D‐Serine metabolism in C6 glioma cells: Involvement of alanine‐serine‐cysteine transporter (ASCT2) and serine racemase (SRR) but not D‐amino acid oxidase (DAO)

Pilleriin Sikka,Rosie Walker,Rebecca Cockayne,Matthew J.A Wood,Philip W.J Burnet,Paul J Harrison

doi:10.1002/jnr.22332

Pilleriin Sikka, Rosie Walker + Show 4 more

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https://doi.org/10.1002/jnr.22332

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Abstract

D-serine is an endogenous N-methyl-D-aspartate (NMDA) receptor coagonist. It is synthesized from L-serine by serine racemase (SRR), but many aspects of its metabolism remain unclear, especially in the forebrain, which lacks active D-amino acid oxidase (DAO), the major D-serine degradative enzyme. Candidate mechanisms include SRR operating in α,β-eliminase mode (converting D-serine to pyruvate) and regulation by serine transport, in which the alanine-serine-cysteine transporter ASCT2 is implicated. Here we report studies in C6 glioma cells, which “simulate” the forebrain, in that the cells express SRR and ASCT2 but lack DAO activity. We measured D-serine, ASCT2, SRR, and DAO expression and DAO activity in two situations: after incubation of cells for 48 hr with serine isomers and after increased or decreased SRR expression by transfection and RNA interference, respectively. Incubation with serine enantiomers decreased [3H]D-serine uptake and ASCT2 mRNA and increased SRR immunoreactivity but did not alter DAO immunoreactivity, and DAO activity remained undetectable. SRR overexpression increased D-serine and pyruvate and decreased [3H]D-serine uptake and ASCT2 mRNA but did not affect DAO. SRR knockdown did not alter any of the parameters. Our data suggest that D-serine transport mediated by ASCT2 contributes prominently to D-serine homeostasis when DAO activity is absent. The factors regulating D-serine are important for understanding normal NMDA receptor function and because D-serine, along with DAO and SRR, is implicated in the pathogenesis and treatment of schizophrenia. © 2010 Wiley-Liss, Inc.

Highlights

The neutral amino acid D-serine acts on the glycine binding site of the N-methyl-D-aspartate receptor (NMDAR) and modulates glutamate-mediated receptor activation
D-serine is synthesized from L-serine by serine racemase (SRR), and it is degraded by D-amino acid oxidase (DAO)
The current study sought to explore the roles of SRR, DAO, and ASCT2 in the modulation of serine isomers in C6 glial cells, which contain undetectable DAO activity

Summary

Introduction

The neutral amino acid D-serine acts on the glycine binding site of the N-methyl-D-aspartate receptor (NMDAR) and modulates glutamate-mediated receptor activation. D-serine is synthesized from L-serine by serine racemase (SRR), and it is degraded by D-amino acid oxidase (DAO) In the brain, both SRR and DAO are primarily but not exclusively localized to glial cells (Wolosker et al, 1999; Mustafa et al, 2004; Kartvelishvily et al, 2006; Miya et al, 2008). SRR may operate in reverse racemase mode, converting D- to L-serine (Foltyn et al, 2005) This multifunctional capability of SRR could be sufficient to regulate D-serine in the forebrain and preclude the need for active DAO. The rat C6 glioma cells lack functional DAO despite expressing it (Park et al, 2006; Burnet et al, 2009) These cells are useful for studying D-serine in a cellular environment that lacks DAO, because in this respect they ‘‘simulate’’ the situation in the forebrain.

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