The XonaChip® offers advantages for culturing neurons differentiated from human stem cells. Differentiated neurons attach and distribute more evenly in the XonaChip® than in silicone-based devices, resulting in healthy cultures that can be maintained for 4-5 weeks or more.
Download Post as PDF

Introduction

Neurons differentiated from human stem cells are increasingly used in neuroscience. The unique extreme polarization of these and other post-mitotic neurons demands an approach to measure and manipulate distinct neuronal compartments. Microfabricated multicompartment devices, pioneered by Xona scientists, have become well-established and well-used research tools for neuroscientists in the last 10-15 years 1-5. These devices compartmentalize neurons and provide a method to physically and chemically manipulate subcellular regions of neurons, including somata, dendrites, axons, and synapses 6-8.

To provide an easy-to-use and fully assembled device, Xona has developed plastic XonaChip®8 (see Introducing XonaChips® for more details). In this current TechNote, researchers at UNC-Chapel Hill differentiated human neural stem cells into glutamatergic neurons and found that XonaChips® improve the long-term culture of these neurons over previous silicone-based compartmentalized devices.

Results & Discussion

Human neural stem cells (NSCs) were differentiated into neurons and neuronal projections entered the axonal compartment after one week (7-10 days) in the chip with differentiation media (Figure 1). The resulting neurons attached and distributed more evenly within the somatic compartment of the chip than in PDMS devices. Neurons in the XonaChips® had healthy bundled axons and neurons were maintained within the chips for 4-5 weeks.

Figure 1. Human stem cell differentiated neurons grown in XonaChips® and side-by-side comparison with PDMS compartmentalized devices.

Methods

XonaChips® were prepared according to the protocol. XC Pre-CoatTM was first used to ensure even wetting of the chip. XC Pre-CoatTM is included with each XonaChips®order. The chip was then coated with Poly-L-Ornithine (20 µg/ml) and laminin (10 µg/ml) before pre-conditioning with NSC media. Poly(dimethylsiloxane) (PDMS) or silicone-based compartmentalized devices were prepared according to Xona’s silicone-based devices protocol. These devices were also coated with Poly-L-Ornithine and laminin before pre-conditioning with NSC media.

H9-derived human neural stem cells (ThermoFisher Scientific, 510088) were thawed according to the manufacturer’s instructions. Approximately 70,000 NSCs were seeded into the right compartment of the XonaChips®. The same number were seeded into the right compartment of the PDMS compartmentalized device. Images were acquired with an inverted phase contrast microscope.

Conclusion

In summary, the XonaChip® is a fully assembled multicompartment device that is easy to use and results in healthy, long-lasting cultures. Importantly, as shown previously, these chips allow microenvironments to be established as with our SND series devices and are equally compatible with high-resolution fluorescence microscopy.

About Xona Microfluidics Inc

Xona Microfluidics Inc is a California LLC based in Temecula, California with R&D facilities in Research Triangle Park, North Carolina. More information can be found at xonamicrofluidics.com.

If you are interested in testing a XonaChips® contact us at info@xona.us

1          Neto, E. et al. Compartmentalized Microfluidic Platforms: The Unrivaled Breakthrough of In Vitro Tools for Neurobiological Research. The Journal of Neuroscience. 36 (46), 11573-11584, doi:10.1523/jneurosci.1748-16.2016, (2016).

2          Bigler, R. L., Kamande, J. W., Dumitru, R., Niedringhaus, M. & Taylor, A. M. Messenger RNAs localized to distal projections of human stem cell derived neurons. Sci Rep. 7 (1), 611, doi:10.1038/s41598-017-00676-w, (2017).

3          Nagendran, T. et al. Distal axotomy enhances retrograde presynaptic excitability onto injured pyramidal neurons via trans-synaptic signaling. Nat Commun. 8 (1), 625, doi:10.1038/s41467-017-00652-y, (2017).

4          Jia, L. et al. MiR-34a Regulates Axonal Growth of Dorsal Root Ganglia Neurons by Targeting FOXP2 and VAT1 in Postnatal and Adult Mouse. Molecular Neurobiology. doi:10.1007/s12035-018-1047-3, (2018).

5          Van Laar, V. S. et al. Evidence for compartmentalized axonal mitochondrial biogenesis: Mitochondrial DNA replication increases in distal axons as an early response to Parkinson’s disease-relevant stress. The Journal of Neuroscience. doi:10.1523/jneurosci.0541-18.2018, (2018).

6          Taylor, A. M. et al. A microfluidic culture platform for CNS axonal injury, regeneration and transport. Nat Methods. 2 (8), 599-605, doi:10.1038/nmeth777, (2005).

7          Taylor, A. M., Dieterich, D. C., Ito, H. T., Kim, S. A. & Schuman, E. M. Microfluidic local perfusion chambers for the visualization and manipulation of synapses. Neuron. 66 (1), 57-68, doi:10.1016/j.neuron.2010.03.022, (2010).

8          Nagendran, T., Poole, V., Harris, J. & Taylor, A. M. Use of Pre-Assembled Plastic Microfluidic Chips for Compartmentalizing Primary Murine Neurons Journal of visualized experiments : JoVE.  (in press).

WordPress Image Lightbox Plugin