Glutamate is an essential excitatory neurotransmitter regulating brain functions. Excitatory amino acid transporter (EAAT)-2 is one of the major glutamate transporters expressed predominantly in astroglial cells and is responsible for 90% of total glutamate uptake.
Glutamate is the predominant excitatory neurotransmitter in the central nervous system in mammalian, critical for normal brain functions including cognition, memory, learning, developmental plasticity, and long-term potentiation, mainly inactivated by the uptake of glutamate transporters. Five mammalian EAAT isoforms (EAAT1, EAAT2, EAAT3, EAAT4, EAAT5) have been characterized with each having different nomenclatures, expression patterns, and uptake kinetics. EAAT2 are principally expressed in astrocytes under normal conditions and localized to the cellular membrane, rarely in neurons. EAAT2 is the most plentiful EAAT and is primarily responsible for up to 80-90 % of total extracellular glutamate uptake activities in the brain, which plays a key role in regulating transmission and preventing excitotoxicity. EAAT2 expression is regulated at the level of transcription(including epigenetic modification), translation, trafficking, transport, and degradation. For regulation at the transcriptional level, endogenous and pharmacological modulators can induce activation or restraint, such as EGF, cAMP, PACAP TGF-β, TNF-α, ceftriaxone, and estrogen related compounds as well as co-culturing astrocytes with neurons. Once glutamate is uptaken by the astrocyte, it is converted into glutamine via glutamine synthetase. Glutamine is then transformed back into the presynaptic neuron and converted back into glutamate via glutaminase. The process is called glutamate-glutamine cycle. Glutamate taken up by EAAT2 can also be oxidized for energy by astrocytes. Research indicates that the function of EAAT2 can be impaired in the disease process of hypoxia, epilepsy and other energy exhaust conditions. EAAT2 may produce trans-shipment of glutamic acid, which leads to the accumulation of extracellular glutamate. The failure of timely removal of glutamate by astrocytes from the synapses results in a pathological process termed glutamate excitotoxicity. Excitotoxicity plays a dominant role in secondary damage following acute pathologies (such as traumatic brain injury, hyperexcitability, seizures and epilepsy, stroke, cerebral and retinal ischemia) and chronic pathologie (such as Amyotrophic lateral sclerosis, Alzheimer’s disease, Huntington’s disease, neuropathic pain and HIV-associated neurocognitive disorder). Quantities of studies have demonstrated that up-regulation of EAAT2 protein offers significant positive effects in many disease models, indicating that it could be a potential therapeutic strategy via EAAT2 activation.
There are pharmacological agents that directly modulate the transport function of EAAT2. Parawixin1 (purified from the spider Parawixia bistriata venom), directly enhances the glutamate uptake function of EAAT2. On the other hand, there are many glutamate transporter inhibitors. Inhibitors are categorized into two classes: competitive inhibitors and noncompetitive inhibitors. The competitive inhibitors include the cyclic molecules L-trans-2,4-pyrrolidinedicarboxylic acid (PDC; nonselective EAATs affinity), and Dihydrokainic acid (DHK; a selective inhibitor for EAAT2), L-threo-beta-benzyloxyaspartate(TBOA)and(3S)-3-[[3-[[4-(Trifluoromethyl)benzoyl]amino]phenyl]methoxy]-L-aspartic acid(TFB-TBOA). While TBOA is a nonspecific subtype inhibitor, TFB-TBOA has higher affinity for EAATs than TBOA. More recently, N-[4-(2-Bromo-4,5-difluorophenoxy) phenyl]-L-asparagine (WAY-213613) has been developed. WAY-213613 has higher potency and selectivity for EAAT2 over EAAT1 and EAAT3. Noncompetitive inhibitors include HIP-B (a EAATs inhibitor which binds at an allosteric binding site) and UCPH-101 (an EAAT1-selective inhibitor that targets a predominantly hydrophobic crevice in the trimerization domain of EAAT1). Blocking of EAAT2 with DHK, shows extended NMDA-receptor mediated excitatory post-synaptic current.
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