On the basis of their relative Glu

On the basis of their relative Glu transport rates and anion currents, EAATs group into two functionally distinct classes, EAAT1–3 being efficient Glu transporters with small associated macroscopic anion currents and EAAT4–5 low-capacity transporters with predominant anion conductance [18]. The differences are due to largely differing Glu transport rates, as the conduction properties of separate EAAT anion channels are very similar [15]. So far, neither the localization of this conduction pathway, the underlying conformation of the transporter, nor the mechanisms of anion permeation are sufficiently understood. EAAT anion conduction appears to play a role in synaptic transmission in neurons [19] and may contribute to the regulation of intracellular chloride concentrations in glial cells [] (see below).
Novel generations of EAAT modulators Considering the immense therapeutic potential in pharmacological intervention into glutamatergic neurotransmission and the fact that other neurotransmitter transporters are targeted by clinically administered drugs against epilepsy, depression and attention deficit hyperactivity disorder [21], EAATs have historically received surprisingly little attention as putative drug targets [2]. EAAT modulation as a therapeutic concept faces the same inherent obstacle as drugs acting through other glutamatergic mechanisms: the considerable risk of inducing adverse effects due to the abundance of glutamatergic neurons in the CNS and the importance of the neurotransmitter for most central processes. Furthermore, most EAAT ligands published to date have been derived from the use of Glu, aspartate or other α-amino acids as lead structures. These efforts have admittedly spawned some important ligands for the transporters, and seemingly simple hiv protease inhibitors can exhibit surprisingly complex pharmacology, as recently demonstrated for the bicyclic Glu analog (+)-HIP-B [22]. Nevertheless, this strategy has some major shortcomings when it comes to developing pharmacological tools. The conserved nature of the substrate binding sites in the five EAATs has, with the exception of a few EAAT2-selective inhibitors, so far been an insurmountable obstacle to the development of truly subtype-selective ligands [2]. Moreover, since both substrates and inhibitors acting through this site will inhibit EAAT-mediated Glu uptake, the therapeutic potential in augmentation of EAAT function cannot be addressed with orthosteric ligands [2]. These realisations have fuelled two alternative approaches for ligand development. Allosteric modulators. Several endogenous nutrients and exogenous compounds have been found to modulate EAATs, however, the majority of these exhibit rather promiscuous pharmacological profiles and/or low potencies at the transporters (Figure 2a) [23, 24, 25, 26, 27]. In recent years we have developed a series of potent EAAT1/GLAST-inhibitors, including UCPH-101 and UCPH-102, from a hit identified in a screening of a small compound library (Figure 2a) [28, 29]. The compounds are highly selective for EAAT1 over the other four EAAT subtypes, and they act as negative allosteric modulators (NAMs) inhibiting Glu uptake through EAAT1 non-competitively through an intra-protomeric site involving regions from both the transport and trimerization domains (Figure 2a) []. Interestingly, close structural analogs from this series exhibit profoundly different kinetic properties as EAAT1 inhibitors, with UCPH-101 and UCPH-102 exhibiting slow and fast unbinding kinetics from the transporter, respectively []. The distinct mechanisms of action and binding sites proposed for a couple of allosteric EAAT modulators [24, 27, 30•] combined with the dramatic ‘reshuffling’ of transporter regions during the transport cycle [3, 8] strongly suggest that the EAAT comprises numerous allosteric sites (Figure 2a). In addition to the higher chance of obtaining subtype-selectivity and the ability to augment EAAT function offered by allosteric ligands, it seems possible to exert very different kinds of transporter inhibition through these sites, as exemplified by the partial inhibition mediated by zinc [23, 24] and the different inhibition kinetics exhibited various NAMs at EAATs [25, 30•]. This ability to tweak the degree or the duration of EAAT modulation could be an important quality in terms of walking the fine line between therapeutic efficacy and adverse effects when targeting the glutamatergic system. Hence, we propose that allosteric modulators hold great potential as pharmacological tools and therapeutics in the EAAT field, and hopefully the relatively simple strategy behind the discovery of UCPH-101 and its analogs will serve as inspiration for others.