Behavioral and biochemical characterization of a mutant mouse strain lacking d-amino acid oxidase activity and its implications for schizophrenia
Introduction
N-methyl-d-aspartate receptors (NMDAR) are expressed at excitatory synapses throughout the central nervous system and mediate a wide range of brain processes, including synaptic plasticity associated with memory formation and learning (Bliss and Collingridge, 1993). NMDAR activation requires binding of the excitatory amino acid l-glutamate to its NR2 subunit and a co-agonist (d-serine or glycine) at the glycine site on the NR1 subunit. d-Serine is abundant in the forebrain, where its expression is correlated with that of NMDARs (Hashimoto and Oka, 1997, Matsui et al., 1995, Schell et al., 1997). It is apparent that in vivo the NMDA glycine site is not saturated in all brain regions (Fuchs et al., 2005), so it is possible that increasing glycine or d-serine levels could potentially modulate responses of the NMDAR.
The NMDAR hypofunction hypothesis of schizophrenia is principally based on the psycho-mimetic effects of the NMDAR antagonists phencyclidine (PCP) and ketamine. PCP induces a schizophrenia-like spectrum of symptoms in healthy subjects (positive, negative and cognitive aspects) and exacerbates symptoms in schizophrenia patients (Javitt and Zukin, 1991, Olney and Farber, 1995). Enhancing NMDAR function could therefore be of therapeutic benefit in schizophrenia. This has been tested clinically with glycine, d-serine and the glycine-site partial agonist d-cycloserine being administered as add-on therapy to conventional anti-psychotics. The results, although mixed, are encouraging with d-serine having benefit for the positive, negative and cognitive domains of schizophrenia (Heresco-Levy, 2005). There are issues with administration of d-serine or glycine, in particular with the large quantities of compound that need to be dosed to increase levels of these amino acids in the brain. Alternative approaches include inhibiting transporters of these amino acids to block cellular uptake, for example, by inhibiting the glycine Glyt1 transporter or the high affinity d-serine transporter Asc-1 (Sur and Kinney, 2004). Additionally, d-serine degradation can be prevented by inhibiting d-amino acid oxidase (DAO), which degrades a range of d-amino acids to their corresponding keto-acids (Konno and Yasumura, 1992).
A strain of mice has been identified lacking DAO activity (ddY/DAO−) due to a single point mutation (G181R) in DAO (Sasaki et al., 1992). ddY/DAO− mice have elevated levels of d-amino acids, including increased d-serine in serum and brain compared to wild-type mice (Hashimoto et al., 1993, Konno and Yasumura, 1983, Morikawa et al., 2001). Recently, they have also been shown to have enhanced hippocampal LTP and improved performance in a test of spatial learning (Maekawa et al., 2005). This suggests inhibiting DAO could have therapeutic impact on the cognitive deficits found in schizophrenics. ddY/DAO− mice provide an important tool in validating DAO inhibition as a therapeutic target for schizophrenia. Therefore, we have further characterized ddY/DAO− mice and shown for the first time that they have increased occupancy of the NMDAR glycine site resulting in increased NMDAR function in vivo. Analysis of the expression of related biochemical partners to DAO shows that there is no apparent compensation in response to a decrease in DAO activity. Combined with further novel behavioral data this provides encouraging support for DAO inhibition as a therapeutic option in treating schizophrenia.
Section snippets
Confirmation that the ddY/DAO− mouse strain lacks DAO activity
To confirm that the mouse strain ddY/DAO− lacks DAO activity, we examined DAO activity by two different methods. Histological studies with DAB staining in the kidney clearly show that there is no enzyme activity in ddY/DAO− mice (Fig. 1A; left-hand panel) compared to ddY/DAO+ (right-hand panel). This was confirmed independently by a tissue homogenate assay of the kidney of both strains. The same assay also showed no DAO activity in the cerebellum of ddY/DAO− mice (Fig. 1B) compared with the
Discussion
This current study establishes for the first time that mice with reduced d-amino acid oxidase activity (ddY/DAO−) have enhanced NMDAR function in vivo. NMDAR hypofunction is currently a leading hypothesis in explaining the etiology of schizophrenia (Olney and Farber, 1995, Heresco-Levy, 2005). Potentiating NMDAR activity through inhibiting DAO would therefore be predicted to have therapeutic utility in the clinical setting.
We have been able to obtain evidence of enhanced NMDAR function in the
Animals
ddY/DAO− and ddY/DAO+ mice were obtained from the laboratory which initially isolated this strain (Konno and Yasumura, 1983)). ddY/DAO− mice have been shown to have a missense mutation (Gly-181–Arg) in the DAO gene and so lack DAO activity (Sasaki et al., 1992).
All animals were group or singly housed (depending on fighting behavior) in solid-bottomed cages with sawdust bedding and environmental enrichment. Food and water were available ad libitum. Temperature and humidity were maintained at
References (55)
- et al.
Methodological variables during analysis of in vivo cerebellar cyclic GMP, an indirect marker of nitric oxide release
J. Pharmacol. Toxicol. Methods
(1994) - et al.
Pharmacological characterization of MDL 105,519, an NMDA receptor glycine site antagonist
Eur. J. Pharmacol.
(1997) - et al.
Synaptic plasticity: LTP and LTD
Curr. Opin. Neurobiol.
(1994) - et al.
Disrupted in Schizophrenia 1 and Nudel form a neurodevelopmentally regulated protein complex: implications for schizophrenia and other major neurological disorders
Mol. Cell. Neurosci.
(2004) - et al.
Memories of NMDA receptors and LTP
Trends Neurosci.
(1995) - et al.
Biological role of d-amino acid oxidase and d-aspartate oxidase. Effects of d-amino acids
J. Biol. Chem.
(1993) - et al.
Actions of d-cycloserine at the N-methyl-d-aspartate-associated glycine receptor site in vivo
Neuropharmacology
(1991) - et al.
d-amino acids in the central nervous system in health and disease
Mol. Genet. Metab.
(2005) - et al.
Inactivation of the glycine transporter 1 gene discloses vital role of glial glycine uptake in glycinergic inhibition
Neuron
(2003) - et al.
Free d-aspartate and d-serine in the mammalian brain and periphery
Prog. Neurobiol.
(1997)
Free d-serine, d-aspartate and d-alanine in central nervous system and serum in mutant mice lacking d-amino acid oxidase
Neurosci. Lett.
Mice lacking d-amino acid oxidase activity display marked attenuation of stereotypy and ataxia induced by MK-801
Brain Res.
d-Amino-acid oxidase is not present in the mouse liver
Biochim. Biophys. Acta
CGS 19755 is a potent and competitive antagonist at NMDA-type receptors
Eur. J. Pharmacol.
Association of DAAO with schizophrenia in the Chinese population
Neurosci. Lett.
Spatial learning and long-term potentiation of mutant mice lacking d-amino-acid oxidase
Neurosci. Res.
LTP and LTD: an embarrassment of riches
Neuron
Determination of free d-aspartic acid, d-serine and d-alanine in the brain of mutant mice lacking d-amino acid oxidase activity
J. Chromatogr., B, Biomed. Sci. Appl.
GLYX-13: a monoclonal antibody-derived peptide that acts as an N-methyl-d-aspartate receptor modulator
Neuropharmacology
NMDA antagonists as neurotherapeutic drugs, psychotogens, neurotoxins, and research tools for studying schizophrenia
Neuropsychopharmacology
A single-base-pair substitution abolishes d-amino-acid oxidase activity in the mouse
Biochim. Biophys. Acta
Cloning and expression of a glycine transporter reveal colocalization with NMDA receptors
Neuron
Exaggerated responses to chronic nociceptive stimuli and enhancement of N-methyl-d-aspartate receptor-mediated synaptic transmission in mutant mice lacking d-amino-acid oxidase
Neurosci. Lett.
Antagonists of excitatory amino acids and cyclic guanosine monophosphate in cerebellum
Neuropharmacology
Involvement of granule, basket and stellate neurons but not Purkinje or Golgi cells in cerebellar cGMP increases in vivo
Life Sci.
Lack of the alanine-serine-cysteine transporter 1 causes tremors, seizures, and early postnatal death in mice
Brain Res.
Spatial memory deficits induced by perinatal treatment of rats with PCP and reversal effect of d-serine
Neuropsychopharmacology
Cited by (94)
Coumarin derivatives as inhibitors of D-amino acid oxidase and monoamine oxidase
2022, Bioorganic ChemistryHuman D-amino acid oxidase: The inactive G183R variant
2018, Biochimica et Biophysica Acta - Proteins and ProteomicsExcitotoxicity
2018, Comprehensive Toxicology: Third EditionModeling Schizophrenia in Animals: Old Challenges and New Opportunities
2016, The Neurobiology of SchizophreniaMouse Models of Schizophrenia: Risk Genes
2016, Handbook of Behavioral Neuroscience
- 1
These authors contributed equally to this study.