D-amino acid oxidase (DAO) is a flavin-dependent enzyme that is expressed in the mammalian brain. It is the metabolising enzyme of several D-amino acids, including D serine, which is an endogenous agonist at the glycine co-agonist site of the glutamatergic NMDA receptor. As such, regulation of D serine levels in the brain by DAO may indirectly modulate the activity of NMDA receptors. The expression and activity of DAO have been reported to be increased in schizophrenia. It has been identified as a putative susceptibility gene for the disorder, and as a potential therapeutic target. This thesis explored three aspects of the interface between DAO and the DA system. First, the expression of DA was investigated in the ventral tegmental area (VTA), the source of the dopaminergic mesocortical pathway. Traditionally, DAO was considered to be an enzyme confined to the hindbrain and to glia, but more recent studies have reported its expression in additional brain regions, and also in neurons. DAO mRNA and protein was found to be expressed in the VTA, and was present in both neurons and glia in this region, whereas in the cerebellum, DAO expression appeared solely glial. DA output from the VTA is regulated by NMDA receptors, and hence expression of DAO in the VTA suggests that it may serve a role in modulating cortical DA via regulation of D serine levels and NMDA receptor function. The second part of this thesis investigated the effects of DAO inhibition and D serine administration on DA levels in the prefrontal cortex (PFC) using in vivo microdialysis. Systemic DAO inhibition and D serine administration resulted in increases in extracellular levels of DA metabolites in the PFC, despite no detectable change in DA. Similarly, DA metabolites in the PFC increased after local application of D serine to the VTA, but no change was detected in DA. However, local DAO inhibition in the VTA resulted in increased levels of both DA and its metabolites, and DAO inhibition combined with D serine administration also produced increases in DA. This suggested that DAO and its regulation of D-serine levels may serve to indirectly modulate mesocortical DA function, and this may be mediated via the VTA. This notion was supported in the final section of this thesis, in which the expression of three DA genes was measured in the PFC of a novel line of DAO knockout mice. In this pilot study, there was evidence for an increase in Comt and Drd2 mRNAs in the knockout mice. As such, constitutive abolition of DAO activity may also alter mesocortical DA function. These studies provide new insights into the presence and role of DAO beyond the hindbrain, and point to a potentially important physiological function in modulating the activity of the mesocortical DA system via the VTA. This could be therapeutically relevant in the context of elevating cortical DA in the treatment of schizophrenia, and may provide supporting evidence for the clinical use of DAO inhibitors.

Recent large-scale genome-wide studies for many human disorders have identified associations with numerous genetic variants. The biological interpretation of these variants presents a major challenge. In particular, the identification of biological pathways underlying the association could provide crucial insights into the disease aetiologies. In this thesis, I used functional genomics approaches to increase our understanding of neuropsychiatric and neurodevelopmental disorders. Firstly, in an integrative analysis of autism spectrum disorder (ASD), I looked into the role of genes targeted by Fragile-X Mental Retardation Protein ("FMRP targets"). I found evidence that FMRP targets contribute to ASD via two distinct aetiologies: (1) ultra-rare and highly penetrant single disruptions of embryonically upregulated FMRP targets ("single-hit aetiology") or (2) the combination of multiple less penetrant disruptions of synaptic FMRP targets ("multiple-hit aetiology"). In particular, I developed a pathway-association test sensitive to multiple-hit aetiologies. Secondly, I carried out an integrative analysis of bipolar disorder, following up a previously identified association with long-term potentiation. The association was not consistent across independent SNP and CNV datasets. Thirdly, I addressed the difficulty in identifying functional relationships between genes by integrating different datasets into a gene functional-linkage network tuned to the nervous system ("NsNet"). NsNet identified functional links between the genes disrupted by de novo loss-of-function mutations in ASD and, separately, in schizophrenia probands more sensitively than a general functional-linkage network. Fourthly, I considered the challenge of interpreting the phenotypic impact of gene disruptions, focusing on the identification of haploinsufficient genes. I constructed a gene haploinsufficiency score based on genome-wide datasets. Compared to existing approaches, the new score performed better in identifying less-studied haploinsufficient genes. This work both extends the methodology to detect the contribution of genetic variation to neuropsychiatric disorders and also yields insights into the variant genes and the pathways that underlie them. Firstly, in an integrative analysis of autism spectrum disorder (ASD), I looked into the role of genes targeted by Fragile-X Mental Retardation Protein ("FMRP targets"). I found evidence that FMRP targets contribute to ASD via two distinct aetiologies: (1) ultra-rare and highly penetrant single disruptions of embryonically upregulated FMRP targets ("single-hit aetiology") or (2) the combination of multiple less penetrant disruptions of synaptic FMRP targets ("multiple-hit aetiology"). In particular, I developed a pathway-association test sensitive to multiple-hit aetiologies. Secondly, I carried out an integrative analysis of bipolar disorder, following up a previously identified association with long-term potentiation. The association was not consistent across independent SNP and CNV datasets. Thirdly, I addressed the difficulty in identifying functional relationships between genes by integrating different datasets into a gene functional-linkage network tuned to the nervous system ("NsNet"). NsNet identified functional links between the genes disrupted by de novo loss-of-function mutations in ASD and, separately, in schizophrenia probands more sensitively than a general functional-linkage network. Fourthly, I considered the challenge of interpreting the phenotypic impact of gene disruptions, focusing on the identification of haploinsufficient genes. I constructed a gene haploinsufficiency score based on genome-wide datasets. Compared to existing approaches, the new score performed better in identifying less-studied haploinsufficient genes. This work both extends the methodology to detect the contribution of genetic variation to neuropsychiatric disorders and also yields insights into the variant genes and the pathways that underlie them.