Synapses are specialised contacts between neurons. At postsynaptic terminals of glutamatergic synapses, protein complexes process and transmit the information received from the presynaptic terminal. Scaffolding proteins, among which members of the disc large homologue (DLG) family are the most abundant, assemble the molecular machinery in the postsynaptic terminal. Recently, two members of the DLG family, postsynaptic density protein 95 (PSD95) and synapse associated protein 102 (SAP102), have been shown to form different types of complexes, thus giving the synapse different signalling capabilities. However, the spatial distribution of these synaptic markers in different synapses remains elusive due to technical challenges. This thesis presents the first applications of a new method, the Genes to Cognition Synaptome Mapping pipeline (G2CSynMapp), to map individual synapses at the whole-brain level, in a quantitative and unbiased manner. This method was used to generate PSD95 and SAP102 synaptome maps – i.e. comprehensive maps of PSD95 and SAP102 positive synapses – in the mouse brain and to achieve three aims: i) characterise PSD95 and SAP102 synapse diversity, ii) measure the trajectory of PSD95 and SAP102 synapse changes during the postnatal lifespan and iii) determine whether PSD95 synaptome is reorganised by mutation. First, I have used G2CSynMapp to generate the first synaptome maps in the adult mouse brain. This reference map of PSD95 and SAP102 positive synapses revealed a highly organised distribution pattern of glutamatergic synapses between anatomical regions. Moreover, it uncovered that synapse populations are very diverse within anatomical regions and can form patches, gradients and input-specific glomeruli. Second, the trajectories of PSD95 and SAP102 synaptomes were mapped across the mouse postnatal lifespan. At birth, synapse densities are low and increase rapidly during the first month of life. During ageing, the density of SAP102 and PSD95 positive synapses decrease gradually. Interestingly, different anatomical regions show different trajectories of synapse density and parameters across the lifespan. Moreover, the packing of PSD95 and SAP102 at synapses have specific pattern of changes. Third, the PSD95 synaptome was found to be reorganised differently in two disease models, PSD93 and SAP102 knock-out mice. In humans, mutations in the genes encoding PSD93 or SAP102 have been involved in schizophrenia and mental retardation, respectively. Of particular interest, opposite changes were identified in the neocortex of the two mutant lines that are reminiscent of their inverse behavioural phenotypes.

Hyperactivation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) by dopamine D₁ receptor (D₁R) in the striatum is a characteristic feature of several neuropsychiatric conditions, including drug addiction and L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesia (LID) within individuals suffering from Parkinson’s disease. However, the current mechanistic gap in understanding of D₁R-mediated regulation of ERK1/2, both in physiology and disease, hampers effective treatment of these conditions. One important factor that is underexplored in this regard is the role played by proteins that physically interact with the intracellular regions of D₁R. Using yeast two-hybrid screens and co-transfected human embryonic kidney 293 (HEK293) cells, our laboratory has recently characterized an interaction between the third intracellular loop of D₁R and synapse-associated protein 102 (SAP102), a member of the membrane-associated guanylate kinase family. Moreover, our lab identified endogenous D₁R-SAP102 complex within rat striatum and hippocampus. Interestingly, SAP102 regulates ERK signaling pathway within the hippocampus, and modulates adenosine A2A receptor-mediated ERK1/2 activation within transfected HEK293 cells. Capitalizing on the above findings, I hypothesized a role for SAP102 in controlling D₁R-mediated ERK1/2 activation. Herein, I demonstrate using co-transfected HEK293 cells that SAP102 alters the temporal activation of ERK1/2 by D₁R. Intriguingly, experiments using the protein kinase A (PKA) inhibitors H89 and protein kinase inhibitor 14-22 amide myristoylated show that SAP102 also facilitates a switch in D₁R-mediated ERK1/2 activation from a PKA-independent to PKA-dependent pathway. Furthermore, SAP102 reduces basal ERK1/2 activation in HEK293 cells, reminiscent of a previously documented role of hippocampal SAP102. To the best of my knowledge, my findings are the first to demonstrate scaffolding protein-mediated switching of a G protein-coupled receptor (GPCR) signaling pathway. Future studies aimed at uncovering the details of this process should provide valuable insight on the mechanisms contributing to D₁R-mediated ERK1/2 activation, and may also offer clues to the existence of similar phenomena for other GPCRs. These studies would also aid development of improved pharmacological treatment options for conditions with dysfunctional D₁R-dependent ERK1/2 activation.

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