Rapid generation of functional engineered 3D human neuronal assemblies: network dynamics evaluated by Micro-Electrodes Arrays

2021 ◽  
Author(s):  
Lorenzo Muzzi ◽  
Donatella Di Lisa ◽  
Pietro Arnaldi ◽  
Davide Aprile ◽  
Laura Pastorino ◽  
...  
Keyword(s):  
Neuronal Networks ◽  
Three Dimensional ◽  
Electrode Array ◽  
Good Control ◽  
Neuronal Population ◽  
Point Of View ◽  
Network Dynamic ◽  
Electrode Arrays ◽  
Neuronal Assemblies

Abstract Objective: In this work we propose a method for producing engineered human derived three-dimensional neuronal assemblies coupled to Micro-Electrode Array (MEA) substrates for studying the electrophysiological activity of such networks. Approach: We used biocompatible chitosan microbeads as scaffold to build 3D networks and to ensure nutrients-medium exchange from the core of the structure to the external environment. We used excitatory neurons derived from human-induced Pluripotent Stem Cells (hiPSCs) co-cultured with astrocytes. By adapting the well-established NgN2 differentiation protocol, we obtained 3D engineered networks with good control over cell density, volume and cell composition. We coupled the 3D neuronal networks to 60-channel Micro Electrode Arrays (MEAs) to evaluate and monitor the functional activity of the neuronal population. In parallel, we generated two-dimensional neuronal networks to compare the results of the two models. Main results: 3D cultures were healthy and functional up to 42 Days In Vitro (DIVs). From the structural point of view, the hiPSC derived neurons were able to adhere to chitosan microbeads and to form a stable 3D assembly thanks to the connections among cells. From a functional point of view, neuronal networks showed spontaneous activity after a couple of weeks. We monitored the functional electrophysiological behavior up to 6 weeks and we compared the network dynamic with 2D models. Significance: We presented for the first time a method to generate 3D engineered cultures with human-derived neurons coupled to MEAs, overcoming some of the limitations related to 2D and 3D neuronal networks and thus increasing the therapeutic target potential of these models for biomedical applications.

scholarly journals L-type voltage-gated calcium channel regulation of in vitro human cortical neuronal networks

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
William Plumbly ◽  
Nick Brandon ◽  
Tarek Z. Deeb ◽  
Jeremy Hall ◽  
Adrian J. Harwood
Keyword(s):  
Calcium Channel ◽  
Neuronal Networks ◽  
Gene Mutations ◽  
Synaptic Function ◽  
Neuronal Firing ◽  
Network Activity ◽  
Electrode Arrays ◽  
Voltage Gated ◽  
Voltage Gated Calcium Channel

Abstract The combination of in vitro multi-electrode arrays (MEAs) and the neuronal differentiation of stem cells offers the capability to study human neuronal networks from patient or engineered human cell lines. Here, we use MEA-based assays to probe synaptic function and network interactions of hiPSC-derived neurons. Neuronal network behaviour first emerges at approximately 30 days of culture and is driven by glutamate neurotransmission. Over a further 30 days, inhibitory GABAergic signalling shapes network behaviour into a synchronous regular pattern of burst firing activity and low activity periods. Gene mutations in L-type voltage gated calcium channel subunit genes are strongly implicated as genetic risk factors for the development of schizophrenia and bipolar disorder. We find that, although basal neuronal firing rate is unaffected, there is a dose-dependent effect of L-type voltage gated calcium channel inhibitors on synchronous firing patterns of our hiPSC-derived neural networks. This demonstrates that MEA assays have sufficient sensitivity to detect changes in patterns of neuronal interaction that may arise from hypo-function of psychiatric risk genes. Our study highlights the utility of in vitro MEA based platforms for the study of hiPSC neural network activity and their potential use in novel compound screening.

scholarly journals L-type voltage-gated calcium channel regulation of in vitro human cortical neuronal networks

2018 ◽  
Author(s):  
William Plumbly ◽  
Nicholas J. Brandon ◽  
Tarek Z. Deeb ◽  
Jeremy Hall ◽  
Adrian J. Harwood
Keyword(s):  
Calcium Channel ◽  
Neuronal Networks ◽  
Gene Mutations ◽  
Synaptic Function ◽  
Neuronal Firing ◽  
Network Activity ◽  
Electrode Arrays ◽  
Voltage Gated ◽  
Voltage Gated Calcium Channel

The combination of in vitro multi-electrode arrays (MEAs) and the neuronal differentiation of stem cells offers the capability to study human neuronal networks from patient or engineered human cell lines. Here, we use MEA-based assays to probe synaptic function and network interactions of hiPSC-derived neurons. Neuronal network behaviour first emerges at approximately 30 days of culture and is driven by glutamate neurotransmission. Over a further 30 days, inhibitory GABergic signalling shapes network behaviour into a synchronous regular pattern of burst firing activity and low activity periods. Gene mutations in L-type voltage gated calcium channel subunit genes are strongly implicated as genetic risk factors for the development of schizophrenia and bipolar disorder. We find that, although basal neuronal firing rate is unaffected, there is a dose-dependent effect of L-type voltage gated calcium channel inhibitors on synchronous firing patterns of our hiPSC-derived neural networks. This demonstrates that MEA assays have sufficient sensitivity to detect changes in patterns of neuronal interaction that may arise from hypo-function of psychiatric risk genes. Our study highlights the utility of in vitro MEA based platforms for the study of hiPSC neural network activity and their potential use in novel compound screening.

Analysis of fitness of prosthetic bridge designed and produced with digital methods

2018 ◽  
Vol 68 (1) ◽  
pp. 32-44
Author(s):  
Daniel Surowiecki ◽  
Jan Wierzba ◽  
Roman Grygoruk ◽  
Kamila Wróbel-Bednarz
Keyword(s):  
Three Dimensional ◽  
Additive Technologies ◽  
Point Of View ◽  
Digital Design ◽  
Three Dimensional Objects ◽  
Average Value ◽  
Conventional Procedure ◽  
Abutment Teeth ◽  
Digital Methods

Introduction. In recent years the digitalization of dentistry has taken a special place in the world of scientific research. With the development of scanning methods and the improvement of software for the analysis and designing of three-dimensional objects, as well as the adaptation of milling techniques and additive technologies for dentistry applications in many clinical cases it is possible to replace the conventional procedure of prosthetic restoration with the digital process. It should be emphasized that the need of further thorough research is recommended especially on the accuracy of elements obtained with intraoral scanning and their production with additive technologies. Aim of the study. Analysis of marginal and internal fit of a prosthetic bridge designed and produced using digital methods. Material and methods. On the model of the jaw abutment teeth have been prepared to make a bridge. Using the intraoral scanner dental arches were scanned. The digital design was performed which was then printed using DMLS technology. The prosthetic restoration was 3D scanned in order to obtain a digital geometry which was processed with specialized software. The bridge fitting analysis to the CAD model of the abutment teeth was performed. Results. Heat map analysis allows to determine the high accuracy of bridge fitting within the abutment teeth. From 50 measurement points an average value of 0.0136 with a standard deviation of 0.0768 was obtained. The minimum value was -0.21, maximum 0.17. Discussion. As it results from the study and reports in the literature in many clinical situations it is possible to perform a completely digital procedure of producing prosthetic restorations. In vitro studies show the potential clinical usefulness of digital methods, however, most authors agree on the need of thorough clinical trials before fully recommending the application of completely digital procedures. The use of the developed methodology allows to perform further research on the accuracy of these technologies. Conclusions. The obtained results indicate the accuracy of producing prosthetic restoration with digital techniques. Application of these technologies allows to produce bridge characterized by high precision. The reverse engineering method can be used to numerically determine parameters which are relevant from the point of view of medical applications.

scholarly journals Three dimensional microenvironments on multi-electrode arrays produce neuronal networks that function like the brain

2018 ◽  
Vol 12 ◽  
Author(s):  
Justin Bourke ◽  
Anita Quigley ◽  
Cathal O'Connell ◽  
Jeremy Crook ◽  
Gordon Wallace ◽  
...  
Keyword(s):  
Neuronal Networks ◽  
Three Dimensional ◽  
Electrode Arrays ◽  
The Brain

scholarly journals Growing neuronal islands on multi-electrode arrays using an Accurate Positioning-μCP device

2015 ◽  
Author(s):  
Robert Samhaber ◽  
Manuel Schottdorf ◽  
Ahmed El Hady ◽  
Kai Broeking ◽  
Andreas Daus ◽  
...  
Keyword(s):  
Cell Culture ◽  
Neuronal Networks ◽  
Standard Cell ◽  
Electrode Arrays ◽  
Unit Recording ◽  
Contact Printing ◽  
Novel Approach ◽  
Custom Made ◽  
Typical Cell

Multi-electrode arrays (MEAs) allow non-invasive multi-unit recording in-vitro from cultured neuronal networks. For sufficient neuronal growth and adhesion on such MEAs, substrate preparation is required. Plating of dissociated neurons on a uniformly prepared MEA's surface results in the formation of spatially extended random networks with substantial inter-sample variability. Such cultures are not optimally suited to study the relationship between defined structure and dynamics in neuronal networks. To overcome these shortcomings, neurons can be cultured with pre-defined topology by spatially structured surface modification. Spatially structuring a MEA surface accurately and reproducibly with the equipment of a typical cell-culture laboratory is challenging. In this paper, we present a novel approach utilizing micro-contact printing (μCP) combined with a custom-made device to accurately position patterns on MEAs with high precision. We call this technique AP-μCP (accurate positioning micro-contact printing). Other approaches presented in the literature using μCP for patterning either relied on facilities or techniques not readily available in a standard cell culture laboratory, or they did not specify means of precise pattern positioning. Here we present a relatively simple device for reproducible and precise patterning in a standard cell-culture laboratory setting. The patterned neuronal islands on MEAs provide a basis for high throughput electrophysiology to study the dynamics of single neurons and neuronal networks.

scholarly journals 3F(eature)s model: modularity, heterogeneity and three-dimensionality to design in vitro neuronal model

2021 ◽  
Vol 4 (s1) ◽  
Author(s):  
Martina Brofiga ◽  
Marietta Pisano ◽  
Mariateresa Tedesco ◽  
Francesca Callegari ◽  
Paolo Massobrio
Keyword(s):  
Three Dimensional ◽  
Neuronal Model ◽  
Neuronal Cultures ◽  
Electrode Arrays ◽  
Experimental Platform ◽  
Electrophysiological Activity ◽  
Micro Electrode Arrays ◽  
S Model

In this work, we present a novel experimental platform to build in vitro interconnected (i.e., modular) heterogeneous (e.g., cortical-hippocampal) and three-dimensional (3D) neuronal cultures plated on Micro-Electrode Arrays (MEAs) to extracellularly record the electrophysiological activity continuously.

Explorative Data Analysis of In-Vitro Neuronal Network Behavior Based on an Unsupervised Learning Approach

2010 ◽  
pp. 241-253
Author(s):  
A. Maffezzoli ◽  
E. Wanke
Keyword(s):  
Neuronal Networks ◽  
Analytical Techniques ◽  
Learning Approach ◽  
Electrode Arrays ◽  
Activity Data ◽  
New Information ◽  
Data Dimensionality Reduction ◽  
Long Time ◽  
Pros And Cons

In the present chapter authors want to expose new insights in the field of Computational Neuroscience at regard to the study of neuronal networks grown in vitro. Such kind of analyses can exploit the availability of a huge amount of data thanks to the use of Multi Electrode Arrays (MEA), a multi-channel technology which allows capturing the activity of several different neuronal cells for long time recordings. Given the possibility of simultaneous targeting of various sites, neuroscientists are so applying such recent technology for various researches. The chapter begins by giving a brief presentation of MEA technology and of the data produced in output, punctuating some of the pros and cons of MEA recordings. Then we present an overview of the analytical techniques applied in order to extrapolate the hidden information from available data. Then we shall explain the approach we developed and applied on MEAs prepared in our cell culture laboratory, consisting of statistical methods capturing the main features of the spiking, in particular bursting, activity of various neuron, and performing data dimensionality reduction and clustering, in order to classify neurons according to their spiking properties having showed correlated features. Finally the chapter wants to furnish to neuroscientists an overview about the quantitative analysis of in-vitro spiking activity data recorded via MEA technology and to give an example of explorative analysis applied on MEA data. Such study is based on methods from Statistics and Machine Learning or Computer Science but at the same time strictly related to neurophysiological interpretations of the putative pharmacological manipulation of synaptic connections and mode of firing, with the final aim to extract new information and knowledge about neuronal networks behavior and organization.

scholarly journals Evaluation of the Efficacy of Dexamethasone-Eluting Electrode Array on the Post-Implant Cochlear Fibrotic Reaction by Three-Dimensional Immunofluorescence Analysis in Mongolian Gerbil Cochlea

2021 ◽  
Vol 10 (15) ◽  
pp. 3315
Author(s):  
Philippine Toulemonde ◽  
Michaël Risoud ◽  
Pierre Emmanuel Lemesre ◽  
Cyril Beck ◽  
Jean Wattelet ◽  
...  
Keyword(s):  
Cochlear Implant ◽  
Dimensional Analysis ◽  
Mongolian Gerbil ◽  
Three Dimensional ◽  
Electrode Array ◽  
Light Sheet ◽  
Tissue Response ◽  
Electrode Arrays ◽  
Immunofluorescence Analysis ◽  
Light Sheet Microscopy

Cochlear implant is the method of choice for the rehabilitation of severe to profound sensorineural hearing loss. The study of the tissue response to cochlear implantation and the prevention of post-cochlear-implant damages are areas of interest in hearing protection research. The objective was to assess the efficacy of dexamethasone-eluting electrode array on endo canal fibrosis formation by three-dimensional immunofluorescence analysis in implanted Mongolian gerbil cochlea. Two trials were conducted after surgery using Mongolian gerbil implanted with dexamethasone-eluting or non-eluting intracochlear electrode arrays. The animals were then euthanised 10 weeks after implantation. The cochleae were prepared (electrode array in place) according to a 29-day protocol with immunofluorescent labelling and tissue clearing. The acquisition was carried out using light-sheet microscopy. Imaris software was then used for three-dimensional analysis of the cochleae and quantification of the fibrotic volume. The analysis of 12 cochleae showed a significantly different mean volume of fibrosis (2.16 × 108 μm3 ± 0.15 in the dexamethasone eluting group versus 3.17 × 108 μm3 ± 0.54 in the non-eluting group) (p = 0.004). The cochlear implant used as a corticosteroid delivery system appears to be an encouraging device for the protection of the inner ear against fibrosis induced by implantation. Three-dimensional analysis of the cochlea by light-sheet microscopy was suitable for studying post-implantation tissue damage.

scholarly journals Human neuronal networks on micro-electrode arrays are a highly robust tool to study disease-specific genotype-phenotype correlations in vitro

2021 ◽  
Author(s):  
B. Mossink ◽  
A.H.A. Verboven ◽  
E.J.H. van Hugte ◽  
T.M. Klein Gunnewiek ◽  
G. Parodi ◽  
...  
Keyword(s):  
Experimental Design ◽  
Neuronal Network ◽  
Neuronal Networks ◽  
Network Activity ◽  
Full Potential ◽  
Electrode Arrays ◽  
Neuronal Network Activity ◽  
Micro Electrode Arrays ◽  
Disease Specific

AbstractMicro-electrode arrays (MEAs) are increasingly used to characterize neuronal network activity of human induced pluripotent stem-cell (hiPSC)-derived neurons. Despite their gain in popularity, MEA recordings from hiPSC-derived neuronal networks are not always used to their full potential in respect to experimental design, execution and data analysis. Therefore, we benchmarked the robustness and sensitivity of MEA-derived neuronal activity patterns derived from ten healthy individual control lines. We provide recommendations on experimental design and analysis to achieve standardization. With such standardization, MEAs can be used as a reliable platform to distinguish (disease-specific) network phenotypes. In conclusion, we show that MEAs are a powerful and robust tool to uncover functional neuronal network phenotypes from hiPSC-derived neuronal networks, and provide an important resource to advance the hiPSC field towards the use of MEAs for disease-phenotyping and drug discovery.

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