Background <p>Environmental stressors and neurotoxicants during prenatal development are known to be risk factors for psychiatric disorders. However, their combined molecular effects remain elusive. Thus, this study aimed to investigate synergistic effects of prenatal stress and lead (Pb) exposure on hippocampal proteome alterations and behavioral outcomes in a rat model.</p> Methods <p>Pregnant Sprague Dawley rats were subjected to repeated variable stress during gestational days 14–21 and were administered 0.2% lead acetate via drinking water throughout gestation and lactation. In addition, a repeated variable stress paradigm was applied to the pregnant rats during the last week of gestation. Male offspring underwent behavioral assessments, including pre-pulse inhibition, social interaction, and open-field tests. Hippocampal protein expression was analyzed using two-dimensional electrophoresis coupled with MALDI-TOF mass spectrometry and validated by western blot analysis.</p> Results <p>Combined exposure significantly altered anxiety-like behavior, reduced social interactions, and impaired sensorimotor gating in the offspring. Proteomic analysis revealed 16 proteins that were differentially expressed in the hippocampus. Notably, western blot validation confirmed the significant downregulation of phosphatidylethanolamine binding protein 1 (Pebp1) and peroxiredoxin-2 (Prdx2).</p> Conclusions <p>This study demonstrates that concurrent prenatal stress and Pb exposure can induce specific molecular signatures in the hippocampal proteome alongside behavioral deficits characteristic of psychiatric disorders. The identification of Pebp1 and Prdx2 as key molecular targets provides new insights into potential therapeutic interventions for environmental exposure-related psychiatric disorders.</p>