Glutamate receptor subunit 2-selective antibody shows a differential distribution of calcium-impermeable AMPA receptors among populations of neurons

Abstract

α‐Amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionate (AMPA) receptors are the major excitatory neurotransmitter receptors of the central nervous system. AMPA receptor complexes that contain the AMPA‐type glutamate receptor subunit 2 (GluR2) are responsible for the low calcium permeability typical of most AMPA receptors, and the absence of GluR2 may be a key factor in neurotoxicity. A polyclonal antibody was produced to a 16 amino acid peptide near the C‐terminus of GluR2 and was affinity‐purified in a three‐step procedure. The antibody did not recognize other AMPA subunits in transfected cells with the use of either Western blots or immunocytochemistry. This highly specific GluR2 antibody was used to provide a specific morphological study of GluR2 protein distribution in neurons and synapses of the rat. GluR2 is prevalent in most principal neurons throughout the telencephalon. Neurons with few or no GluR2 subunits include two major types: 1) some populations of interneurons of the telencephalon and of some other areas and 2) many populations of principal neurons in the brainstem and spinal cord. Immunofluorescence showed that GluR2 immunolabeling was widespread, including in dendrites and puncta, in the hippocampus and neocortex. Where they were present, GluR2 subunits colocalized with other AMPA receptor subunits in individual neurons. Electron microscopy of the hippocampus showed GluR2‐bearing, calcium‐impermeable AMPA receptors postsynaptic to dendrite synapses of forebrain principal neurons. In addition, electron microscopy of the neocortex showed significant staining in postsynaptic profiles. Electron microscopy of the cerebellum revealed the presence of GluR2 subunits in the postsynaptic profiles of many parallel fiber/Purkinje cell spine synapses, whereas electron microscopy of the spinal cord showed substantial staining in the postsynaptic profiles of dorsal horn synapses, but not in ventral horn synapses. Both ultrastructural and immunofluorescence data showed that calcium‐impermeable AMPA receptors are widespread in dendrite arborizations. J. Comp. Neurol. 385:456–476, 1997.