The substrate specificity of a neuronal glutamate transporter is determined by the nature of the coupling ion

Glutamate transporters are essential for terminating synaptic transmission. Glutamate is translocated together with three sodium ions. In the neuronal glutamate transporter EAAC1, lithium can replace sodium. To address the question of whether the coupling ion interacts with the 'driven' substrate during co-transport, the kinetic parameters of transport of the three substrates, l-glutamate and d- and l-aspartate by EAAC-1 in sodium- and lithium-containing media were compared. The major effect of the substitution of sodium by lithium was on K_m. In the presence of sodium, the values for K_m and I_max of these substrates were similar. In the presence of lithium, the K_m for l-aspartate was increased around 13-fold. Remarkably, the corresponding increase for l-glutamate and d-aspartate was much larger, around 130-fold. In marked contrast, the K_i values for a non-transportable substrate analogue were similar in the presence of either sodium or lithium. The preference for l-aspartate in the presence of lithium was also observed when electrogenic transport of radioactive substrates was monitored in EAAC1-containing proteoliposomes. Our results indicate that, subsequent to substrate binding, the co-transported solutes interact functionally in the binding pocket of the transporter.