Compelling evidence suggests that major depression is associated with dysfunction of the brain glutamatergic transmission, and that the glutamatergic N-methyl-d-aspartate (NMDA) receptor plays a role in antidepressant activity. Recent post-mortem studies demonstrate that depression is associated with altered concentrations of proteins associated with NMDA receptor signalling in the brain. The present study investigated glutamate signalling proteins in the amygdala from depressed subjects, given strong evidence for amygdala pathology in depression. Lateral amygdala samples were obtained from 1314 pairs of age- sex-, and post-mortem-interval-matched depressed and psychiatrically healthy control subjects. Concentrations of NR1 and NR2A subunits of the NMDA receptor, as well as NMDA receptor-associated proteins such as post-synaptic density protein-95 (PSD-95) and neuronal nitric oxide synthase (nNOS) were measured by Western immunoblotting. Additionally, levels of enzymes involved in glutamate metabolism, including glutamine synthetase and glutamic acid decarboxylase (GAD-67), were measured in the same amygdala samples. NR2A protein levels were markedly and significantly elevated (+115%, p=0.03) in depressed subjects compared to controls. Interestingly, PSD-95 levels were also highly elevated (+128%, p=0.01) in the same depressed subjects relative to controls. Amounts of NR1, nNOS, glutamine synthetase, and GAD-67 were unchanged. Increased levels of NR2A and PSD-95 suggest that glutamate signalling at the NMDA receptor in the amygdala is disrupted in depression.

The N-methyl-D-aspartate receptor (NMDAR) hypofunctional hypothesis is believed to explain one of the contributing factors to schizophrenia. This hypothesis suggests the dysregulation of NMDAR, a protein responsible for receiving signals from the synapses between neurons, is the cause of some of the symptoms seen in schizophrenia. The post synaptic density protein 95 (PSD95) uses its PDZ-domains to help facilitate the received signal from NMDAR to α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) which in turn transmits the signal through the neuron. One way to increase the function of NMDAR could be to increase its affinity towards PDZ-domains of PSD95 using a small molecule. Fragment based drug design (FBDD) is one way to screen for molecules that modulates the NMDAR-PDZ interaction. This work describes the development of differential scanning fluorimetry (DSF) and surface plasmon resonance (SPR) assays using a fusion protein to screen for molecules that potentiate the interaction between NMDAR and AMPAR as well as methods assisting in the prioritization of hits based on both affinity, selectivity, and mechanism. The developed assays were used to screen a library containing 768 compounds. Screen positives and other compounds of interest were triaged and evaluated based on affinity, selectivity, and ability to modulated peptide binding resulting in eight confirmed hits that interacts with the two PDZ-domains of PSD95 investigated. As part of this work, the dissociation constant (KD) was determined for a panel of peptides representing versions of the truncated NMDAR GluN2b-subunit C-terminal towards PDZ1 and 2 of PSD95.