REGIONAL AND ONTOGENIC EXPRESSION OF THE NMDA RECEPTOR SUBUNIT NR2D PROTEIN IN RAT-BRAIN USING A SUBUNIT-SPECIFIC ANTIBODY

A polyclonal antibody for the NMDA receptor subunit NR2D has been deve loped that identifies an similar to 160-kDa band on immunoblots from N R2D transfected cells and CNS tissues. No cross-reactivity is seen wit h other NMDA receptor subunits. The NR2D receptor subunit is N-glycosy lated in both brain and transfected cells. Transfected cells expressin g NR2D are immunofluorescently labeled, whereas untransfected cells or cells transfected with other NMDA receptor subunit cDNAs are not. Sim ilarly, the NR2D subunit is selectively and quantitatively immunopreci pitated, whereas the NR1, NR2A, or NR2B subunit is not. The relative d ensities of the NR2D subunit in nine areas of postnatal day 7 and adul t rat brains have been determined by quantitative immunoblotting. NR2D was expressed at highest levels in the thalamus, midbrain, medulla, a nd spinal cord, whereas intermediate levels of this subunit were found in the cortex and hippocampus. Low or undetectable levels were seen i n the olfactory bulb, striatum, and cerebellum. Following a peak after the first week of birth, NR2D protein levels decreased by about twofo ld in adulthood in all rat brain regions examined. More complete ontog enic profiles were determined for the diencephalon, telencephalon, and spinal cord where similar ontogenic patterns were seen. NR2D protein is present at high levels at embryonic stages of development, rises to a peak at postnatal day 7, and decreases but remains measurable durin g late postnatal life. This study demonstrates the generation and char acterization of an antibody selective for the NR2D NMDA receptor subun it as well as a determination of the distribution and ontogenic profil e of this subunit in rat brain. The results suggest that native NMDA r eceptors containing the NR2D subunit may have functional roles not onl y in the young brain but also in adult brain.