Microfluidic Chip for Site‐Specific Neuropharmacological Treatment and Activity Probing of 3D Neuronal “Optonet” Cultures

Marom, Anat; Mahto, Sanjeev Kumar; Shor, Erez; Tenenbaum-Katan, Janna; Sznitman, Josué; Shoham, Sharon

doi:10.1002/adhm.201400643

Cited by 11 publications

(9 citation statements)

References 36 publications

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“…Here, temperature was found to be a powerful modulator of spontaneous activity patterns; when reduced, wave patterns with diverse spatial characteristics were observed. Similar effects were previously observed under cholinergic pharmacological modulation (Marom et al, 2015), which induced propagation of linear and circular waves at velocities up to 2 μm/s. Here, when lowering the temperature, waves propagated at similar velocities (periods of 2–6 s of waves were seen in culture, see Figure 7B).…”

Section: Discussionsupporting

confidence: 86%

“…In our previous studies, optonets were subjected to varying environmental conditions (see Dana et al, 2014; Marom, 2014; Marom et al, 2015 for the effects of different scaffold materials, support cells and pharmacological environments). Here, temperature was found to be a powerful modulator of spontaneous activity patterns; when reduced, wave patterns with diverse spatial characteristics were observed.…”

Section: Discussionmentioning

confidence: 99%

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Spontaneous Activity Characteristics of 3D “Optonets”

et al. 2017

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Sporadic spontaneous network activity emerges during early central nervous system (CNS) development and, as the number of neuronal connections rises, the maturing network displays diverse and complex activity, including various types of synchronized patterns. These activity patterns have major implications on both basic research and the study of neurological disorders, and their interplay with network morphology tightly correlates with developmental events such as neuronal differentiation, migration and establishment of neurotransmitter phenotypes. Although 2D neural cultures models have provided important insights into network activity patterns, these cultures fail to mimic the complex 3D architecture of natural CNS neural networks and its consequences on connectivity and activity. A 3D in-vitro model mimicking early network development while enabling cellular-resolution observations, could thus significantly advance our understanding of the activity characteristics in the developing CNS. Here, we longitudinally studied the spontaneous activity patterns of developing 3D in-vitro neural network “optonets,” an optically-accessible bioengineered CNS model with multiple cortex-like characteristics. Optonet activity was observed using the genetically encodable calcium indicator GCaMP6m and a 3D imaging solution based on a standard epi-fluorescence microscope equipped with a piezo-electric z-stage and image processing-based deconvolution. Our results show that activity patterns become more complex as the network matures, gradually exhibiting longer-duration events. This report characterizes the patterns over time, and discusses how environmental changes affect the activity patterns. The relatively high degree of similarity between the network's spontaneously generated activity patterns and the reported characteristics of in-vivo activity, suggests that this is a compelling model system for brain-in-a chip research.

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Section: Discussionsupporting

confidence: 86%

Section: Discussionmentioning

confidence: 99%

Spontaneous Activity Characteristics of 3D “Optonets”

et al. 2017

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“…Indeed, the 3D scaffold allows the formation of chemical gradients within the network, as soluble and slowly diffusing molecules could be locally trapped within the hydrogel polymer network. The integration of microfluidic systems 62 could permit the study of the effects of controlled local release of chemical compounds in a 3D geometry. In contrast, such gradients could not be easily established and maintained in the bath of 2D neuronal cultures within Petri dishes.…”

Section: Discussionmentioning

confidence: 99%

Fast wide-volume functional imaging of engineered in vitro brain tissues

Palazzolo

Moroni

Soloperto

et al. 2017

Sci Rep

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The need for in vitro models that mimic the human brain to replace animal testing and allow high-throughput screening has driven scientists to develop new tools that reproduce tissue-like features on a chip. Three-dimensional (3D) in vitro cultures are emerging as an unmatched platform that preserves the complexity of cell-to-cell connections within a tissue, improves cell survival, and boosts neuronal differentiation. In this context, new and flexible imaging approaches are required to monitor the functional states of 3D networks. Herein, we propose an experimental model based on 3D neuronal networks in an alginate hydrogel, a tunable wide-volume imaging approach, and an efficient denoising algorithm to resolve, down to single cell resolution, the 3D activity of hundreds of neurons expressing the calcium sensor GCaMP6s. Furthermore, we implemented a 3D co-culture system mimicking the contiguous interfaces of distinct brain tissues such as the cortical-hippocampal interface. The analysis of the network activity of single and layered neuronal co-cultures revealed cell-type-specific activities and an organization of neuronal subpopulations that changed in the two culture configurations. Overall, our experimental platform represents a simple, powerful and cost-effective platform for developing and monitoring living 3D layered brain tissue on chip structures with high resolution and high throughput.

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“…[268,269] By transfecting neurons with GCaMP and culturing them in 3D hydrogel matrices, multi-planar images provide spatial specificity and the functional activity of the developed 3D neural networks, particularly in response to drug input. [270,271] Additional fluorescent protein-based probes such as red GECIs (RCaMP) provide the option of using different colors within the same cell or to distinguish cell populations (e.g., neurons and astrocytes) and may be integrated with optogenetic approaches. [272] The emergence of optogenetics, starting from the use of a light-sensitive ion channel protein (Channelrhodopsin-2, ChR2), as a means of optically controlling the activity of neuronal populations has provided researchers with another tool to activate or inhibit the activity of specific neurons noninvasively in culture with high temporal and spatial resolution.…”

Section: Technology and Systems Integration In Microfluidic Platformsmentioning

confidence: 99%

“…[ 268,269 ] By transfecting neurons with GCaMP and culturing them in 3D hydrogel matrices, multi‐planar images provide spatial specificity and the functional activity of the developed 3D neural networks, particularly in response to drug input. [ 270,271 ] Additional fluorescent protein‐based probes such as red GECIs (RCaMP) provide the option of using different colors within the same cell or to distinguish cell populations (e.g., neurons and astrocytes) and may be integrated with optogenetic approaches. [ 272 ]…”

Section: Engineering Approaches To 3d Brain Modelsmentioning

confidence: 99%

Innovations in 3D Tissue Models of Human Brain Physiology and Diseases

Lovett

Nieland

Dingle

et al. 2020

Adv Funct Materials

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3D laboratory tissue cultures have emerged as an alternative to traditional 2D culture systems that do not recapitulate native cell behavior. The discrepancy between in vivo and in vitro tissue-cell-molecular responses impedes understanding of human physiology in general and creates roadblocks for the discovery of therapeutic solutions. Two parallel approaches have emerged for the design of 3D culture systems. The first is biomedical engineering methodology, including bioengineered materials, bioprinting, microfluidics, and bioreactors, used alone or in combination, to mimic the microenvironments of native tissues. The second approach is organoid technology, in which stem cells are exposed to chemical and/or biological cues to activate differentiation programs that are reminiscent of human (prenatal) development. This review article describes recent technological advances in engineering 3D cultures that more closely resemble the human brain. The contributions of in vitro 3D tissue culture systems to new insights in neurophysiology, neurological diseases, and regenerative medicine are highlighted. Perspectives on designing improved tissue models of the human brain are offered, focusing on an integrative approach merging biomedical engineering tools with organoid biology.

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Microfluidic Chip for Site‐Specific Neuropharmacological Treatment and Activity Probing of 3D Neuronal “Optonet” Cultures

Cited by 11 publications

References 36 publications

Spontaneous Activity Characteristics of 3D “Optonets”

Spontaneous Activity Characteristics of 3D “Optonets”

Fast wide-volume functional imaging of engineered in vitro brain tissues

Innovations in 3D Tissue Models of Human Brain Physiology and Diseases

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