The excitatory amino acid transporters (EAATs) play key roles in the regulation of CNS L-glutamate, especially related to synthesis, signal termination, synaptic spillover, and excitotoxic protection. Inhibitors available to delineate EAAT pharmacology and function are essentially limited to those that non-selectively block all EAATs or those that exhibit a substantial preference for EAAT2. Thus, it is difficult to selectively study the other subtypes, particularly EAAT1 and EAAT3. Structure activity studies on a series of β-substituted aspartate analogues identify L-β-benzyl-aspartate (L-β-BA) as among the first blockers that potently and preferentially inhibits the neuronal EAAT3 subtype. Kinetic analysis of 3H-Daspartate uptake into C17.2 cells expressing the hEAATs demonstrate that L-threo-β-BA is the more potent diastereomer, acts competitively, and exhibits a 10-fold preference for EAAT3 compared to EAAT1 and EAAT2. Electrophysiological recordings of EAAT-mediated currents in Xenopus oocytes identify L-β-BA as a non-substrate inhibitor. Analyzing L-threo-β-BA within the context of a novel EAAT2 pharmacophore model suggests: (1) a highly conserved positioning of the electrostatic carboxyl and amino groups, (2) nearby regions that accommodate select structural modifications (cyclopropyl rings, methyl groups, oxygen atoms), and (3) a unique region occupied by the benzyl moieties of L-TBOA, L-β-threo-BA and related analogs. It is plausible that the preference of L-β-threo-BA and L-TBOA for EAAT3 and EAAT2, respectively, could reside in the latter two pharmacophore regions.
|Advisor:||Esslinger, Christopher S.|
|Commitee:||Bridges, Richard J., Grimes, Mark L., Kavanaugh, Michael P., Lurie, Diana I.|
|School:||University of Montana|
|School Location:||United States -- Montana|
|Source:||DAI-B 69/12, Dissertation Abstracts International|
|Subjects:||Pharmacology, Organic chemistry|
|Keywords:||Benzyl aspartate, EAAT3, Glutamate, Transporters|
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