Noradrenergic innervation of major brain areas, such as the hypothalamus and cerebral cortex, originates in part from the locus coeruleus (LC) located in the hindbrain. The LC is theoretically under the control of excitatory glutamatergic afferents, since this area can be stimulated by exogenous glutamate. Therefore, this study examined the localization of ionotropic glutamate receptors in the LC of perfused fixed rhesus monkey brains. Both the NMDA and AMPA classes of receptors were examined by immunocytochemistry (ICC) and in situ hybridization (ISH). The LC was localized initially in coronal sections by ICC for tyrosine hydroxylase (Boehringer-Mannheim), the rate limiting enzyme in the catecholamine synthesis pathway. Neighboring sections were then immunostained for NR1, the NMDA receptor subunit found in all NMDA receptor complexes, and for GluR1 and GluR2/3, major subunits of AMPA receptors (Chemicon Int.). In all cases, immunostaining for NR1, GluR1 and GluR2/3 subunits were found in neurons of the LC. To corroborate these results, ISH for NR1, GluR1, GluR2 and GluR3 subunits were performed on sections containing the LC. Coronal sections were initially postfixed, digested with proteinase K, acetylated, then hybridized overnight at 60 C with 35S-labelled cRNA probes generated from rat cDNA templates. Sections were subsequently washed, treated with RNase A, then further rinsed, with a final stringency wash at 70 C in 0.1 X SSC. The LC was visualized on exposed -max films (Amersham) and emulsion-dipped slides (Kodak). Again, all subunits examined were found in the LC, with NR1 and GluR1 giving the most robust signals. Both GluR2 and GluR3 mRNA were also found in LC neurons, corroborating the immunostaining seen with the GluR2/3 antibody. (Findings from this study were presented in 1996 at the Annual Meeting of the Society for Neuroscience, held in Washington D.C.).
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