Immunobiochemical characterization of the NMDA-receptor subunit NR1 in the developing and adult rat brain

Abstract

To investigate the developmental and regional expression of the NR1-subunit of the NMDA-receptor on the protein level, two polyclonal antisera [NR1(N) and NR1(C)] were raised against fusion proteins derived from the N- and C-terminal domain of the NR1-subunit, respectively. In Western blots of rat brain membranes, both antisera specifically recognized a single protein band with an apparent molecular size of 115 kDa. The regional distribution of the NR1-subunit immunoreactivity was analyzed in the developing and adult rat brain using sections blotted onto nitrocellulose membranes for immunostaining. With the NR1(N)-antiserum, strongest signals were detected in hippocampus, followed by cortex, striatum and thalamus, and weaker staining was observed in tectum, brainstem and cerebellum of adult brain. The NR1(C)-immunoreactivity exhibited a similar distribution, except that the staining in thalamus, tectum, brainstem and cerebellum was faint or virtually absent. The distinct pattern of NR1(N)- and NR1(C)-immunoreactivity arose during postnatal development. At birth, moderate staining with both NR1-subunit antisera was observed throughout the brain increasing strongly in most brain regions until postnatal day 21. In some brain areas, however, the NR1(C)-, in contrast to the NR1(N)-staining, decreased postnatally e.g. in thalamus, tectum and brainstem. The restricted staining intensity of the NR1(C)-antiserum in particular areas of adult and developing brain appears to reflect the emergence of C-terminal splice variants of the NR1-subunit which are not recognized by the NR1(C)-antiserum.