Date of Award
Microfluidics, Neuron, Glia, Drosophila, Co-Culture, Neuromuscular Junction
Neuron-glia communication is crucial to the development, plasticity, and repair of the nervous system (NS). While neurons are well known to conduct electrical impulses that transfer biological information and stimuli throughout the NS, our understanding of the roles of glia continues to evolve from when the cells were largely believed to act solely for neuronal support. Recent decades of research has shown that glia can alter metabolism, conduct impulses and change phenotype for NS repair. NG interactions have, thereby, become heavily researched in varied areas of biomedical engineering, including embryogenesis, neural regeneration, growth, and intracellular synaptic activity. However, while NG interactions are known to regulate survival, differentiation, communication, and targeted migration of neural cells, the molecular signals that orchestrate these behaviors remain incompletely understood. As a result, many emerging studies have embraced microfluidics to regulate the spatial and temporal stimuli delivered to neural cell groups and measure subsequent NG responses.
The overall objective of this thesis was to examine emergent NG behavior in response to chemical stimuli within controlled microfluidic environments. Experiments examined NG behavior in the central and peripheral NS critical to neural repair. In the first model, we examined the behavior of transformed glial progenitors (in the form of Medulloblastoma (MB)), known to emulate developmental processes, to external stimuli using controlled microenvironments. We used a microfluidic system called the bridged mlane, which allows for steady-state, 1D, controllable concentration gradients along the length of its’ microchannel. The system was used to evaluate in vitro migratory responses of MB-derived cells to external signaling from Epidermal Growth Factor (EGF) and stromal cell-derived factor 1-alpha (SDF-1). Data demonstrated that MB cells exhibit dosage-dependent chemotaxis towards increasing concentration gradients. However, as glial behaviors are intricately linked with that of neuronal cells, we next used a more comprehensive neural model to examine the collective behavior of neural progenitors in response to chemotactic stimulation. Experiments examined the collective behavior of NG progenitor cell populations in response to stimulation via fibroblast growth factor (FGF) gradients using a developmental model of the central nervous system (CNS) in the Drosophila Melanogaster, 3rd instar larvae stage. Surprisingly, our data demonstrated that cells migrated larger distances and with higher directionality within collective groups of both neuronal and glial progenitors than in populations of glia only. Taken together, these results helped elucidate different modalities for directed movement that can be used for therapeutic techniques that leverage the interdependent NG relationship.
The last model examined NG contributions to the formation of neuromuscular junctions (NMJ) in the peripheral nervous system (PNS). The glial component of the NMJ, the Schwann cells (SCs), are essential to NMJ development and function including remodeling and regeneration. SCs are critical for PNS regeneration, where studies have shown SC are able to trans-differentiate in order to create glial bridges that bypass non-functional neuronal nodes and isolate damaged neurons. However recent NMJ models mainly focus on motor neurons (MN) and muscle cells (MCs), some in vitro work has been utilized to study SCs, but their overall roles still remain to be well-defined and studied. To that end, the experiments used a compartmentalized microfluidic platform to demonstrate reproducible differentiation of skeletal myotubes with increased viability and length following the time-dependent addition of neuronal and glial cells. We lastly probed the guidance cues and migratory patterns of NGs towards various growth factors to elucidate emergent NMJ response. Our data illustrated there is a co-culture effect on receptor expression dependent on stimulation time. The data point to SCs as key players in stabilizing and maintaining in vitro NMJ models that will aid the development and testing of emerging therapies for neuromuscular dysfunction.
Singh, Tanya, "Neuron-Glial (NG) Interactions: A Microfluidic Examination of NG Emergent Responses for Repair" (2019). CUNY Academic Works.