EAAC1, A HIGH-AFFINITY GLUTAMATE TRANSPORTER, IS LOCALIZED TO ASTROCYTES AND GABAERGIC NEURONS BESIDES PYRAMIDAL CELLS IN THE RAT CEREBRAL-CORTEX

High-affinity uptake of glutamate from the synaptic cleft plays a cruc ial role in regulating neuronal activity in physiological and patholog ical conditions. We have used affinity-purified specific polyclonal an tibodies raised against a synthetic peptide corresponding to the C-ter minal region of rabbit and rat EAAC1. a glutamate (Glu) transporter be lieved to be exclusively neuronal, to investigate its cellular and sub cellular localization and whether it is expressed exclusively in gluta matergic cells of infragranular layers, as suggested by previous studi es. Light microscopic studies revealed that EAAC1 immunoreactivity (ir ) is localized to neurons and punctate elements in the neuropil. EAAC1 -positive neurons were more numerous in layers II-III and V-VI, i.e. t hroughout all projection layers. Most EAAC1-positive neurons were pyra midal, although nonpyramidal cells were also observed. Some EAAC1-posi tive non-pyramidal neurons stained positively with an antiserum to GAD , thus demonstrating that EAAC1 is not confined to glutamatergic neuro ns. Non-neuronal EAAC1-positive cells were also observed in the white matter, and some of them stained positively with an antiserum to GFAP. Ultrastructural studies showed that EAAC1-ir was in neuronal cell bod ies, dendrites and dendritic spines. but not in axon terminals, i.e. e xclusively postsynaptic. Analysis of the type of axon terminals synaps ing on EAAC1-ir profiles showed that 97% of them formed asymmetric con tacts, thus indicating that EAAC1 is located at the very sites of exci tatory amino acid release. Unexpectedly, EAAC1-ir was also found in a few astrocytic processes located in both the gray and the white matter . The localization of EAAC1 may explain the pathological symptoms that follow EAAC knockout (seizures and mild toxicity), as seizures could be due to the loss of EAAC1-mediated fine regulation of neuronal excit ability at axodendritic and axospinous synapses, whereas the mild toxi city may be related to the functional inactivation of astrocytic EAAC1 .