412. Study of behavioural, molecular, morphological and electrophysiological effects of chronic stress in mice

Abstract

Abstract Background Chronic stress profoundly affects brain function and constitutes a major risk factor for both psychiatric and systemic pathologies. Understanding its impact necessitates an integrated approach linking behavioural, molecular, and neural mechanisms. While biological variables such as sex and circadian timing are known to shape the stress responses, how these factors interact with underlying synaptic and functional alterations requires further elucidation. Aims & Objectives This study investigates sex- and circadian-dependent effects of chronic stress on behaviour. Furthermore, we analysed changes in hippocampal synaptoproteome composition, translatome, and neuronal morphology in female mice stressed in the light phase, and alterations of brain activity in male mice stressed in the light phase. Method Male and female C57BL/6 mice were subjected to a Chronic Restraint Stress (CRS) paradigm (2 h/day for 21 days) during either the light or dark phase. A battery of behavioural tests assessed depressive-like (Tail Suspension Test, Forced Swim Test), anxiety-like (Open Field Test), anhedonic-like (Sucrose splash test) phenotypes and cognition/memory impairments (Novel Object Recognition Test). Bases on behavioural outcomes, female mice stressed in the light phase were selected for proteomic profiling of hippocampal synaptosomes to identify molecular pathways compromised by stress, while dendritic architecture, specifically in terms of dendritic length and dendritic spine density of CA1 pyramidal neurons and DG granule cells, is being evaluated via Golgi Staining. Furthermore, the Translating Ribosome Affinity Purification (TRAP) technique was employed to isolate and characterize cell-type-specific translatomes (ribosome-associated mRNA), elucidating distinct translational signatures within discrete hippocampal subpopulations. Lastly, EEG recordings through EEG telemetric system at the end of each week of the CRS protocol and during OFT and NORT are assessing stress-induced alterations in neural oscillations during emotional and cognitive tasks in male mice stressed in the light phase. Results Our findings reveal distinct behavioural and molecular stratifications dependent on both sex and circadian phase. Female mice showed increased stress sensitivity in both behavioural domains and body weight trajectories compared to males. Female mice exposed to stress during the light phase displayed a depressive-like phenotype without overlapping anxiety traits while, when stressed during the dark phase, developed an anxious-like behaviour. Conversely, male mice developed depressive and anhedonic behaviours only if stressed in the light phase but not in the dark phase. Proteomic analyses of hippocampal synaptosomes from female mice stressed in the light phase identified stress-induced modulation of synaptic proteins. Ongoing molecular, morphological, and electrophysiological investigations are expected to further delineate how chronic stress reshapes neuronal connectivity and activity dynamics. Discussion & Conclusions By combining behavioural, proteomic, morphological, and functional approaches, this study provides an innovative, multi-scale perspective on the impact of chronic stress on the brain. The results underscore the complex interplay among biological variables and molecular networks, offering novel insights into mechanisms of stress.