.Glutamate is the predominant excitatory neurotransmitter in the mammalian central nervous system. Glutamate is normally cleared from the synaptic cleft by high-affinity, sodium-dependent glutamate transporters located in both neurons and glia. Glutamate transport malfunction can lead to the accumulation of excessive glutamate in the synapse, with subsequent neurotoxicity. EAAT2 is an astroglial glutamate transporter and is the predominant glutamate transporter. A loss of this protein has been found in the affected areas of Alzheimer's disease (AD) as well as in amyotrophic lateral sclerosis (ALS). Antisense knockdown studies as well as EAAT2 null mice have demonstrated that loss of EAAT2 protein can lead to excitotoxic neuronal degeneration. What could account for the selective loss of EAAT2 protein in these neurodegenerative diseases? The investigator's recent studies in ALS have demonstrated that the loss of EAAT2 is due to aberrant mRNAs, possibly as a consequence of abnormal splicing. In this study it is proposed to test the possibility that the aberrant mRNAs could account for the loss of EAAT2 in AD. Preliminary results demonstrate that aberrant EAAT2 mRNAs are present in AD specimens. The investigator will determine the abundance of the aberrant EAAT2 mRNA species and evaluate the prevalence of the aberrant EAAT2 mRNAs in AD patients and whether they are correlated with loss of EAAT2 protein. The investigator also proposes to investigate whether there are aberrant EAAT2 mRNAs in transgenic mice with APP (amyloid precursor protein) mutations. Can aberrant EAAT2 mRNAs lead to loss of EAAT2 protein and contribute to neurodegeneration in vivo? To answer this they will generate transgenic mice manifesting astrocyte-specific expression of the human wild-type and aberrant EAAT2 mRNAs. What mechanism accounts for these aberrant splicing defects? They will determine whether there are acquired somatic mutations in the EAAT2 gene. The investigator proposes to develop astrocyte cultures from AD brain and determine if the aberrant EAAT2 mRNAs can be produced in vitro.
