprints/86820 doi:10.7554/eLife.86820 Close
7.	Duru, Jens; Maurer, Benedikt; Doran, Ciara Giles; Jelitto, Robert; Küchler, Joël; Ihle, Stephan J; Ruff, Tobias; John, Robert; Genocchi, Barbara; Vörös, János: Investigation of the input-output relationship of engineered neural networks using high-density microelectrode arrays. In: SSRN, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Duru2023, title = {Investigation of the input-output relationship of engineered neural networks using high-density microelectrode arrays}, author = {Jens Duru and Benedikt Maurer and Ciara Giles Doran and Robert Jelitto and Joël Küchler and Stephan J. Ihle and Tobias Ruff and Robert John and Barbara Genocchi and János Vörös}, url = {https://www.ssrn.com/abstract=4427959}, doi = {DOI: 10.2139/ssrn.4427959}, year = {2023}, date = {2023-04-24}, journal = {SSRN}, abstract = {Bottom-up neuroscience utilizes small, engineered biological neural networks to study neuronal activity in systems of reduced complexity. We present a platform that establishes up to six independent networks formed by primary rat neurons on planar complementary metal–oxide–semiconductor (CMOS) microelectrode arrays (MEAs). We introduce an approach that allows repetitive stimulation and recording of network activity at any of the over 700 electrodes underlying a network. We demonstrate that the continuous application of a repetitive super-threshold stimulus yields a reproducible network answer within a 15 ms post-stimulus window. This response can be tracked with high spatiotemporal resolution across the whole extent of the network. Moreover, we show that the location of the stimulation plays a significant role in the networks’ early response to the stimulus. By applying a stimulation pattern to all network-underlying electrodes in sequence, the sensitivity of the whole network to the stimulus can be visualized. We demonstrate that microchannels reduce the voltage stimulation threshold and induce the strongest network response. By varying the stimulation amplitude and frequency we reveal discrete network transition points. Finally, we introduce vector fields to follow stimulation-induced spike propagation pathways within the network. Overall we show that our defined neural networks on CMOS MEAs enable us to elicit highly reproducible activity patterns that can be precisely modulated by stimulation amplitude, stimulation frequency and the site of stimulation.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Bottom-up neuroscience utilizes small, engineered biological neural networks to study neuronal activity in systems of reduced complexity. We present a platform that establishes up to six independent networks formed by primary rat neurons on planar complementary metal–oxide–semiconductor (CMOS) microelectrode arrays (MEAs). We introduce an approach that allows repetitive stimulation and recording of network activity at any of the over 700 electrodes underlying a network. We demonstrate that the continuous application of a repetitive super-threshold stimulus yields a reproducible network answer within a 15 ms post-stimulus window. This response can be tracked with high spatiotemporal resolution across the whole extent of the network. Moreover, we show that the location of the stimulation plays a significant role in the networks’ early response to the stimulus. By applying a stimulation pattern to all network-underlying electrodes in sequence, the sensitivity of the whole network to the stimulus can be visualized. We demonstrate that microchannels reduce the voltage stimulation threshold and induce the strongest network response. By varying the stimulation amplitude and frequency we reveal discrete network transition points. Finally, we introduce vector fields to follow stimulation-induced spike propagation pathways within the network. Overall we show that our defined neural networks on CMOS MEAs enable us to elicit highly reproducible activity patterns that can be precisely modulated by stimulation amplitude, stimulation frequency and the site of stimulation. Close https://www.ssrn.com/abstract=4427959 doi:DOI: 10.2139/ssrn.4427959 Close
8.	Xu, He Jax; Yao, Yao; Yao, Fenyong; Chen, Jiehui; Li, Meishi; Yang, Xianfa; Li, Sheng; Lu, Fangru; Hu, Ping; He, Shuijin; Peng, Guangdun; Jing, Naihe: Generation of functional posterior spinal motor neurons from hPSCs-derived human spinal cord neural progenitor cells. In: Cell Regeneration, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Xu2023, title = {Generation of functional posterior spinal motor neurons from hPSCs-derived human spinal cord neural progenitor cells}, author = {He Jax Xu and Yao Yao and Fenyong Yao and Jiehui Chen and Meishi Li and Xianfa Yang and Sheng Li and Fangru Lu and Ping Hu and Shuijin He and Guangdun Peng and Naihe Jing}, url = {https://cellregeneration.springeropen.com/articles/10.1186/s13619-023-00159-6}, doi = {10.1186/s13619-023-00159-6}, year = {2023}, date = {2023-03-23}, journal = {Cell Regeneration}, abstract = {Spinal motor neurons deficiency results in a series of devastating disorders such as amyotrophic lateral sclerosis (ALS), spinal muscular atrophy (SMA) and spinal cord injury (SCI). These disorders are currently incurable, while human pluripotent stem cells (hPSCs)-derived spinal motor neurons are promising but suffered from inappropriate regional identity and functional immaturity for the study and treatment of posterior spinal cord related injuries. In this study, we have established human spinal cord neural progenitor cells (hSCNPCs) via hPSCs differentiated neuromesodermal progenitors (NMPs) and demonstrated the hSCNPCs can be continuously expanded up to 40 passages. hSCNPCs can be rapidly differentiated into posterior spinal motor neurons with high efficiency. The functional maturity has been examined in detail. Moreover, a co-culture scheme which is compatible for both neural and muscular differentiation is developed to mimic the neuromuscular junction (NMJ) formation in vitro. Together, these studies highlight the potential avenues for generating clinically relevant spinal motor neurons and modeling neuromuscular diseases through our defined hSCNPCs.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Spinal motor neurons deficiency results in a series of devastating disorders such as amyotrophic lateral sclerosis (ALS), spinal muscular atrophy (SMA) and spinal cord injury (SCI). These disorders are currently incurable, while human pluripotent stem cells (hPSCs)-derived spinal motor neurons are promising but suffered from inappropriate regional identity and functional immaturity for the study and treatment of posterior spinal cord related injuries. In this study, we have established human spinal cord neural progenitor cells (hSCNPCs) via hPSCs differentiated neuromesodermal progenitors (NMPs) and demonstrated the hSCNPCs can be continuously expanded up to 40 passages. hSCNPCs can be rapidly differentiated into posterior spinal motor neurons with high efficiency. The functional maturity has been examined in detail. Moreover, a co-culture scheme which is compatible for both neural and muscular differentiation is developed to mimic the neuromuscular junction (NMJ) formation in vitro. Together, these studies highlight the potential avenues for generating clinically relevant spinal motor neurons and modeling neuromuscular diseases through our defined hSCNPCs. Close https://cellregeneration.springeropen.com/articles/10.1186/s13619-023-00159-6 doi:10.1186/s13619-023-00159-6 Close
9.	Radivojevic, Milos; Punga, Anna Rostedt: Functional imaging of conduction dynamics in cortical and spinal axons. In: BioRxiv, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Radivojevic2023, title = {Functional imaging of conduction dynamics in cortical and spinal axons}, author = {Milos Radivojevic and Anna Rostedt Punga}, url = {https://www.biorxiv.org/content/10.1101/2023.02.28.530461v1}, doi = {https://doi.org/10.1101/2023.02.28.530461}, year = {2023}, date = {2023-03-01}, journal = {BioRxiv}, abstract = {Mammalian axons are specialized for transmitting action potentials to targets within the central and peripheral nervous system. A growing body of evidence suggests that, besides signal conduction, axons play essential roles in neural information processing, and their malfunctions are common hallmarks of neurodegenerative diseases. The technologies available to study axonal function and structure integrally limit the comprehension of axon neurobiology. High-density microelectrode arrays (HD-MEAs) allow for accessing axonal action potentials at high spatiotemporal resolution, but provide no insights on axonal morphology. Here we demonstrate a method for electrical visualization of axonal morphologies based on extracellular action potentials recorded from cortical and motor neurons using HD-MEAs. The method enabled us to reconstruct up to 5-centimeter-long axonal arbors and directly monitor axonal conduction across thousands of recording sites. We reconstructed 1.86 meters of cortical and spinal axons in total and found specific features in their structure and function.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Mammalian axons are specialized for transmitting action potentials to targets within the central and peripheral nervous system. A growing body of evidence suggests that, besides signal conduction, axons play essential roles in neural information processing, and their malfunctions are common hallmarks of neurodegenerative diseases. The technologies available to study axonal function and structure integrally limit the comprehension of axon neurobiology. High-density microelectrode arrays (HD-MEAs) allow for accessing axonal action potentials at high spatiotemporal resolution, but provide no insights on axonal morphology. Here we demonstrate a method for electrical visualization of axonal morphologies based on extracellular action potentials recorded from cortical and motor neurons using HD-MEAs. The method enabled us to reconstruct up to 5-centimeter-long axonal arbors and directly monitor axonal conduction across thousands of recording sites. We reconstructed 1.86 meters of cortical and spinal axons in total and found specific features in their structure and function. Close https://www.biorxiv.org/content/10.1101/2023.02.28.530461v1 doi:https://doi.org/10.1101/2023.02.28.530461 Close
10.	Cai, Hongwei; Ao, Zheng; Tian, Chunhui; Wu, Zhuhao; Liu, Hongcheng; Tchieu, Jason; Gu, Mingxia; Mackie, Ken; and Guo, Feng: Brain Organoid Computing for Artificial Intelligence. In: bioRxiv, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Cai2023, title = {Brain Organoid Computing for Artificial Intelligence}, author = {Hongwei Cai and Zheng Ao and Chunhui Tian and Zhuhao Wu and Hongcheng Liu and Jason Tchieu and Mingxia Gu and Ken Mackie and and Feng Guo}, url = {https://www.biorxiv.org/content/10.1101/2023.02.28.530502v1}, doi = {10.1101/2023.02.28.530502}, year = {2023}, date = {2023-03-01}, journal = {bioRxiv}, abstract = {Brain-inspired hardware emulates the structure and working principles of a biological brain and may address the hardware bottleneck for fast-growing artificial intelligence (AI). Current brain-inspired silicon chips are promising but still limit their power to fully mimic brain function for AI computing. Here, we develop Brainoware, living AI hardware that harnesses the computation power of 3D biological neural networks in a brain organoid. Brain-like 3D in vitro cultures compute by receiving and sending information via a multielectrode array. Applying spatiotemporal electrical stimulation, this approach not only exhibits nonlinear dynamics and fading memory properties but also learns from training data. Further experiments demonstrate real-world applications in solving non-linear equations. This approach may provide new insights into AI hardware. }, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Brain-inspired hardware emulates the structure and working principles of a biological brain and may address the hardware bottleneck for fast-growing artificial intelligence (AI). Current brain-inspired silicon chips are promising but still limit their power to fully mimic brain function for AI computing. Here, we develop Brainoware, living AI hardware that harnesses the computation power of 3D biological neural networks in a brain organoid. Brain-like 3D in vitro cultures compute by receiving and sending information via a multielectrode array. Applying spatiotemporal electrical stimulation, this approach not only exhibits nonlinear dynamics and fading memory properties but also learns from training data. Further experiments demonstrate real-world applications in solving non-linear equations. This approach may provide new insights into AI hardware. Close https://www.biorxiv.org/content/10.1101/2023.02.28.530502v1 doi:10.1101/2023.02.28.530502 Close
11.	Lin, Waka; Shiomoto, Shusaku; Yamada, Saki; Watanabe, Hikaru; Kawashima, Yudai; Eguchi, Yuichi; Muramatsu, Koichi; Sekino, Yuko: Dendritic spine formation and synapse maturation in transcription factor-induced human iPSC-derived neurons. In: iScience, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Lin2023, title = {Dendritic spine formation and synapse maturation in transcription factor-induced human iPSC-derived neurons}, author = {Waka Lin and Shusaku Shiomoto and Saki Yamada and Hikaru Watanabe and Yudai Kawashima and Yuichi Eguchi and Koichi Muramatsu and Yuko Sekino}, url = {https://pubmed.ncbi.nlm.nih.gov/37034988/}, year = {2023}, date = {2023-02-27}, journal = {iScience}, abstract = {Synaptic maturation is reportedly limited in human induced pluripotent stem cell (iPSC)-derived neurons. Notably, their ability to reach postnatal-like stages and form dendritic spines has been difficult to demonstrate unless using long-term cultured organoids. Recent transcription factor (TF)-based induction methods allow the accelerated generation of differentiated neurons, which offers an unprecedented opportunity to address further progression into late developmental stages. Herein, we report on a comprehensive time-course study of TF-induced iPSC neurons cultured in vitro through an intrinsic maturation program following neurogenesis. Moreover, we determined the transcriptional and morphological sequences of key developmental events associated with spinogenesis, including the conversion of drebrin to its brain-specific isoform A and the N-methyl-D-aspartate (NMDA) receptor subunit switch. TF-induced iPSC neurons successfully acquired structural and functional synaptic maturity, which will critically expand their utility in modeling higher brain functions and disorders.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Synaptic maturation is reportedly limited in human induced pluripotent stem cell (iPSC)-derived neurons. Notably, their ability to reach postnatal-like stages and form dendritic spines has been difficult to demonstrate unless using long-term cultured organoids. Recent transcription factor (TF)-based induction methods allow the accelerated generation of differentiated neurons, which offers an unprecedented opportunity to address further progression into late developmental stages. Herein, we report on a comprehensive time-course study of TF-induced iPSC neurons cultured in vitro through an intrinsic maturation program following neurogenesis. Moreover, we determined the transcriptional and morphological sequences of key developmental events associated with spinogenesis, including the conversion of drebrin to its brain-specific isoform A and the N-methyl-D-aspartate (NMDA) receptor subunit switch. TF-induced iPSC neurons successfully acquired structural and functional synaptic maturity, which will critically expand their utility in modeling higher brain functions and disorders. Close https://pubmed.ncbi.nlm.nih.gov/37034988/ Close
12.	Qian, Junming; Guan, Xiaonan; Xie, Bing; Xu, Chuanyun; Niu, Jacqueline; Tang, Xin; Li, Charles H; Colecraft, Henry M; Jaenisch, Rudolf; Liu, Shawn X: Multiplex epigenome editing of MECP2 to rescue Rett syndrome neurons. In: Science Translational Medicine, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Qian2023, title = {Multiplex epigenome editing of MECP2 to rescue Rett syndrome neurons}, author = {Junming Qian and Xiaonan Guan and Bing Xie and Chuanyun Xu and Jacqueline Niu and Xin Tang and Charles H. Li and Henry M. Colecraft and Rudolf Jaenisch and X. Shawn Liu}, url = {https://www.science.org/doi/10.1126/scitranslmed.add4666}, doi = {10.1126/scitranslmed.add4666}, year = {2023}, date = {2023-01-18}, journal = {Science Translational Medicine}, abstract = {Rettsyndrome(RTT)isanX-linkedneurodevelopmental disorder caused byloss-of-function heterozygous mutationsofmethyl CpG-binding protein2(MECP2) ontheXchromosome inyoungfemales. Reactivationofthe silent wild-type MECP2 allelefromtheinactiveXchromosome (Xi)represents apromising therapeutic opportunity forfemale patients withRTT.Here,weapplied amultiple xepigenome editing approachtoreactivate MECP2 fromXiinRTThuman embryonicstemcells(hESCs) andderivedneurons.Demethyla tionofthe MECP2 promoter bydCas9-T et1withtarget single-guide RNAreactivatedMECP2 fromXiinRTThESCs without detectable off-target effects atthetranscriptional level.Neuronsderivedfrommethyla tion-edited RTThESCs maintained MECP2 reactivationandreversedthesmaller somasizeandelectrophysiological abnormalities, twohallmarks ofRTT.InRTTneurons,insulationofthemethyla tion-edited MECP2 locusbydCpf1-CT CF (acatalytically deadCpf1fusedwithCCCTC-binding factor)withtarget CRISPR RNAenhanced MECP2 reactivationandrescued RTT-relatedneuronaldefects, providing aproof-of-concept studyforepigenome editing to treatRTTandpotentially otherdominant X-linkeddiseases.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Rettsyndrome(RTT)isanX-linkedneurodevelopmental disorder caused byloss-of-function heterozygous mutationsofmethyl CpG-binding protein2(MECP2) ontheXchromosome inyoungfemales. Reactivationofthe silent wild-type MECP2 allelefromtheinactiveXchromosome (Xi)represents apromising therapeutic opportunity forfemale patients withRTT.Here,weapplied amultiple xepigenome editing approachtoreactivate MECP2 fromXiinRTThuman embryonicstemcells(hESCs) andderivedneurons.Demethyla tionofthe MECP2 promoter bydCas9-T et1withtarget single-guide RNAreactivatedMECP2 fromXiinRTThESCs without detectable off-target effects atthetranscriptional level.Neuronsderivedfrommethyla tion-edited RTThESCs maintained MECP2 reactivationandreversedthesmaller somasizeandelectrophysiological abnormalities, twohallmarks ofRTT.InRTTneurons,insulationofthemethyla tion-edited MECP2 locusbydCpf1-CT CF (acatalytically deadCpf1fusedwithCCCTC-binding factor)withtarget CRISPR RNAenhanced MECP2 reactivationandrescued RTT-relatedneuronaldefects, providing aproof-of-concept studyforepigenome editing to treatRTTandpotentially otherdominant X-linkeddiseases. Close https://www.science.org/doi/10.1126/scitranslmed.add4666 doi:10.1126/scitranslmed.add4666 Close
13.	Kim, Eunhee; Jeon, Sungwoong; Yang, Yoon-Sil; Jin, Chaewon; Kim, Jin-young; Oh, Yong- Seok; Rah, Jong-Cheol; and Choi, Hongsoo: A Neurospheroid-Based Microrobot for Targeted Neural Connections in a Hippocampal Slice. In: Advanced Materials, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{EunheeKim2023, title = {A Neurospheroid-Based Microrobot for Targeted Neural Connections in a Hippocampal Slice}, author = {Eunhee Kim and Sungwoong Jeon and Yoon-Sil Yang and Chaewon Jin and Jin-young Kim and Yong- Seok Oh and Jong-Cheol Rah and and Hongsoo Choi}, url = {https://onlinelibrary.wiley.com/doi/10.1002/adma.202208747?af=R}, doi = {https://doi.org/10.1002/adma.202208747}, year = {2023}, date = {2023-01-14}, journal = {Advanced Materials}, abstract = {Functional restoration by the re-establishment of cellular or neural connections remains a major challenge in targeted cell therapy and regenerative medicine. Recent advances in magnetically powered microrobots have shown potential for use in controlled and targeted cell therapy. In this study, a magnetic neurospheroid (Mag-Neurobot) that could form both structural and functional connections with an organotypic hippocampal slice (OHS) was assessed using an ex vivo model as a bridge toward in vivo application. The Mag-Neurobot consists of hippocampal neurons and superparamagnetic nanoparticles (SPIONs); it is precisely and skillfully manipulated by an external magnetic field. Furthermore, the results of patch-clamp recordings of hippocampal neurons indicated that neither the neuronal excitabilities nor the synaptic functions of SPION-loaded cells were significantly affected. Analysis of neural activity propagation using high-density multi-electrode arrays showed that the delivered Mag-Neurobot was functionally connected with the OHS. The applications of this study include functional verification for targeted cell delivery through the characterization of novel synaptic connections and the functionalities of transported and transplanted cells. The success of the Mag-Neurobot opens up new avenues of research and application; it offers a test platform for functional neural connections and neural regenerative processes through cell transplantation.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Functional restoration by the re-establishment of cellular or neural connections remains a major challenge in targeted cell therapy and regenerative medicine. Recent advances in magnetically powered microrobots have shown potential for use in controlled and targeted cell therapy. In this study, a magnetic neurospheroid (Mag-Neurobot) that could form both structural and functional connections with an organotypic hippocampal slice (OHS) was assessed using an ex vivo model as a bridge toward in vivo application. The Mag-Neurobot consists of hippocampal neurons and superparamagnetic nanoparticles (SPIONs); it is precisely and skillfully manipulated by an external magnetic field. Furthermore, the results of patch-clamp recordings of hippocampal neurons indicated that neither the neuronal excitabilities nor the synaptic functions of SPION-loaded cells were significantly affected. Analysis of neural activity propagation using high-density multi-electrode arrays showed that the delivered Mag-Neurobot was functionally connected with the OHS. The applications of this study include functional verification for targeted cell delivery through the characterization of novel synaptic connections and the functionalities of transported and transplanted cells. The success of the Mag-Neurobot opens up new avenues of research and application; it offers a test platform for functional neural connections and neural regenerative processes through cell transplantation. Close https://onlinelibrary.wiley.com/doi/10.1002/adma.202208747?af=R doi:https://doi.org/10.1002/adma.202208747 Close
14.	Kim, Taehoon; Chen, Dexiong; Hornauer, Philipp; Emmenegger, Vishalini; Bartram, Julian; Ronchi, Silvia; Hierlemann, Andreas; Schröter, Manuel; Roqueiro, Damian: Predicting in vitro single-neuron firing rates upon pharmacological perturbation using Graph Neural Networks. In: Frontiers in Neuroinformatics, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Taehoon2023, title = {Predicting in vitro single-neuron firing rates upon pharmacological perturbation using Graph Neural Networks}, author = {Taehoon Kim and Dexiong Chen and Philipp Hornauer and Vishalini Emmenegger and Julian Bartram and Silvia Ronchi and Andreas Hierlemann and Manuel Schröter and Damian Roqueiro}, url = {https://www.frontiersin.org/articles/10.3389/fninf.2022.1032538/full}, doi = {10.3389/fninf.2022.1032538}, year = {2023}, date = {2023-01-11}, journal = {Frontiers in Neuroinformatics}, abstract = {Modern Graph Neural Networks (GNNs) provide opportunities to study the determinants underlying the complex activity patterns of biological neuronal networks. In this study, we applied GNNs to a large-scale electrophysiological dataset of rodent primary neuronal networks obtained by means of high-density microelectrode arrays (HD-MEAs). HD-MEAs allow for long-term recording of extracellular spiking activity of individual neurons and networks and enable the extraction of physiologically relevant features at the single- neuron and population level. We employed established GNNs to generate a combined representation of single-neuron and connectivity features obtained from HD-MEA data, with the ultimate goal of predicting changes in single-neuron firing rate induced by a pharmacological perturbation. The aim of the main prediction task was to assess whether single-neuron and functional connectivity features, inferred under baseline conditions, were informative for predicting changes in neuronal activity in response to a perturbation with Bicuculline, a GABAA receptor antagonist. Our results suggest that the joint representation of node features and functional connectivity, extracted from a baseline recording, was informative for predicting firing rate changes of individual neurons after the perturbation. Specifically, our implementation of a GNN model with inductive learning capability (GraphSAGE) outperformed other prediction models that relied only on single-neuron features. We tested the generalizability of the results on two additional datasets of HD-MEA recordings–a second dataset with cultures perturbed with Bicuculline and a dataset perturbed with the GABAA receptor antagonist Gabazine. GraphSAGE models showed improved prediction accuracy over other prediction models. Our results demonstrate the added value of taking into account the functional connectivity between neurons and the potential of GNNs to study complex interactions between neurons.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Modern Graph Neural Networks (GNNs) provide opportunities to study the determinants underlying the complex activity patterns of biological neuronal networks. In this study, we applied GNNs to a large-scale electrophysiological dataset of rodent primary neuronal networks obtained by means of high-density microelectrode arrays (HD-MEAs). HD-MEAs allow for long-term recording of extracellular spiking activity of individual neurons and networks and enable the extraction of physiologically relevant features at the single- neuron and population level. We employed established GNNs to generate a combined representation of single-neuron and connectivity features obtained from HD-MEA data, with the ultimate goal of predicting changes in single-neuron firing rate induced by a pharmacological perturbation. The aim of the main prediction task was to assess whether single-neuron and functional connectivity features, inferred under baseline conditions, were informative for predicting changes in neuronal activity in response to a perturbation with Bicuculline, a GABAA receptor antagonist. Our results suggest that the joint representation of node features and functional connectivity, extracted from a baseline recording, was informative for predicting firing rate changes of individual neurons after the perturbation. Specifically, our implementation of a GNN model with inductive learning capability (GraphSAGE) outperformed other prediction models that relied only on single-neuron features. We tested the generalizability of the results on two additional datasets of HD-MEA recordings–a second dataset with cultures perturbed with Bicuculline and a dataset perturbed with the GABAA receptor antagonist Gabazine. GraphSAGE models showed improved prediction accuracy over other prediction models. Our results demonstrate the added value of taking into account the functional connectivity between neurons and the potential of GNNs to study complex interactions between neurons. Close https://www.frontiersin.org/articles/10.3389/fninf.2022.1032538/full doi:10.3389/fninf.2022.1032538 Close
15.	Sato, Yuya; Yamamoto, Hideaki; Kato, Hideyuki; Tanii, Takashi; Sato, Shigeo; Hirano-Iwata, Ayumi: Microfluidic cell engineering on high-density microelectrode arrays for assessing structure-function relationships in living neuronal networks. In: Frontiers in Neuroscience, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Sato2023, title = {Microfluidic cell engineering on high-density microelectrode arrays for assessing structure-function relationships in living neuronal networks}, author = {Yuya Sato and Hideaki Yamamoto and Hideyuki Kato and Takashi Tanii and Shigeo Sato and Ayumi Hirano-Iwata}, url = {https://www.frontiersin.org/articles/10.3389/fnins.2022.943310/full}, doi = {doi: 10.3389/fnins.2022.943310}, year = {2023}, date = {2023-01-09}, journal = {Frontiers in Neuroscience}, abstract = {Neuronal networks in dissociated culture combined with cell engineering technology offer a pivotal platform to constructively explore the relationship between structure and function in living neuronal networks. Here, we fabricated defined neuronal networks possessing a modular architecture on high-density microelectrode arrays (HD-MEAs), a state-of-the-art electrophysiological tool for recording neural activity with high spatial and temporal resolutions. We first established a surface coating protocol using a cell-permissive hydrogel to stably attach a polydimethylsiloxane microfluidic film on the HD-MEA. We then recorded the spontaneous neural activity of the engineered neuronal network, which revealed an important portrait of the engineered neuronal network–modular architecture enhances functional complexity by reducing the excessive neural correlation between spatially segregated modules. The results of this study highlight the impact of HD- MEA recordings combined with cell engineering technologies as a novel tool in neuroscience to constructively assess the structure-function relationships in neuronal networks.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Neuronal networks in dissociated culture combined with cell engineering technology offer a pivotal platform to constructively explore the relationship between structure and function in living neuronal networks. Here, we fabricated defined neuronal networks possessing a modular architecture on high-density microelectrode arrays (HD-MEAs), a state-of-the-art electrophysiological tool for recording neural activity with high spatial and temporal resolutions. We first established a surface coating protocol using a cell-permissive hydrogel to stably attach a polydimethylsiloxane microfluidic film on the HD-MEA. We then recorded the spontaneous neural activity of the engineered neuronal network, which revealed an important portrait of the engineered neuronal network–modular architecture enhances functional complexity by reducing the excessive neural correlation between spatially segregated modules. The results of this study highlight the impact of HD- MEA recordings combined with cell engineering technologies as a novel tool in neuroscience to constructively assess the structure-function relationships in neuronal networks. Close https://www.frontiersin.org/articles/10.3389/fnins.2022.943310/full doi:doi: 10.3389/fnins.2022.943310 Close
16.	Bartram, Julian; Franke, Felix; Kumar, Sreedhar Saseendran; Buccino, Alessio Paolo; Xue, Xiaohan; Gänswein, Tobias; Schröter, Manuel; Kim, Taehoon; Kasuba, Krishna Chaitanya; Hierlemann, Andreas: Parallel reconstruction of the excitatory and inhibitory inputs received by single neurons reveals the synaptic basis of recurrent spiking. In: bioRxiv, 2023. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Bartram2023, title = {Parallel reconstruction of the excitatory and inhibitory inputs received by single neurons reveals the synaptic basis of recurrent spiking}, author = {Julian Bartram and Felix Franke and Sreedhar Saseendran Kumar and Alessio Paolo Buccino and Xiaohan Xue and Tobias Gänswein and Manuel Schröter and Taehoon Kim and Krishna Chaitanya Kasuba and Andreas Hierlemann}, url = {https://www.biorxiv.org/content/10.1101/2023.01.06.523018v2}, doi = {https://doi.org/10.1101/2023.01.06.523018}, year = {2023}, date = {2023-01-08}, journal = {bioRxiv}, abstract = {Self-sustained recurrent activity in cortical networks is thought to be important for multiple crucial processes, including circuit development and homeostasis. However, the precise relationship between synaptic input patterns and spiking output of individual neurons remains unresolved during spontaneous network activity. Here, using whole-network high-density microelectrode array (HD-MEA) recordings and patch clamping, we developed a novel experimental approach and analytical tools that provide a comprehensive long-term input-output characterization of individual neurons in cortical cell cultures. We found that, during in vivo-like network activity with excitation(E)-inhibition(I) balance, postsynaptic spiking coincided with the maxima of rapid, network state-dependent fluctuations in the input E/I ratio. Our approach also uncovered the underlying circuit architecture and we identified a few key inhibitory inputs – often from special hub neurons – that were instrumental in mediating these E/I ratio changes. Balanced network theory predicts dynamical regimes governed by input fluctuation and featuring a fast neuronal responsiveness. Our findings – obtained in self-organized neuronal cultures – suggest that the emergence of these favorable regimes and associated network architectures is an inherent property of all cortical networks.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Self-sustained recurrent activity in cortical networks is thought to be important for multiple crucial processes, including circuit development and homeostasis. However, the precise relationship between synaptic input patterns and spiking output of individual neurons remains unresolved during spontaneous network activity. Here, using whole-network high-density microelectrode array (HD-MEA) recordings and patch clamping, we developed a novel experimental approach and analytical tools that provide a comprehensive long-term input-output characterization of individual neurons in cortical cell cultures. We found that, during in vivo-like network activity with excitation(E)-inhibition(I) balance, postsynaptic spiking coincided with the maxima of rapid, network state-dependent fluctuations in the input E/I ratio. Our approach also uncovered the underlying circuit architecture and we identified a few key inhibitory inputs – often from special hub neurons – that were instrumental in mediating these E/I ratio changes. Balanced network theory predicts dynamical regimes governed by input fluctuation and featuring a fast neuronal responsiveness. Our findings – obtained in self-organized neuronal cultures – suggest that the emergence of these favorable regimes and associated network architectures is an inherent property of all cortical networks. Close https://www.biorxiv.org/content/10.1101/2023.01.06.523018v2 doi:https://doi.org/10.1101/2023.01.06.523018 Close
17.	Han, Xiaobo; Matsuda, Naoki; Ishibashi, Yuto; Odawara, Aoi; Takahashi, Sayuri; Tooi, Norie; Kinoshita, Koshi; Suzuki, Ikuro: A functional neuron maturation device provides convenient application on microelectrode array for neural network measurement. In: Biomaterials Research, 2022. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Han2022, title = {A functional neuron maturation device provides convenient application on microelectrode array for neural network measurement}, author = {Xiaobo Han and Naoki Matsuda and Yuto Ishibashi and Aoi Odawara and Sayuri Takahashi and Norie Tooi and Koshi Kinoshita and Ikuro Suzuki }, url = {https://biomaterialsres.biomedcentral.com/articles/10.1186/s40824-022-00324-z}, doi = {https://doi.org/10.1186/s40824-022-00324-z}, year = {2022}, date = {2022-12-20}, journal = {Biomaterials Research}, abstract = {Background Microelectrode array (MEA) systems are valuable for in vitro assessment of neurotoxicity and drug efficiency. However, several difficulties such as protracted functional maturation and high experimental costs hinder the use of MEA analysis requiring human induced pluripotent stem cells (hiPSCs). Neural network functional parameters are also needed for in vitro to in vivo extrapolation. Methods In the present study, we produced a cost effective nanofiber culture platform, the SCAD device, for long-term culture of hiPSC-derived neurons and primary peripheral neurons. The notable advantage of SCAD device is convenient application on multiple MEA systems for neuron functional analysis. Results We showed that the SCAD device could promote functional maturation of cultured hiPSC-derived neurons, and neurons responded appropriately to convulsant agents. Furthermore, we successfully analyzed parameters for in vitro to in vivo extrapolation, i.e., low-frequency components and synaptic propagation velocity of the signal, potentially reflecting neural network functions from neurons cultured on SCAD device. Finally, we measured the axonal conduction velocity of peripheral neurons. Conclusions: Neurons cultured on SCAD devices might constitute a reliable in vitro platform to investigate neuron functions, drug efficacy and toxicity, and neuropathological mechanisms by MEA.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Background Microelectrode array (MEA) systems are valuable for in vitro assessment of neurotoxicity and drug efficiency. However, several difficulties such as protracted functional maturation and high experimental costs hinder the use of MEA analysis requiring human induced pluripotent stem cells (hiPSCs). Neural network functional parameters are also needed for in vitro to in vivo extrapolation. Methods In the present study, we produced a cost effective nanofiber culture platform, the SCAD device, for long-term culture of hiPSC-derived neurons and primary peripheral neurons. The notable advantage of SCAD device is convenient application on multiple MEA systems for neuron functional analysis. Results We showed that the SCAD device could promote functional maturation of cultured hiPSC-derived neurons, and neurons responded appropriately to convulsant agents. Furthermore, we successfully analyzed parameters for in vitro to in vivo extrapolation, i.e., low-frequency components and synaptic propagation velocity of the signal, potentially reflecting neural network functions from neurons cultured on SCAD device. Finally, we measured the axonal conduction velocity of peripheral neurons. Conclusions: Neurons cultured on SCAD devices might constitute a reliable in vitro platform to investigate neuron functions, drug efficacy and toxicity, and neuropathological mechanisms by MEA. Close https://biomaterialsres.biomedcentral.com/articles/10.1186/s40824-022-00324-z doi:https://doi.org/10.1186/s40824-022-00324-z Close
18.	Lent, Jonas Van; Vendredy, Leen; Adriaenssens, Elias; Authier, Tatiana Da Silva; Asselbergh, Bob; Kaji, Marcus; Weckhuysen, Sarah; Bosch, Ludo Van Den; Baets, Jonathan; Timmerman, Vincent: Downregulation of PMP22 ameliorates myelin defects in iPSC-derived human organoid cultures of CMT1A. In: Brain, 2022. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{VanLent2022, title = {Downregulation of PMP22 ameliorates myelin defects in iPSC-derived human organoid cultures of CMT1A}, author = {Jonas Van Lent and Leen Vendredy and Elias Adriaenssens and Tatiana Da Silva Authier and Bob Asselbergh and Marcus Kaji and Sarah Weckhuysen and Ludo Van Den Bosch and Jonathan Baets and Vincent Timmerman}, url = {https://academic.oup.com/brain/advance-article/doi/10.1093/brain/awac475/6895197?login=false}, doi = {https://doi.org/10.1093/brain/awac475}, year = {2022}, date = {2022-12-12}, journal = {Brain}, abstract = {Charcot-Marie-Tooth (CMT) disease is the most common inherited disorder of the peripheral nervous system. CMT1A accounts for 40-50% of all cases and is caused by a duplication of the PMP22 gene on chromosome 17, leading to dysmyelination in the peripheral nervous system. Patient-derived models to study such myelination defects are lacking as the in vitro generation of human myelinating Schwann cells has proven to be particularly challenging. Here, we present an iPSC-derived organoid culture, containing various cell types of the peripheral nervous system, including myelinating human Schwann cells, which mimics the human peripheral nervous system. Single-cell analysis confirmed the peripheral nervous system-like cellular composition and provides insight into the developmental trajectory. We used this organoid-model to study disease signatures of CMT1A, revealing early ultrastructural myelin alterations, including increased myelin periodic line distance and hypermyelination of small axons. Furthermore, we observed the presence of onion bulb-like formations in a later developmental stage. These hallmarks were not present in the for CMT1A-corrected isogenic line or in a CMT2A iPSC line, supporting the notion that these alterations are specific to CMT1A. Downregulation of PMP22 expression using short-hairpin RNAs or a combinatorial drug consisting of baclofen, naltrexone hydrochloride and D-sorbitol, was able to ameliorate the myelin defects in CMT1A-organoids. In summary, this self-organizing organoid model is able to capture biologically meaningful features of the disease and capture the physiological complexity, forms an excellent model to study demyelinating diseases, and supports the therapeutic approach of reducing PMP22 expression.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Charcot-Marie-Tooth (CMT) disease is the most common inherited disorder of the peripheral nervous system. CMT1A accounts for 40-50% of all cases and is caused by a duplication of the PMP22 gene on chromosome 17, leading to dysmyelination in the peripheral nervous system. Patient-derived models to study such myelination defects are lacking as the in vitro generation of human myelinating Schwann cells has proven to be particularly challenging. Here, we present an iPSC-derived organoid culture, containing various cell types of the peripheral nervous system, including myelinating human Schwann cells, which mimics the human peripheral nervous system. Single-cell analysis confirmed the peripheral nervous system-like cellular composition and provides insight into the developmental trajectory. We used this organoid-model to study disease signatures of CMT1A, revealing early ultrastructural myelin alterations, including increased myelin periodic line distance and hypermyelination of small axons. Furthermore, we observed the presence of onion bulb-like formations in a later developmental stage. These hallmarks were not present in the for CMT1A-corrected isogenic line or in a CMT2A iPSC line, supporting the notion that these alterations are specific to CMT1A. Downregulation of PMP22 expression using short-hairpin RNAs or a combinatorial drug consisting of baclofen, naltrexone hydrochloride and D-sorbitol, was able to ameliorate the myelin defects in CMT1A-organoids. In summary, this self-organizing organoid model is able to capture biologically meaningful features of the disease and capture the physiological complexity, forms an excellent model to study demyelinating diseases, and supports the therapeutic approach of reducing PMP22 expression. Close https://academic.oup.com/brain/advance-article/doi/10.1093/brain/awac475/6895197[...] doi:https://doi.org/10.1093/brain/awac475 Close
19.	Tran, Hoang-Dai; Shin, Min-Kyoung; Denman, Charlotte; Han, Run-Run; Kuhn, Bernd; Arbuthnott, Gordon; Jo, Junghyun: Generation of Human Striatal-Midbrain Assembloids From Human Pluripotent Stem Cells to Model Alpha-Synuclein Propagation. In: Sneak Peek - Cell Press, 2022. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Tran2022, title = {Generation of Human Striatal-Midbrain Assembloids From Human Pluripotent Stem Cells to Model Alpha-Synuclein Propagation}, author = {Hoang-Dai Tran and Min-Kyoung Shin and Charlotte Denman and Run-Run Han and Bernd Kuhn and Gordon Arbuthnott and Junghyun Jo}, url = {https://papers.ssrn.com/sol3/papers.cfm?abstract_id=4288935}, doi = {http://dx.doi.org/10.2139/ssrn.4288935}, year = {2022}, date = {2022-12-05}, journal = {Sneak Peek - Cell Press}, abstract = {Animal models of the pathology of Parkinson’s disease (PD) have provided most of the treatments to date, but the disease is restricted to human patients. In vitro models using human pluripotent stem cell-derived neural organoids have provided improved access to study PD etiology. Here, we generated human striatal and midbrain organoids and assembled both regionalized neural organoids to form human striatal-midbrain assembloids (hSMAs), mimicking a part of basal ganglia. Both the nigrostriatal and striatonigral pathways are present and electrophysiologically active in the hSMAs. hSMA development in the presence of increased alpha-synuclein (α-syn) from SNCA overexpression, induced nigrostriatal system damage, which is typical of the disease. Using the α-syn-mKO2 reporter and bimolecular fluorescence complementation system, we demonstrated that fluorescent α-syn is transported from the striatal area tothe dopaminergic (DA) neurons of the midbrain area. Furthermore, insoluble α-syn aggregated over time in DA neurons similar to Lewy bodies in human patients. Such assembloids are a compelling new platform to develop novel PD therapeutic strategies.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Animal models of the pathology of Parkinson’s disease (PD) have provided most of the treatments to date, but the disease is restricted to human patients. In vitro models using human pluripotent stem cell-derived neural organoids have provided improved access to study PD etiology. Here, we generated human striatal and midbrain organoids and assembled both regionalized neural organoids to form human striatal-midbrain assembloids (hSMAs), mimicking a part of basal ganglia. Both the nigrostriatal and striatonigral pathways are present and electrophysiologically active in the hSMAs. hSMA development in the presence of increased alpha-synuclein (α-syn) from SNCA overexpression, induced nigrostriatal system damage, which is typical of the disease. Using the α-syn-mKO2 reporter and bimolecular fluorescence complementation system, we demonstrated that fluorescent α-syn is transported from the striatal area tothe dopaminergic (DA) neurons of the midbrain area. Furthermore, insoluble α-syn aggregated over time in DA neurons similar to Lewy bodies in human patients. Such assembloids are a compelling new platform to develop novel PD therapeutic strategies. Close https://papers.ssrn.com/sol3/papers.cfm?abstract_id=4288935 doi:http://dx.doi.org/10.2139/ssrn.4288935 Close
20.	Akarca, Danyal; Dunn, Alexander W E; Hornauer, Philipp J; Ronchi, Silvia; Fiscella, Michele; Wang, Congwei; Terrigno, Marco; Jagasia, Ravi; Vértes, Petra E; Mierau, Susanna B; Paulsen, Ole; Eglen, Stephen J; Hierlemann, Andreas; Astle, Duncan E; Schröter, Manuel: Homophilic wiring principles underpin neuronal network topology in vitro. In: BioRxiv, 2022. (Type: Journal Article \| Abstract \| Links \| BibTeX) @article{Akarca2022, title = {Homophilic wiring principles underpin neuronal network topology in vitro}, author = {Danyal Akarca and Alexander W. E. Dunn and Philipp J. Hornauer and Silvia Ronchi and Michele Fiscella and Congwei Wang and Marco Terrigno and Ravi Jagasia and Petra E. Vértes and Susanna B. Mierau and Ole Paulsen and Stephen J. Eglen and Andreas Hierlemann and Duncan E. Astle and Manuel Schröter}, url = {https://www.biorxiv.org/content/10.1101/2022.03.09.483605v2.abstract}, doi = {https://doi.org/10.1101/2022.03.09.483605}, year = {2022}, date = {2022-12-01}, journal = {BioRxiv}, abstract = {Economic efficiency has been a popular explanation for how networks self-organize within the developing nervous system. However, the precise nature of the economic negotiations governing this putative organizational principle remains unclear. Here, we address this question further by combining large-scale electrophysiological recordings, to characterize the functional connectivity of developing neuronal networks in vitro, with a generative modeling approach capable of simulating network formation. We find that the best fitting model uses a homophilic generative wiring principle in which neurons form connections to other neurons which are spatially proximal and have similar connectivity patterns to themselves. Homophilic generative models outperform more canonical models in which neurons wire depending upon their spatial proximity either alone or in combination with the extent of their local connectivity. This homophily-based mechanism for neuronal network emergence accounts for a wide range of observations that are described, but not sufficiently explained, by traditional analyses of network topology. Using rodent and human monolayer and organoid cultures, we show that homophilic generative mechanisms can accurately recapitulate the topology of emerging cellular functional connectivity, representing an important wiring principle and determining factor of neuronal network formation in vitro.}, keywords = {}, pubstate = {published}, tppubtype = {article} } Close Economic efficiency has been a popular explanation for how networks self-organize within the developing nervous system. However, the precise nature of the economic negotiations governing this putative organizational principle remains unclear. Here, we address this question further by combining large-scale electrophysiological recordings, to characterize the functional connectivity of developing neuronal networks in vitro, with a generative modeling approach capable of simulating network formation. We find that the best fitting model uses a homophilic generative wiring principle in which neurons form connections to other neurons which are spatially proximal and have similar connectivity patterns to themselves. Homophilic generative models outperform more canonical models in which neurons wire depending upon their spatial proximity either alone or in combination with the extent of their local connectivity. This homophily-based mechanism for neuronal network emergence accounts for a wide range of observations that are described, but not sufficiently explained, by traditional analyses of network topology. Using rodent and human monolayer and organoid cultures, we show that homophilic generative mechanisms can accurately recapitulate the topology of emerging cellular functional connectivity, representing an important wiring principle and determining factor of neuronal network formation in vitro. Close https://www.biorxiv.org/content/10.1101/2022.03.09.483605v2.abstract doi:https://doi.org/10.1101/2022.03.09.483605 Close