Neural network formation is a complex process involving axon outgrowth and guidance. Axon guidance is facilitated by structural and molecular cues from the surrounding microenvironment. Micro-fabrication techniques can be employed to produce microfluidic chips with a highly controlled microenvironment for neural cells enabling longitudinal studies of complex processes associated with network formation. In this work, we demonstrate a novel open microfluidic chip design that encompasses a freely variable number of nodes interconnected by axon-permissible tunnels, enabling structuring of multi-nodal neural networks in vitro. The chip employs a partially open design to allow high level of control and reproducibility of cell seeding, while reducing shear stress on the cells. We show that by culturing dorsal root ganglion cells (DRGs) in our microfluidic chip, we were able to structure a neural network in vitro. These neurons were compartmentalized within six nodes interconnected through axon growth tunnels. Furthermore, we demonstrate the additional benefit of open top design by establishing a 3D neural culture in matrigel and a neuronal aggregate 3D culture within the chips. In conclusion, our results demonstrate a novel microfluidic chip design applicable to structuring complex neural networks in vitro, thus providing a versatile, highly relevant platform for the study of neural network dynamics applicable to developmental and regenerative neuroscience.

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The human brain is a remarkable computing machine, i.e. vastly parallel, self-organizing, robust, and energy efficient. To gain a better understanding into how the brain works, a cyborg (cybernetic organism, a combination of machine and living tissue) is currently being made in an interdisciplinary effort, known as the Cyborg project. In this paper we describe how living cultures of neurons (biological neural networks) are successfully grown in-vitro over Micro-Electrode Arrays (MEAs), which allow them to be interfaced to a robotic body through electrical stimulation and neural recordings. Furthermore, we describe the bio-and nano-technological procedures utilized for the culture of such dissociated neural networks and the interface software and hardware framework used for creating a closed-loop hybrid neuro-system. A Reservoir Computing (RC) approach is used to harness the computational power of the neuronal culture.

It has been hypothesized that the brain optimizes its capacity for computation by self-organizing to a critical point. The dynamical state of criticality is achieved by striking a balance such that activity can effectively spread through the network without overwhelming it and is commonly identified in neuronal networks by observing the behavior of cascades of network activity termed “neuronal avalanches.” The dynamic activity that occurs in neuronal networks is closely intertwined with how the elements of the network are connected and how they influence each other's functional activity. In this review, we highlight how studying criticality with a broad perspective that integrates concepts from physics, experimental and theoretical neuroscience, and computer science can provide a greater understanding of the mechanisms that drive networks to criticality and how their disruption may manifest in different disorders. First, integrating graph theory into experimental studies on criticality, as is becoming more common in theoretical and modeling studies, would provide insight into the kinds of network structures that support criticality in networks of biological neurons. Furthermore, plasticity mechanisms play a crucial role in shaping these neural structures, both in terms of homeostatic maintenance and learning. Both network structures and plasticity have been studied fairly extensively in theoretical models, but much work remains to bridge the gap between theoretical and experimental findings. Finally, information theoretical approaches can tie in more concrete evidence of a network's computational capabilities. Approaching neural dynamics with all these facets in mind has the potential to provide a greater understanding of what goes wrong in neural disorders. Criticality analysis therefore holds potential to identify disruptions to healthy dynamics, granted that robust methods and approaches are considered.

In this work, we report the preliminary analysis of the electrophysiological behavior of in vitro neuronal networks to identify when the networks are in a critical state based on the size distribution of network-wide avalanches of activity. The results presented here demonstrate the importance of selecting appropriate parameters in the evaluation of the size distribution and indicate that it is possible to perturb networks showing highly synchronized-or supercritical-behavior into the critical state by increasing the level of inhibition in the network. The classification of critical versus non-critical networks is valuable in identifying networks that can be expected to perform well on computational tasks, as criticality is widely considered to be the state in which a system is best suited for computation. This type of analysis is expected to enable the identification of networks that are well-suited for computation and the classification of networks as perturbed or healthy. This study is part of a larger research project, the overarching aim of which is to develop computational models that are able to reproduce target behaviors observed in in vitro neuronal networks. These models will ultimately be used to aid in the realization of these behaviors in nanomagnet arrays to be used in novel computing hardwares.