Microfluidic culture platform for studying neuronal response to mild to very mild axonal stretch injury.
A new model for studying localised axonal stretch injury is presented, using a microfluidic device to selectively culture axons on a thin, flexible poly (dimethylsiloxane) membrane which can be deflected upward to stretch the axons. A very mild (0.5% strain) or mild stretch injury (5% strain) was applied to primary cortical neurons after 7 days growth in vitro. The extent of distal degeneration was quantified using the degenerative index (DI, the ratio of fragmented axon area to total axon area) of axons fixed at 24 h and 72 h post injury (PI), and immunolabelled for the axon specific, microtubule associated protein-tau. At 24 h PI following very mild injuries (0.5%), the majority of the axons remained intact and healthy with no significant difference in DI when compared to the control, but at 72 h PI, the DI increased significantly (DI = 0.11 ± 0.03). Remarkably, dendritic beading in the somal compartment was observed at 24 h PI, indicative of dying back degeneration. When the injury level was increased (5% stretch, mild injury), microtubule fragmentation along the injured axons was observed, with a significant increase in DI at 24 h PI (DI = 0.17 ± 0.02) and 72 h PI (DI = 0.18 ± 0.01), relative to uninjured axons. The responses observed for both mild and very mild injuries are similar to those observed in the in vivo models of traumatic brain injury, suggesting that this model can be used to study neuronal trauma and will provide new insights into the cellular and molecular alterations characterizing the neuronal response to discrete axonal injury.
Yiing C
YC
Yap
Menzies Research Institute, University of Tasmania , Private Bag 23, Hobart, Tasmania 7000, Australia.
Tracey C
TC
Dickson
Wicking Dementia Research and Education Centre, University of Tasmania , Private Bag 23, Hobart, Tasmania 7000, Australia.
Anna E
AE
King
Australian Center for Research on Separation Science (ACROSS), School of Physical Sciences, University of Tasmania , Private Bag 75, Hobart, Tasmania 7001, Australia.
Michael C
MC
Breadmore
Pharmacy School of Medicine, ACROSS, University of Tasmania , Private Bag 26, Hobart, Tasmania 7001, Australia.
Rosanne M
RM
Guijt