BACKGROUND: Tuberous sclerosis complex (TSC) is a rare monogenic disorder 
characterized by benign tumors in multiple organs as well as a high prevalence 
of epilepsy, intellectual disability and autism. TSC is caused by inactivating 
mutations in the TSC1 or TSC2 genes. Heterozygocity induces hyperactivation of 
mTOR which can be inhibited by mTOR inhibitors, such as rapamycin, which have 
proven efficacy in the treatment of TSC-associated symptoms. The aim of the 
present study was (1) to identify molecular changes associated with social and 
cognitive deficits in the brain tissue of Tsc1+/- mice and (2) to investigate 
the molecular effects of rapamycin treatment, which has been shown to ameliorate 
genotype-related behavioural deficits.
METHODS: Molecular alterations in the frontal cortex and hippocampus of Tsc1+/- 
and control mice, with or without rapamycin treatment, were investigated. A 
quantitative mass spectrometry-based shotgun proteomic approach (LC-MSE) was 
employed as an unbiased method to detect changes in protein levels. Changes 
identified in the initial profiling stage were validated using selected reaction 
monitoring (SRM). Protein Set Enrichment Analysis was employed to identify 
dysregulated pathways.
RESULTS: LC-MSE analysis of Tsc1+/- mice and controls (n = 30) identified 51 
proteins changed in frontal cortex and 108 in the hippocampus. Bioinformatic 
analysis combined with targeted proteomic validation revealed several 
dysregulated molecular pathways. Using targeted assays, proteomic alterations in 
the hippocampus validated the pathways "myelination", "dendrite," and "oxidative 
stress", an upregulation of ribosomal proteins and the mTOR kinase. LC-MSE 
analysis was also employed on Tsc1+/- and wildtype mice (n = 34) treated with 
rapamycin or vehicle. Rapamycin treatment exerted a stronger proteomic effect in 
Tsc1+/- mice with significant changes (mainly decreased expression) in 231 and 
106 proteins, respectively. The cellular pathways "oxidative stress" and 
"apoptosis" were found to be affected in Tsc1+/- mice and the cellular 
compartments "myelin sheet" and "neurofilaments" were affected by rapamycin 
treatment. Thirty-three proteins which were altered in Tsc1+/- mice were 
normalized following rapamycin treatment, amongst them oxidative stress related 
proteins, myelin-specific and ribosomal proteins.
CONCLUSIONS: Molecular changes in the Tsc1+/- mouse brain were more prominent in 
the hippocampus compared to the frontal cortex. Pathways linked to myelination 
and oxidative stress response were prominently affected and, at least in part, 
normalized following rapamycin treatment. The results could aid in the 
identification of novel drug targets for the treatment of cognitive, social and 
psychiatric symptoms in autism spectrum disorders. Similar pathways have also 
been implicated in other psychiatric and neurodegenerative disorders and could 
imply similar disease processes. Thus, the potential efficacy of mTOR inhibitors 
warrants further investigation not only for autism spectrum disorders but also 
for other neuropsychiatric and neurodegenerative diseases.