Generation of Human Motor Units with Functional Neuromuscular Junctions in Microfluidic Devices
Scope of the method
- Human health
- Basic Research
- In vitro - Ex vivo
- Human derived cells / tissues / organs
- Microfluidic device
- Human iPSC-derived motor neuron
- Human primary mesoangioblast-derived myotube
- Neuromuscular junction
- live-cell imaging
- Scanning electron microscopy
- Amyotrophic lateral sclerosis
- Neurite outgrowth
- Neurite regrowth
- Tubastatin A
This study aimed to create a versatile and reproducible in vitro model of a human motor unit with functional neuromuscular junctions (NMJs). Therefore, human induced pluripotent stem cell (hiPSC)-derived motor neurons and human primary mesoangioblast (MAB)-derived myotubes were co-cultured in commercially available microfluidic devices. The use of fluidically isolated micro-compartments allows for the maintenance of cell-specific microenvironments while permitting cell-to-cell contact through microgrooves. By applying a chemotactic and volumetric gradient, the growth of motor neuron-neurites through the microgrooves promoting myotube interaction and the formation of NMJs were stimulated. These NMJs were identified immunocytochemically through co-localization of motor neuron presynaptic marker synaptophysin (SYP) and postsynaptic acetylcholine receptor (AChR) marker α-bungarotoxin (Btx) on myotubes and characterized morphologically using scanning electron microscopy (SEM). The functionality of the NMJs was confirmed by measuring calcium responses in myotubes upon depolarization of the motor neurons. The motor unit generated using standard microfluidic devices and stem cell technology can aid future research focusing on NMJs in health and disease.
- Laminar flow cabinet
- Cell-culture incubator
- Published in peer reviewed journal
Pros, cons & Future potential
This method uses commercially available microfluidic devices and standard stem cell technology, which increases reproducibility.
The model is validated for research in amyotrophic lateral sclerosis, but can also be used in other fields where motor units or neuromuscular junctions are of interest.
References, associated documents and other information
- - Stoklund Dittlau K et al.. Human motor units in microfluidic devices are impaired by FUS mutations and improved by HDAC6 inhibition. Stem Cell Reports, 2021 Sep 14;16(9):2213-2227. doi: 10.1016/j.stemcr.2021.03.029
- - Stoklund Dittlau K et al. Generation of Human Motor Units with Functional Neuromuscular Junctions in Microfluidic Devices. J Vis Exp. 2021 Sep 7;(175). doi: 10.3791/62959. Protocol includes a professional instruction video.
Contact personKatarina Stoklund Dittlau
OrganisationsVlaams Instituut voor Biotechnologie (VIB)
VIB-Center for Brain and Disease Research, Laboratory of Neurobiology
Katholieke Universiteit Leuven (KUL)
Department of Neurosciences
Neurosciences - KU Leuven
Department of Neurosciences