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In vitro models that are relevant in their physiological behavior and allow spatially segregated interrogation of motor neuron somata and axons, are critical in advancing our understanding of peripheral nerve injury and disease, and in developing novel curing or regenerating therapies. Accordingly, there is an increased research interest in studying axonal repair after peripheral nerve injury, as well as in studying axonal pathology in ALS. Although exact causes are still unknown, aberrances in axonal stability, axonal transport, and axon growth dynamics have been reported amongst other hallmarks 5, 6, 7, 8, 9, 10. In amyotrophic lateral sclerosis (ALS), upper and lower motor neurons undergo progressive degeneration leading to muscle denervation, paralysis and death within 3–5 years after diagnosis 4, 5. Disease is another cause of disrupted motor neuron and axon function. Although peripheral nerves are able to regenerate after injury, complete recovery does not occur 1, 2, 3. Impaired motor neuron function due to nerve injury or disease, affects voluntary movement and can cause paralysis. Motor neurons project their axons to innervate other neurons and smooth- and skeletal muscle, allowing muscles in the body to contract. The model will thus prove ideal for studying axonal biology and disease, drug discovery and regenerative medicine. The platform comprises 40 chips arranged underneath a microtiter plate providing both throughput and compatibility to standard laboratory equipment. Importantly, outgrowing axons could be attracted and repelled through a gradient of axonal guidance cues, such as semaphorins. The model was shown susceptible to excitotoxicity upon exposure with excess glutamate and showed formation of stress granules upon excess glutamate or sodium arsenite exposure, mimicking processes common in motor neuron diseases.
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Axonal outgrowth could be precisely quantified and was shown to respond to the chemotherapeutic drug vincristine in a highly reproducible dose-dependent manner. Neurons extend their axons into an adjacent layer of gel, whereas dendrites and soma remain predominantly in the somal compartment, as verified by immunofluorescent staining. iPSC-derived motor neurons are cultured in extracellular matrix gel in a microfluidic platform. We report a method to generate a 3D motor neuron model with segregated and directed axonal outgrowth.