Association of AMPA receptors with a subset of glutamate receptor-interacting protein in vivo

The NMDA and AMPA classes of ionotropic glutamate receptors are concentrate d at postsynaptic sites in excitatory synapses. NMDA receptors interact via their NR2 subunits with PSD-95/SAP90 family proteins, whereas AMPA recepto rs bind via their GluR2/3 subunits to glutamate receptor-interacting protei n (GRIP), AMPA receptor-binding protein (ABP), and protein interacting with C kinase 1 (PICK1). We report here a novel cDNA (termed ABP-L/GRIP2) that is virtually identical to ABP except for additional GRIP-like sequences at the N-terminal and G-terminal ends. Like GRIP (which we now term GR(PI), AE P-L/GRIP2 contains a seventh PDZ domain at its C terminus. Using antibodies that recognize both these proteins, we examined the subcellular localizati on of GRIP1 and ABP-L/GRIP2 (collectively termed GRIP) and their biochemica l association with AMPA receptors. Immunogold electron microscopy revealed the presence of GRIP at excitatory synapses and also at nonsynaptic membran es and within intracellular compartments. The association of native GRIP an d AMPA receptors was confirmed biochemically by coimmunoprecipitation from rat brain extracts. A majority of detergent-extractable GluR2/3 was complex ed with GRIP in the brain. However, only approximately half of GRIP was ass ociated with AMPA receptors. Unexpectedly, immunocytochemistry of cultured hippocampal neurons and rat brain at the light microscopic level showed enr ichment of GRIP in GABAergic neurons and in GABAergic nerve terminals. Thus GRIP is associated with inhibitory as well as excitatory synapses. Collect ively, these findings support a role for GRIP in the synaptic anchoring of AMPA receptors but also suggest that GRIP has additional functions unrelate d to the binding of AMPA receptors.