Macroscopic and microscopic properties of a cloned glutamate transporter/chloride channel

The behavior of a Cl2 channel associated with a glutamate transporter was studied using intracellular and patch recording techniques in Xenopus oocytes injected with human EAAT1 cRNA. Channels could be activated by application of glutamate to either face of excised membrane patches. The channel exhibited strong selectivity for amphipathic anions and had a minimum pore diameter of;5Å. Glutamate flux exhibited a much greater temperature dependence than Cl2 flux. Station-ary and nonstationary noise analysis was consistent with a sub-femtosiemen Cl2 conductance and a maximum channel Po,, 1. The glutamate binding rate was similar to estimates for receptor binding. After glutamate binding, channels activated rapidly followed by a relaxation phase. Differences in the mac-roscopic kinetics of channels activated by concentration jumps of L-glutamate or D-aspartate were correlated with differences in uptake kinetics, indicating a close correspondence of channel gating to state transitions in the transporter cycle. Key words: glutamate transporter; uptake; kinetics; astrocyte; postsynaptic; chloride channel Glutamate is the primary excitatory neurotransmitter at central synapses, and its effects on receptors are terminated by diffusion and by the actions of glutamate transporters. These molecules are members of a large amino acid transporter gene family (Malandro and Kilberg, 1996), and they exhibit discrete anatomical localiza-tions. GLAST (EAAT1) and GLT-1 (EAAT2) are found primar-ily in glial cells, whereas EAAC1 (EAAT3), EAAT4, and EAAT5 are primarily expressed in neuronal cells (Rothstein et al., 1994; Lehre et al., 1995; Yamada et al., 1996; Eliasof et al., 1998). Glutamate transport is electrogenic and coupled to sodium and proton influx and potassium efflux (Kanner and Sharon, 1978