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Title: Investigating AMPA Receptor Trafficking In Post-Ischaemic Hippocampal Neurons
Author: Dixon, Rebecca Mary
ISNI:       0000 0001 3426 1370
Awarding Body: University of Bristol
Current Institution: University of Bristol
Date of Award: 2007
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Regulated AMPA receptor trafficking at excitatory synapses plays an important role in changes in synaptic strength such as long-term potentiati,on (LTP) and long-term depression (LTD). GluR2 is a key AMPA receptor subunit present in the majority of AMPA receptors in the adult rat hippocampus. In the absence of GluR2, AMPARs exhibit enhanced Ca2+ influx and decreased current at positive potentials due to an intracellular polyamine block. I investigated AMPA receptor trafficking in hippocampal neurons following oxygen-glucose deprivation (OGD), an in vitro model of ischaemia. I carried out whole-cell recordings from CA1 pyramidal neurons and measured the rectification index (RI: +40 mV EPSC/-70 mV EPSC) before and after DGD. My results show a 30 minute period of DGD results in a decrease in RI, suggesting replacement of GluR2containing AMPARs with GluR2-lacking AMPARs. To investigate direct changes in AMPAR subunit composition during OGD, surface biotinylation and Western blot analysis were carried out on dissociated hippocampal cultures. A 30 minute period ofOGD led to a reduction in surface GluR2 protein levels, but no change in surface GluRI, in support of the electrophysiology results. This data suggests a rapid switch in AMPAR subunit composition is occurring following DGD, which may be linked to the more prolonged expression of GluR2-lacking AMPARs that occurs 24-48 hours following ischaemia. To investigate the molecular mechanism for this trafficking event, I postsynaptically infused peptides that interfere with GluR2 C-terminal interactions with PDZ domain proteins, via the patch pipette. Infusion of pep2-EVKI, which selectively blocks the PICKI-GluR2 interaction, prevented the OGD-induced decrease in RI, whilst a control peptide, pep2SVKE, had no effect. Furthermore, expressing pep2-EVKI from Sindbis virus blocked the DGD-mediated decrease in surface GluR2 in dissociated hippocampal cultures. This shows PICKl is a crucial mediator of rapid AMPA receptor trafficking in post-ischaemic hippocampal neurons, and presents the GluR2-PICKl interaction as a potential therapeutic target for ischaemic stroke.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available