D-serine is an endogenous and obligatory coagonist for the glycine site of the N-methyl-D-aspartate receptor in the mammalian brain. D-serine is synthesized from L-serine by serine racemase; immunohistochemical studies have long been believed to indicate that serine racemase and D-serine occur predominantly in astrocytes. However, we have recently demonstrated in the primary cultures that both the mRNA and protein levels of serine racemase are higher in neurons than in astrocytes. Here we report the application of in situ hybridization based on tyramide signal amplification for the detection of serine racemase mRNA in sections of the adult rat brain. Serine racemase mRNA could be demonstrated in a large number of neurons throughout the brain, especially in the forebrain such as the cerebral cortex, striatum, and hippocampus. This is the first study to demonstrate the exact localization of serine racemase mRNA at the cellular or tissue level. These results suggest that neuron-derived D-serine could modulate neurotransmission via the glycine site of N-methyl-D-aspartate receptors.

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Abstract. Recent studies indicate that an endogenous co-agonist for an N-methyl-D-aspartate (NMDA) receptor-related glycine site, D-serine, is synthesized by serine racemase and is metabolized by D-amino acid oxidase (DAO) and that acute treatment with morphine augments the gene expression of serine racemase and DAO in rat brain. To further elucidate the mechanism underlying the activation of NMDA receptors following chronic opioid administration, we have evaluated the effects of the chronic administration of morphine on the mRNA and protein expressions of serine racemase and DAO and on the contents of D-serine in several areas of the rat brain. Repeated administration of morphine for 30 days produced a significant augmentation of both the mRNA and protein expressions of serine racemase in all the brain regions, whereas no significant change in the protein expression of DAO was observed in all the brain regions. Furthermore, the chronic administration caused a slight but significant elevation in the concentration of D-serine in the cortex, striatum, and hippocampus. These results indicate the elevated D-serine level following the chronic morphine treatment could at least in part be involved in the activation of NMDA receptors via the glycine site.

IntroductionD-Serine acts as an obligatory and endogenous coagonist for the N-methyl-D-aspartate (NMDA) receptorrelated glycine site in the mammalian brain (Hashimoto et al., 1993;Matsui et al., 1995;Schell et al., 1995;Hashimoto and Oka, 1997;Danysz and Parsons, 1998). D-Serine is synthesized by serine racemase (Srr), which converts L-serine to D-serine. The regional profile of Srr closely resembles those of endogenous D-serine and NMDA receptors, which are highest in the forebrain and lowest in the hindbrain (Yoshikawa et al., 2004). Previous immunohistochemical studies have demonstrated that D-serine and Srr were localized exclusively in glial fibrillary acidic protein (GFAP)-positive astrocytes (Hashimoto and Oka, 1997;Schell et al., 1997; Wolosker et al., 1999;Mustafa et al., 2004). However, recent studies using immunohistochemistry, Western blot, and in situ hybridization have demonstrated that D-serine (Yasuda et al., 2001;Puyal et al., 2006) and Srr (Kartvelishvily et al., 2006;Yoshikawa et al., 2006Yoshikawa et al., , 2007 are signifi cantly expressed in neurons of tissue sections, and of primary cultures of the rat brain as compared with astrocytes.Compared with the brain, the retina contains fewer cell types, many of which have recognizable morphologies and consistent positions, making any investigation of the mechanisms for the central nervous system more The serine racemase mRNA is expressed in both neurons and glial cells of the rat retina