The substituted aspartate analogue L-beta-threo-benzyl-aspartate preferentially inhibits the neuronal excitatory amino acid transporter EAAT3.
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 beta-substituted aspartate analogues identify L-beta-benzyl-aspartate (L-beta-BA) as among the first blockers that potently and preferentially inhibits the neuronal EAAT3 subtype. Kinetic analysis of D-[(3)H]aspartate uptake into C17.2 cells expressing the hEAATs demonstrate that L-beta-threo-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-beta-BA as a non-substrate inhibitor. Analyzing L-beta-threo-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 L-beta-threo-BA occupied by the benzyl moieties of L-TBOA, L-beta-threo-BA and related analogues. It is plausible that the preference of L-beta-threo-BA and L-TBOA for EAAT3 and EAAT2, respectively, could reside in the latter two pharmacophore regions.
Esslinger
C Sean
CS
COBRE Center for Structural and Functional Neuroscience, Departments of Biomedical & Pharmaceutical Sciences, University of Montana, Missoula, MT 59812, USA.
Agarwal
Shailesh
S
Gerdes
John
J
Wilson
Paul A
PA
Davis
Erin S
ES
Awes
Alicia N
AN
O'Brien
Erin
E
Mavencamp
Teri
T
Koch
Hans P
HP
Poulsen
David J
DJ
Rhoderick
Joseph F
JF
Chamberlin
A Richard
AR
Kavanaugh
Michael P
MP
Bridges
Richard J
RJ
D027322:Amino Acid Transport System X-AG / Q000037:antagonists & inhibitors*; D000818:Animals; D001224:Aspartic Acid / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology*; D002461:Cell Line, Transformed; D004305:Dose-Response Relationship, Drug; D004558:Electric Stimulation / Q000379:methods; D027341:Excitatory Amino Acid Transporter 1 / Q000502:physiology; D027342:Excitatory Amino Acid Transporter 2 / Q000502:physiology; D050580:Excitatory Amino Acid Transporter 3 / Q000037:antagonists & inhibitors* / Q000502:physiology; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D051379:Mice; D008958:Models, Molecular; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D009865:Oocytes; D018408:Patch-Clamp Techniques / Q000379:methods; D014162:Transfection / Q000379:methods; D014316:Tritium / Q000493:pharmacokinetics; D014981:Xenopus
D027322:Amino Acid Transport System X-AG; D027341:Excitatory Amino Acid Transporter 1; D027342:Excitatory Amino Acid Transporter 2; D050580:Excitatory Amino Acid Transporter 3; C508335:L-beta-threo-benzyl-aspartate; C493399:Slc1a1 protein, mouse; C120673:benzyloxyaspartate; D014316:Tritium; D001224:Aspartic Acid