The complex structural and functional organisation of the brain warrants the application of high-throughput approaches to study its functional alterations in physiological and pathological conditions. Such approaches have greatly benefited from advances in proteomics and genomics, and from their combination with computational modelling. They have been particularly instrumental for the analysis of processes such as the post-translational modification (PTM) of proteins, a critical biological process in the nervous system that remains not well studied. Protein PTMs are dynamic covalent marks that can be induced by activity and allow the maintenance of a trace of this activity. In the nucleus, they can modulate histone proteins and the components of the transcriptional machinery, and thereby contribute to regulating gene expression. PTMs do however need to be tightly controlled for proper chromatin functions. This review provides a synopsis of methods available to study PTMs and protein expression based on high-throughput mass spectrometry (MS), and covers basic concepts of traditional 'shot-gun'-based MS. It describes classical and emerging proteomic approaches such as multiple reaction monitoring and electron transfer dissociation, and their application to the analyses of nuclear processes in the brain.