![]() ![]() We then validated a protocol wherein the electrical activity of motor neural cultures is measured directly by a voltage sensitive dye and a microplate reader without causing damage to the cells. Neurons function by transmitting electrical signals to one another and being able to assess the development of electrical signaling serves is an important verification step when engineering neural tissues. In this method, spherical indenters are positioned on top of the fibrin samples, generating an indentation depth that is then correlated with elastic modulus. Here we validate a direct method for acquiring elastic modulus of fibrin using a modified Hertz model for thin films. Neurons require soft substrates to differentiate and mature, however measuring the elastic moduli of soft substrates remains difficult to accurately measure using standard protocols such as atomic force microscopy or shear rheology. The protocols obtain elastic moduli of very soft fibrin hydrogel scaffolds and voltage readings from motor neuron cultures. ![]() These protocols characterize the mechanical properties of engineered neural tissues and measure their electrophysical activity. We have designed and validated a set of robust and non-toxic protocols for directly evaluating the properties of engineered neural tissue. We anticipate that our findings may help developing better models for the study of cerebellar dysfunctions, while providing an advance toward the development of autologous replacement strategies for treating cerebellar degenerative diseases. Using a defined basal medium optimized for neuronal cell culture, we successfully promoted the differentiation of cerebellar precursors without the need for co-culturing. Here we describe a novel differentiation strategy that uses defined medium to generate Purkinje cells, granule cells, interneurons, and deep cerebellar nuclei projection neurons, that self-formed and differentiated into electrically active cells. ![]() Moreover, in vitro generation of Purkinje cells required co-culture systems, which may introduce unknown components to the system. However, previous studies only produced limited amounts of Purkinje cells. As an alternative to animal models, cerebellar neurons differentiated from pluripotent stem cells have been used. ![]() A major limitation in cerebellar research has been the lack of adequate disease models. The cerebellum plays a critical role in all vertebrates, and many neurological disorders are associated with cerebellum dysfunction. This new model system demonstrates the potential for enabling an increased understanding of molecular mechanisms in human rabies, which could lead to improved control methods. This study therefore defines the first stem cell-derived ex-vivo model system to study rabies pathogenesis in human neurons. In addition, we highlight specific differences in cellular pathogenesis between laboratory-adapted and field strain lyssavirus. We show key cellular features of rabies infection in our human neural cultures, including upregulation of inflammatory chemokines, lack of neuronal apoptosis, and axonal transmission of viruses in neuronal networks. In this study, we utilize advances in stem cell technology to characterize rabies infection in human stem cell-derived neurons. Lack of appropriate ex-vivo models for studying rabies infection in human neurons has contributed to this knowledge gap. Many aspects of rabies pathogenesis in human neurons are not well understood. Rabies is a zoonotic neurological infection caused by lyssavirus that continues to result in devastating loss of human life.
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