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A sizeable proportion of drug attrition is due to drug-induced seizures. Current available animal models frequently fail to predict human seizure liability. Therefore, there is a need for in vitro alternatives, preferably based on human-derived neurons to circumvent interspecies translation. The increasing number of commercially available human induced pluripotent stem cell (hiPSC)-derived neuronal models holds great promise for replacing rodent primary cultures. We therefore tested three different hiPSC-derived neuronal models for their applicability for in vitro seizure liability assessment. Using immunofluorescent staining and multi-well micro-electrode arrays we show that all models develop functional neuronal networks that exhibit spontaneous activity and (network) bursting behavior. Developmental patterns differ between the models, probably due to differences in model composition and seeding density. Nevertheless, neuronal activity and (network) bursting can be reproducibly modulated with the seizurogenic compounds strychnine, picrotoxin (PTX) and 4-aminopyridine (4-AP). However, the sensitivity and degree of chemical-induced effects differs between the models, which can likely be explained by differences in seeding density, maturation and different ratios of inhibitory and excitatory cell types. Importantly, compared to rat primary cortical neurons, the hiPSC-derived neuronal models were equally, or even better in the case of 4-AP, suited to detect seizurogenicity. Overall, our data indicate that hiPSC-derived neuronal models may in the future be used as a first screening tool for in vitro seizure liability assessment. However, before hiPSC-derived neuronal models can fully replace animal experiments, more compounds should be tested and the available models must be further characterized to fully understand their applicability.

Synthetic hydrogel materials can recapitulate the natural cell microenvironment; however, it is equally necessary that the gels maintain cell viability and phenotype while permitting reisolation without stress, especially for use in the stem cell field. Here, we describe a family of synthetically accessible, squaramide-based tripodal supramolecular monomers consisting of a flexible tris(2-aminoethyl)amine (TREN) core that self-assemble into supramolecular polymers and eventually into self-recovering hydrogels. Spectroscopic measurements revealed that monomer aggregation is mainly driven by a combination of hydrogen bonding and hydrophobicity. The self-recovering hydrogels were used to encapsulate NIH 3T3 fibroblasts as well as human-induced pluripotent stem cells (hiPSCs) and their derivatives in 3D. The materials reported here proved cytocompatible for these cell types with maintenance of hiPSCs in their undifferentiated state essential for their subsequent expansion or differentiation into a given cell type and potential for facile release by dilution due to their supramolecular nature.