EAATs for stroke: to modulate or not to modulate

Abstract ID 95456 Poster Board 324 Excitatory amino acid transporters (EAATs) are critical proteins in the CNS that regulate synaptic glutamate levels, crucially preventing excitotoxicity. The astrocytic transporter EAAT2 is responsible for the majority of glutamate clearance in the CNS. Aberrant EAAT2 activity and glutamatergic signaling occurs in many neuropsychiatric disorders. Our work focused on ischemic stroke, a condition that urgently needs new treatments. In ischemic stroke, excessive levels of released glutamate cause excitotoxicity, leading to secondary damage which ultimately results in cognitive deficits. We have developed novel allosteric modulators (AMs) of EAATs, including selective-EAAT2 positive allosteric modulators (PAMs), and non-specific and broad-acting analog AMs. We hypothesize the pharmacological activity of AMs is determined by differential interactions with critical amino acid residues located between the transporter's scaffold and transport domains. We have two main goals: to further understand the mechanism of these AMs, and to study their effects in an in vitro stroke model. Computational modeling predictions suggest some amino acid residues on EAAT2 that are critical to mediating the action of NA-014, an EAAT2-specific PAM. Dose-response assays evaluating the effect of NA-014 and other AMs offered further insights on which residues are important for their action, and what chemical moieties confer EAAT subtype selectivity and pharmacological action. Additionally, we evaluated potential translatability of EAATs PAMs in a model of ischemic stroke. We hypothesize that these compounds can restore glutamatergic homeostasis by augmenting glutamate clearance. We evaluated NA-014 in an in vitro model of ischemic stroke, oxygen glucose deprivation, in primary neuron-glia cultures, and found that it demonstrated neuroprotective properties. Collectively, these studies expand our mechanistic understanding of the EAAT AMs and demonstrate their clinical utility for ischemic stroke.