Synaptoproteomics of an animal model of depression with gene-environment interaction

of the general population, its pathogenesis is still essentially unknown. Recent studies highlighted the important influence of environmental stress factors on an individual’s genetic predisposition to develop mood disorders. Indeed, the experience of stressful events in childhood, such as neglect, abuse or parent loss, was found to increase the risk for the development of depression in adult life. The Flinders Sensitive Line (FSL) rats are a well-validated animal model of depression carrying genetic vulnerability associated to distinct features of pathology and to responsiveness to antidepressant drugs. To reproduce early life stress events the FSL rats and their control, the Flinders Resistant Line (FRL) rats, were subjected to a maternal separation (MS) protocol (180 min/day from postnatal days 2 to 14). Treatment with the antidepressant (AD) escitalopram (ESC, 25 mg/kg/day), was carried out at weeks 11-14 of age. Global analysis of protein expression is a powerful approach to gain insight into the molecular mechanisms underlying vulnerability to psychiatric disorders and the long-term action of drug treatments. Many of the biological targets of AD are localized at synapses; thus to reduce the complexity of the proteome analyzed and to enrich for less abundant proteins, purified nerve terminals (synaptosomes) from prefrontal/frontal cortex (P/FC) and hippocampus (HC) of FSL and FRL rats were used. Synaptosomes from 8 rats per group were purified by differential centrifugation on Percoll gradients and analyzed by two-dimensional polyacrylamide gel electrophoresis (2DE). Statistical analysis of 2DE maps from P/FC synaptosomes revealed 37 proteins differently regulated in basal FSL vs FRL rats. MS significantly dysregulated 48 proteins in FSL, and 24 proteins in FRL P/FC synaptosomes. Chronic ESC treatment differently regulated 33 protein spots in basal FSL, and 7 protein spots in FSL subjected to MS. Statistical analysis of 2DE maps from HC synaptosomes revealed 15 protein spots differentially regulated in FSL vs. FRL rats. ESC regulated 13 spots in basal FSL. MS regulated 17 spots in FRL and 14 in FSL, subsequent pharmacological treatment with ESC regulated, in FSL rats maternally separated, 12 protein spots. Interestingly, the overall number of protein spots significantly up- or down-regulated in the different experimental groups, is fewer in HC than in P/FC synaptosomes. Contrary to P/FC, stress of MS differently regulated in HC almost the same number of proteins in FRL and FSL. However, similar to the results obtained in P/FC, MS regulated different set of proteins in the two lines of rats. Protein spots differently regulated in the various comparisons were excised from gels and identified by mass spectrometry analysis, by comparison with SwissProt and NCBI databases. In the various comparisons between groups, proteins related to synaptic function (NSF, αCaMKII, Munc-18, Dynamin-1, Clathrin light chain B) and to stress response or oxidative stress and apoptotic pathways (Prohibitin, 14-3-3 protein, Peroxiredoxine 6, Aconitase hydratase, Voltage-dependent anion selective channel protein 1) were identified. Heim, C., Nemeroff C.B. 2001 The role of childhood trauma in the neurobiology of mood and anxiety disorders: preclinical and clinical studies. Biol. Psychiatry 49: 1023-1039 Overstreet D.H., Friedman E., Mathé A.A., Yadid G. 2005 The Flinders Sensitive Line rat: a selectively bred putative animal model of depression. Neurosci. Biobehav. Rev. 29: 739-759 Vercauteren F.G.G., Bergeron J.J.M., Vandesande F., Arckens L., Quirion R. 2004 Proteomics approaches in brain research and neuropharmacology. Eur. J. Pharmacol. 500: 385-398