Quantitation of NMDA receptor (NMDAR1) mRNA levels in the adult and developing rat CNS

A rapid and sensitive solution hybridization assay was used to quantitate N- methyl- d- aspartate (NMDA) receptor mRNA levels in the central nervous system (CNS) of rat, mouse and human. A riboprobe labelled with 32P was prepared from a plasmid containing a 1413 base sequence from the cDNA for the functional rat NMDA receptor subunit, NMDAR1. Using a full length sense transcript as the calibration standard, the assay reliably measures 8 pg of NMDAR1 mRNA. When expressed as pg of NMDAR1 mRNA/μg total cellular RNA, the highest levels in the adult rat CNS are in the olfactory bulb (20.9 pg/υg RNA) and the lowest levels are in the spinal cord (5.2 pg/υg RNA). Intermediate levels were found in frontal cortex, hippocampus, cerebellum and whole brain. In the mouse CNS the highest levels of NMDAR1 mRNA were found in the olfactory bulb (12.9 pg equivalents/υg RNA), followed closely by hippocampus, frontal cortex and cerebellum. Mouse spinal cord (4.4 pg equivalents/υg RNA) had the lowest levels of NMDAR1 mRNA. The NMDAR1 riboprobe hybridizes with the same size transcripts in Poly(A) + RNA from rat, mouse and human brain. In the developing rat, NMDAR1 mRNA levels in frontal cortex and hippocampus increased nearly 3 fold from postnatal day 3 to day 15 and approximately doubled from day 15 to day 67 (adult). Therefore, from postnatal day 3 to adult (day 67) frontal cortex and hippocampus levels of NMDAR1 mRNA increased nearly 6 fold. In contrast, cerebellum and brain stem showed no change in NMDAR1 mRNA levels when postnatal days 3 and 15 were compared while the levels were increased 2 fold when day 15 and day 67 were compared. These changes were not a function of alterations in total cellular RNA. These results demonstrate significant differences in frontal cortex and hippocampus when compared to cerebellum and brain stem in the postnatal development of NMDAR1 mRNA.