Date of Award
Fall 2012
Document Type
Dissertation
Degree Name
Doctor of Philosophy (PhD)
Department
Biological Sciences
First Advisor
Lobner, Doug
Second Advisor
Baker, David
Third Advisor
Buchanan, James
Abstract
The main focus of the studies in this thesis involves examining the role of cystine/glutamate exchange (system xC-) in neuronal death in primary cortical cell culture, with an emphasis on how glial function affects neuronal cell death. System xC- is a sodium-independent transporter that mediates cystine uptake and glutamate release. It accounts for most of the cystine uptake in astrocytes in mature cultures, providing the rate limiting substrate for synthesis of the main endogenous antioxidant glutathione. The glutamate released by system xC- may lead to excessive extracellular glutamate and cause excitotoxicity.
β-N-methylamino-L-alanine (BMAA) is a non-protein amino acid that may be involved in neurodegenerative diseases. We found that BMAA induced oxidative stress by competing with cystine at system xC- leading to depletion of glutathione. BMAA also drives system xC- mediated glutamate release, which may contribute to its induction of excitotoxicity.
Fibroblast growth factor-2 (FGF-2) is involved in multiple processes in the central nervous system, including plasticity, neurogenesis, differentiation, and neuronal survival. Also, alterations in FGF-2 and its signaling have been implicated in neurodegenerative diseases and psychiatric disorders. We found that FGF-2 greatly increased cystine uptake through system xC- in astrocyte-enriched primary cultures, but not in neuronal or microglial cultures. Our data showed that FGF-2 increased cystine uptake by upregulating system xC- by acting on FGFR1, and signaling through the PI3K/Akt and MEK/ERK pathways.
FGF-2 treatment for 48 hours caused significant neuronal death only in mixed neuronal and glial cultures, but not in neuronal-enriched or astrocyte-enriched cultures. Blocking system xC-, or AMPA/kainate receptors, eliminated the neuronal death induced by FGF-2 treatment. Therefore, it is likely that 48 hour FGF-2 treatment induces AMPA receptor mediated toxicity through increased glutamate release from astrocytes due to increased system xC- function. However, we cannot exclude the possibility that FGF-2 treatment sensitizes the neurons to normal system xC- mediated glutamate release.
Together the results indicate that 1) competitive substrates of system xC-, such as BMAA, that do not lead to glutathione production are particularly toxic; and 2) upregulation of system xC- on astrocytes may be toxic to surrounding neurons.