Date of Award

2015

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Bioengineering

First Advisor

Beth A. Winkelstein

Abstract

Peripheral neural trauma is known to induce blood-spinal cord barrier (BSCB) breakdown, but if and how BSCB breakdown contributes to pain is unknown. The studies in this thesis use rat models of cervical nerve root insults (compressive and inflammatory) to investigate which components of injury contribute to BSCB disruption and if BSCB disruption controls the pain responses. Further, whether serum-derived thrombin extravasates into the spinal parenchyma during BSCB breakdown and contributes to the associated spinal astrocyte activation and pain through its activation of PAR1 is investigated. BSCB breakdown occurs transiently early after root compression exclusively when pain develops; blocking BSCB breakdown with activated protein C prevents pain. During increased BSCB permeability, thrombin acts enzymatically in the spinal cord. By inhibiting spinal thrombin with intrathecal hirudin before a painful compression, and separately by administering exogenous rat thrombin intrathecally to naïve rats, spinal thrombin is found to contribute to BSCB breakdown, spinal astrocyte activation and pain. Additionally, spinal thrombin initiates spinal neuroinflammation and pain through its activation of PAR1, which is determined using pharmacological inhibition of spinal PAR1 with SCH79797. In contrast to mammalian thrombin, studies using the same painful in vivo model and complementary endothelial and astrocytic in vitro experiments also demonstrate that salmon thrombin exerts opposite effects on vascular permeability, astrocytic inflammation, and pain. Using synthetic fluorogenic peptides and in silico protein models, the vascular protection and anti-inflammatory effects of salmon thrombin are hypothesized to be controlled by its unique affinity for protein C over PAR1. Lastly, spinal astrocytic and endothelial expression of the intermediate filament, vimentin, are shown to undergo a delayed increase at day 7 after a painful compression, suggesting that spinal cells may modulate their cellular mechanics in response to a painful neural injury. These studies identify how one pathological response, BSCB breakdown, that occurs in the spinal cord early after painful injury initiates a host of immediate and delayed biochemical and biomechanical responses that contribute to pain development. Findings further establish the powerful, yet diverse, role of spinal thrombin and its ability to initiate pathological or protective effects based on the signaling cascades it activates.

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