Modelling central and peripheral nervous system connectivity using compartmentalised microfluidic systems

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The mechanisms that control the development of chronic nociception are not yet fully understood. Despite recent advances, the complex interplay achieved by different neuronal types in nociceptive circuits has not been adequately modelled in vitro, with cell culture experiments ignoring the incredible anatomical polarization of dorsal root ganglia (DRG) neurons.In this context, there is a clear need to implement physiologically relevant in vitro systems that could allow the investigation of cellular and molecular mechanisms in a context of multi-cellular circuit complexity. Culture platforms with fluidically isolated compartments allow the functional separation of neuronal domains in more physiologically relevant contexts. We have already demonstrated the ability to replicate the "pseudo-unipolar" nature of DRG neurons in functional experiments by measuring Ca2+ signals in the cell body compartment of microfluidic systems. We will further develop these novel culture systems by sequentially adding levels of network complexity. Dorsal horn primary neurons and keratinocytes will be seeded into the lateral compartments of three-channel microfluidic devices that have DRG neurons placed in the middle channel. This ststem will model the "central" and "peripheral" domains of DRG neuron circuitry, but with greater experimental accesibility.Our preliminary work identified key miRNAs that are changed in hyperalgesic priming, an in vivo model for studying the transition from acute to chronic pain. Although local protein expression has been suggested in hyperalgesic priming mechanisms, the role of miRNAs has not been investigated. Using the proposed system, miRNA inhibitors will be added into the peripheral and central compartments while neuron excitability and expression of synaptic markers will be evaluated. This will allow us to determine the potential role of miRNAs in the control of protein expression and local regulation of neuronal circuits during nociception.