Characterisation of the pathogenic effects of the in vivo expression of an ALS-linked mutation in D-amino acid oxidase: Phenotype and loss of spinal cord motor neurons

Amyotrophic lateral sclerosis (ALS) is the most common adult-onset neuromuscular disorder characterised by selective loss of motor neurons leading to fatal paralysis. Current therapeutic approaches are limited in their effectiveness. Substantial advances in understanding ALS disease mechanisms has come from the identification of pathogenic mutations in dominantly inherited familial ALS (FALS). We previously reported a coding mutation in D-amino acid oxidase (DAO(R199W)) associated with FALS. DAO metabolises D-serine, an essential co-agonist at the N-Methyl-D-aspartic acid glutamate receptor subtype (NMDAR). Using primary motor neuron cultures or motor neuron cell lines we demonstrated that expression of DAO(R199W), promoted the formation of ubiquitinated protein aggregates, activated autophagy and increased apoptosis. The aim of this study was to characterise the effects of DAO(R199W) in vivo, using transgenic mice overexpressing DAO(R199W). Marked abnormal motor features, e.g. kyphosis, were evident in mice expressing DAO(R199W), which were associated with a significant loss (19%) of lumbar spinal cord motor neurons, analysed at 14 months. When separated by gender, this effect was greater in females (26%; p< 0.0132). In addition, we crossed the DAO(R199W) transgenic mouse line with the SOD1(G93A) mouse model of ALS to determine whether the effects of SOD1(G93A) were potentiated in the double transgenic line (DAO(R199W)/SOD1(G93A)). Although overall survival was not affected, onset of neurological signs was significantly earlier in female double transgenic animals than their female SOD1(G93A) littermates (125 days vs 131 days, P = 0.0239). In summary, some significant in vivo effects of DAO(R199W) on motor neuron function (i.e. kyphosis and loss of motor neurons) were detected which were most marked in females and could contribute to the earlier onset of neurological signs in double transgenic females compared to SOD1(G93A) littermates, highlighting the importance of recognizing gender effects present in animal models of ALS.