Toward Brain-on-a-Chip: Human Induced Pluripotent Stem Cell-Derived Guided Neuronal Networks in Tailor-Made 3D Nanoprinted Microscaffolds

Abstract

Brain-on-a-chip (BoC) concepts should consider three-dimensional (3D) scaffolds to mimic the 3D nature of the human brain not accessible by conventional planar cell culturing. Furthermore, the essential key to adequately address drug development for human pathophysiological diseases of the nervous system, such as Parkinson’s or Alzheimer’s, is to employ human induced pluripotent stem cell (iPSC)-derived neurons instead of neurons from animal models. To address both issues, we present electrophysiologically mature human iPSC-derived neurons cultured in BoC applicable microscaffolds prepared by direct laser writing. 3D nanoprinted tailor-made elevated cavities interconnected by freestanding microchannels were used to create defined neuronal networksas a proof of conceptwith two-dimensional topology. The neuronal outgrowth in these nonplanar structures was investigated, among others, in terms of neurite length, size of continuous networks, and branching behavior using z-stacks prepared by confocal microscopy and cross-sectional scanning electron microscopy images prepared by focused ion beam milling. Functionality of the human iPSC-derived neurons was demonstrated with patch clamp measurements in both current- and voltage-clamp mode. Action potentials and spontaneous excitatory postsynaptic currentsfundamental prerequisites for proper network signalingprove full integrity of these artificial neuronal networks. Considering the network formation occurring within only a few days and the versatile nature of direct laser writing to create even more complex scaffolds for 3D network topologies, we believe that our study offers additional approaches in human disease research to mimic the complex interconnectivity of the human brain in BoC studies.

Culturing human iPSC-derived neural progenitor cells on nanowire arrays: mapping the impact of nanowire length and array pitch on proliferation, viability, and membrane deformation

“…[16][17][18][19] The influence of the substrate in terms of chemical and topological properties has been tested, for example, for adhesion, proliferation, viability, migration, and guidance of seeded cells. [20][21][22][23][24][25][26] In this context, a particular subset of substrates featuring upright arranged high aspect ratio nanostructures-so-called nanowire (NW) arraysplay an increasingly important role. [27][28][29][30][31][32][33] Nanowire arrays can affect biological parameters such as cellular growth, viability, morphology, and mechanotransduction machinery, [34][35][36][37][38][39] while electrophysiological parameters are maintained.…”

Current status of ischemic stroke treatment: From thrombolysis to potential regenerative medicine

“…For example, as damage/loss of endothelial glycocalyx contributes to additional brain damage during reperfusion; perhaps artificial glycocalyx could be transplanted to ameliorate this in the future [ 106 ]. Additionally, in vitro disease modeling, such as brain/body-on-a-chip methods, is also useful, especially in determining the effect of cytokines and the efficacy of conditioned media [ 47 , 109 ]. IPSCs are particularly useful for this purpose, given their endless proliferative abilities and their ability to tailor disease modeling to any patient [ 109 ].…”

“…Additionally, in vitro disease modeling, such as brain/body-on-a-chip methods, is also useful, especially in determining the effect of cytokines and the efficacy of conditioned media [ 47 , 109 ]. IPSCs are particularly useful for this purpose, given their endless proliferative abilities and their ability to tailor disease modeling to any patient [ 109 ].…”

Organs-on-a-chip models for biological research

“…Through these features, OOCs provide improved consistency of tissue structure and phenotypes for studies of organ-level functions, while often incorporating only a few cell types found in the native organ. Examples of OOCs include heart muscle (Mathur et al, 2015;Ronaldson-Bouchard et al, 2018;Zhao et al, 2019;Tiburcy, 2017), liver ((Kostrzewski et al, 2019;Schepers et al, 2016) alveolar unit of the lung (Huh et al, 2010), brain (Harberts et al, 2020) and blood-brain barrier (Vatine et al, 2019), kidney glomerulus and proximal tubule (Zhou et al, 2016;Homan et al, 2016), neuromuscular junction (Afshar Bakooshli et al, 2019), vascular network (Zhang et al, 2016), skin (Abaci et al, 2018), retina (Achberger et al, 2019), pancreas (Bauer et al, 2017), gut (Kim et al, 2016), bone marrow (Chou et al, 2020) placenta (Blundell et al, 2016), and tumors (Lai et al, 2020), all of which are being used to study tissue maturation, regeneration, and disease.…”