Development of two-photon polymerised scaffolds for optical interrogation and neurite guidance of human iPSC-derived cortical neuronal networks

Crowe, James A; El-Tamer, Ayman; Nagel, David A.; Koroleva, Anastasia; Madrid‐Wolff, Jorge; Olarte, Omar E.; Sokolovsky, S.G.; Estévez-Priego, Estefanía; Ludl, Adriaan-Alexander; Soriano, Jordi; Loza-Álvarez, Pablo; Chichkov, Boris N.; Hill, Eric J.; Parri, H. Rheinallt; Rafailov, Edik U.

doi:10.1039/c9lc01209e

Cited by 21 publications

(17 citation statements)

References 96 publications

(122 reference statements)

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“…Triangles: The triangle-shaped obstacles were designed to mimic the geometry of experimental scaffold structures (Crowe et al, 2020 ). Triangles were of isosceles shape with 50 μm height and 20 μm width.…”

Section: Methodsmentioning

confidence: 99%

“…They were designed to facilitate the trapping or deflection of axons (crosses), to gently modulate connectivity across the network (circles) and to dictate the directionality of connectivity (triangles). We selected these shapes in view of recent experimental studies aimed at guiding neuronal connectivity through microfabrication technology (Crowe et al, 2020 ). We observed that the obstacles molded structural connectivity at short and long length scales.…”

Section: Introductionmentioning

confidence: 99%

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Impact of Physical Obstacles on the Structural and Effective Connectivity of in silico Neuronal Circuits

Ludl

Soriano

2020

Front. Comput. Neurosci.

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Scaffolds and patterned substrates are among the most successful strategies to dictate the connectivity between neurons in culture. Here, we used numerical simulations to investigate the capacity of physical obstacles placed on a flat substrate to shape structural connectivity, and in turn collective dynamics and effective connectivity, in biologically-realistic neuronal networks. We considered μ-sized obstacles placed in mm-sized networks. Three main obstacle shapes were explored, namely crosses, circles and triangles of isosceles profile. They occupied either a small area fraction of the substrate or populated it entirely in a periodic manner. From the point of view of structure, all obstacles promoted short length-scale connections, shifted the in- and out-degree distributions toward lower values, and increased the modularity of the networks. The capacity of obstacles to shape distinct structural traits depended on their density and the ratio between axonal length and substrate diameter. For high densities, different features were triggered depending on obstacle shape, with crosses trapping axons in their vicinity and triangles funneling axons along the reverse direction of their tip. From the point of view of dynamics, obstacles reduced the capacity of networks to spontaneously activate, with triangles in turn strongly dictating the direction of activity propagation. Effective connectivity networks, inferred using transfer entropy, exhibited distinct modular traits, indicating that the presence of obstacles facilitated the formation of local effective microcircuits. Our study illustrates the potential of physical constraints to shape structural blueprints and remodel collective activity, and may guide investigations aimed at mimicking organizational traits of biological neuronal circuits.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Impact of Physical Obstacles on the Structural and Effective Connectivity of in silico Neuronal Circuits

Ludl

Soriano

2020

Front. Comput. Neurosci.

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“…Calcium imaging experiment, as adapted from Ozdener et al [ 12 ], was, accordingly, performed to investigate whether cultured BTBCs still retain their physiological functions of taste responsiveness post their isolation and cryopreservation. The dynamic calcium imaging experiments were conducted and analysed according to standard approaches used in our laboratory [ 18 , 19 ]. Cells were exposed for 50–100 ms depending on loading efficiency of particular culture, this was kept constant for each experiment.…”

Section: Methodsmentioning

confidence: 99%

Novel, Fully Characterised Bovine Taste Bud Cells of Fungiform Papillae

Ftuwi

Parri

Mohammed

2021

Cells

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Current understanding of functional characteristics and biochemical pathways in taste bud cells have been hindered due the lack of long-term cultured cells. To address this, we developed a holistic approach to fully characterise long term cultured bovine taste bud cells (BTBCs). Initially, cultured BTBCs were characterised using RT-PCR gene expression profiling, immunocytochemistry, flowcytometry and calcium imaging, that confirmed the cells were mature TBCs that express taste receptor genes, taste specific protein markers and capable of responding to taste stimuli, i.e., denatonium (2 mM) and quinine (462.30 μM). Gene expression analysis of forty-two genes implicated in taste transduction pathway (map04742) using custom-made RT-qPCR array revealed high and low expressed genes in BTBCs. Preliminary datamining and bioinformatics demonstrated that the bovine α-gustducin, gustatory G-protein, have higher sequence similarity to the human orthologue compared to rodents. Therefore, results from this work will replace animal experimentation and provide surrogate cell-based throughput system to study human taste transduction.

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“…In most biocompatible photoresist systems such as hydrogel-based natural or artificial materials, they also need to be soluble and thermally stable in the monomer aqueous microenvironments [42]. Currently, a lot of PIs, such as rose bengal, methylene blue, lithium phenyl-2,4, 6-trimethyibenzoylphosphinate (LAP) and Irgacure 369, are available commercially and commonly used for fabricating micro/nano-structures for biomedical applications [26,36,[50][51][52]. In Table 1, PIs for the TPP fabrication are summarized along with their water solubility, δ TPA , and biomedical applications as reported in some studies.…”

Section: Introductionmentioning

confidence: 99%

Two-photon polymerization nanolithography technology for fabrication of stimulus-responsive micro/nano-structures for biomedical applications

Huang

Tsui

Deng

et al. 2020

Nanotechnology Reviews

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Micro/nano-fabrication technology via two-photon polymerization (TPP) nanolithography is a powerful and useful manufacturing tool that is capable of generating two dimensional (2D) to three dimensional (3D) arbitrary micro/nano-structures of various materials with a high spatial resolution. This technology has received tremendous interest in cell and tissue engineering and medical microdevices because of its remarkable fabrication capability for sophisticated structures from macro- to nano-scale, which are difficult to be achieved by traditional methods with limited microarchitecture controllability. To fabricate precisely designed 3D micro/nano-structures for biomedical applications via TPP nanolithography, the use of photoinitiators (PIs) and photoresists needs to be considered comprehensively and systematically. In this review, widely used commercially available PIs are first discussed, followed by elucidating synthesis strategies of water-soluble initiators for biomedical applications. In addition to the conventional photoresists, the distinctive properties of customized stimulus-responsive photoresists are discussed. Finally, current limitations and challenges in the material and fabrication aspects and an outlook for future prospects of TPP for biomedical applications based on different biocompatible photosensitive composites are discussed comprehensively. In all, this review provides a basic understanding of TPP technology and important roles of PIs and photoresists for fabricating high-precision stimulus-responsive micro/nano-structures for a wide range of biomedical applications.

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Development of two-photon polymerised scaffolds for optical interrogation and neurite guidance of human iPSC-derived cortical neuronal networks

Abstract: We identified photopolymers for 2-photon polymerisation with biocompatibility for human iPSC-derived neural network development. Generation of microscale scaffold topologies enabled neurite guidance, demonstrating use in reproducing aligned networks.

Cited by 21 publications

References 96 publications

Impact of Physical Obstacles on the Structural and Effective Connectivity of in silico Neuronal Circuits

Impact of Physical Obstacles on the Structural and Effective Connectivity of in silico Neuronal Circuits

Novel, Fully Characterised Bovine Taste Bud Cells of Fungiform Papillae

Two-photon polymerization nanolithography technology for fabrication of stimulus-responsive micro/nano-structures for biomedical applications

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