All ETDs from UAB

Advisory Committee Chair

Harald Sontheimer

Advisory Committee Members

Dale Benos

Michael Brenner

Lisa Guay-Woodford

John Hablitz

Document Type


Date of Award


Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine


According to the Central Brain Tumor Registry of the United States, the most common primary brain tumors are gliomas, tumors composed of cells of glial origin, most commonly astrocytes and oligodendrocytes. The most aggressive of these tumors are characterized by hyperproliferation, marked cellular and nuclear atypia, extensive infiltration into surrounding normal brain tissue, and large areas of cell and tissue death. Previous data published by our lab and others suggest that these biological processes may involve regulated cell volume changes. Using cell volume regulation in the presence of an anisosmotic challenge as a model for cell swelling and shrinkage, cell volume changes have been shown to involve the movement of molecules, or osmolytes, across the plasma membrane through channels and transporters. Water is osmotically obliged to follow the net movement of molecules, resulting in a net flux of water across the plasma membrane and an overall change in cell volume. As the most abundant anion in biological systems, chloride has been shown to be involved in the volume regulation of several cell types. However, chloride-mediated volume changes in human glioma cells have not been extensively studied. It is the primary goal of this dissertation to examine the mechanisms employed by human glioma cells to dynamically regulate their cell volume at rest, in the presence of anisosmotic conditions, and importantly during the biological processes of apoptosis and migration. We confirm the expression of the voltage gated chloride channels ClC-2, 3, and 5 in human glioma cell lines and patient biopsies. In addition, we ii demonstrate the expression of the cation chloride cotransporters, KCC1, KCC3, and NKCC1. Using a variety of techniques, including electrophysiology, Coulter Counter, and chloride-sensitive fluorescent dyes, we establish that the resting intracellular chloride concentration and cell volume are maintained by the basal activity of chloride cotransporters. While pharmacological inhibition of these cotransporters suggests that they are also involved in cell volume regulation during an aniosomotic challenge, chloride efflux through channels plays a more significant role in post-hyposmotic volume decrease. Similarly, inhibition of chloride channels, but not chloride cation cotransporters, inhibits the cell condensation that occurs in the presence of apoptotic stimuli. Cells treated with chloride channel inhibitors also demonstrated limited caspase 3 activity and DNA fragmentation, suggesting that the volume decrease is necessary for apoptosis. Finally, transwell migration of human glioma cells was blocked in the presence of chloride channel and transport inhibitors, suggesting that the mechanisms involved in cell shrinkage are necessary in glioma cell migration.



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