Characterization of a Putative Molecular Activator of Spreading Depolarization Generated by the Ischemic Brain
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Authors
Lowry, Chloe
Date
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thesis
Language
eng
Keyword
Spreading depolarization , Stroke , Ischemia , Na+/K+-ATPase , Palytoxin , Ouabain , Red blood cells
Alternative Title
Abstract
Immediately following brain ischemia, failure of the Na+/K+-ATPase leads to a formidable inward cationic current through an unidentified channel which drives spreading depolarization (SD). Recurrent SD events contribute to tissue injury following stroke or traumatic brain injury and therefore worsen clinical outcomes. Unfortunately, the molecular scenario underlying SD initiation and propagation is unknown. Palytoxin (PLTX) is a marine poison that converts the Na+/K+-ATPase into an open, depolarizing channel. Nanomolar concentrations of PLTX induce SD in live brain slices. Therefore, we used this compound to investigate if metabolically stressed gray matter releases an endogenous molecular activator of SD (SDa), which opens the Na+/K+-ATPase in a PLTX-like manner to promote SD. We also examined whether the putative SDa might evoke afferent pain and investigated its possible chemical identity. To this end, we generated a solution containing biomolecules (including the putative SDa) released from rat brain slices undergoing SD due to oxygen-glucose deprivation (OGD) or hyperthermia (Ht). Using an ex vivo hemolysis assay, we first confirmed that PLTX evoked red blood cell (RBC) swelling and lysis in a concentration-dependent manner. We found that Post-SD(OGD) and Post-SD(Ht) fractions also increased swelling and hemolysis, although a very low amount of PLTX priming (0.005–0.02 nM) was required. These results suggest that the putative SDa acts on the Na+/K+-ATPase to disrupt ionic gradients, causing RBC swelling that progresses to hemolysis. Next, we assessed whether Post-SD(OGD) and Post-SD(Ht) fractions induced mechanical or thermal hypersensitivity in mice. The results suggest that the putative SDa does not activate nociceptors during SD to evoke the subsequent migraine pain. Finally, mass spectrometry analysis of brain slices midway through SD or following ischemic stroke identified several possible lipid species associated with SD propagation, one or more of which could be SDa candidates. Overall, our findings further support the concept of metabolically stressed gray matter releasing a putative SDa that initiates and promotes SD propagation, and provides insight into its mechanism of action and possible chemical identity. Ultimately, the isolation and identification of such a compound may yield novel therapeutic targets to reduce SDs and the subsequent tissue damage following ischemic brain injury.
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ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.