MOLECULAR DISTINCTION OF 3 N-METHYL-D-ASPARTATE-RECEPTOR SUBTYPES IN-SITU AND DEVELOPMENTAL RECEPTOR MATURATION DEMONSTRATED WITH THE PHOTOAFFINITY LIGAND I-125 LABELED CGP-55802A

Activation of N-methyl-D-aspartate (NMDA) receptors is essential for s ynaptic plasticity in the central nervous system and contributes to ne uronal death under various pathological conditions. Although several s ubunits have been cloned, the structure of NMDA receptors in situ is u nresolved. By using a photoreactive antagonist with nanomolar affinity to the NMDA-binding site, three types of receptors were differentiate d by their pattern of photoaffinity-labeled subunits. In adult brain, a protein of 175-kDa was photoreactive that displayed a profile of lig and binding and autoradiographical distribution corresponding to NMDA receptors. In contrast, in early postnatal brain, proteins of both 175 kDa and 115 kDa were photolabeled. This labeling pattern is switched to that of adult brain around postnatal day 10, pointing to a structur al maturation of NMDA receptors. A third type of receptor could be ide ntified in cerebellar granule cell cultures, where NMDA receptors medi ate trophic effects and photolabeling was exclusively targeted to a 11 5-kDa protein. To identify the proteins labeled in situ, recombinant r eceptors were subjected to photolabeling. When the NR1 subunit was coe xpressed with either the NR2A, NR2B, or NR2C subunit, only the combina tion of NR1/NR2A was photoreactive. Both the NR1 and NR2A subunits wer e photolabeled, corresponding in size to the proteins labeled in situ. However, the lack of subunit-selectivity in photolabeling the NR1/NR2 A combination suggests the presence of additional receptor components in situ to explain the subunit-selective photoreactivity in adult brai n (175 kDa) and in cerebellar granule cells (115 kDa). The subunit com bination NR1/NR2A by itself appears insufficient to describe a major p opulation of NMDA receptors, in particular, in adult brain.