SPATIAL BRAIN PROTEOMIC ANALYSIS IN A CLOZAPINE-TREATED SCHIZOPHRENIA-LIKE MOUSE MODEL

Abstract Background Schizophrenia is a complex psychiatric disorder characterized by a spectrum of cognitive, emotional, and behavioral symptoms. Despite substantial research efforts and the accumulation of evidence, the definitive molecular mechanisms underlying schizophrenia remain elusive, with up to 30% of patients exhibiting treatment-resistance. Clozapine (CLZ), an atypical antipsychotic, has demonstrated remarkable effectiveness in the treatment on treatment-resistant schizophrenia, especially in cases where other antipsychotics have been ineffective. Elucidating the molecular mechanisms of clozapine can contribute to the development of effective treatment strategies; however, the impact on the expression of brain molecules involved in its pharmacological actions is not fully understood. Aims & Objectives This study aims to investigate spatial alterations in the brain proteome of a schizophrenia-like mouse model administered phencyclidine (PCP) or subsequently treated with CLZ, and to obtain insights into the neurobiological pathways that underlie the therapeutic effects of CLZ. Methods PCP (10 mg/kg, s.c.) or saline (10 mL/kg, s.c.) was injected once a day for 14 consecutive days to the 6- week-old male ICR strain mice. After PCP or saline administration, CLZ (10 mg/kg or 30 mg/kg, p.o.) or vehicle (10 mL/kg, p.o.) was treated once a day for 7 consecutive days. Twenty-four hours after the last administration, the prefrontal cortex (PFC), nucleus accumbens (NAc), striatum (STR), hippocampus (HIP), and amygdala (AMY) were collected and subjected to proteomic analysis. Protein extraction, purification, and digestion from the brain samples were carried out, followed by nanoflow liquid chromatography-tandem mass spectrometry for comprehensive proteome profiling. Results We performed pathway analysis on differentially expressed proteins (DEPs) and confirmed their biological significance. As a result, DEPs related to the interaction of the two factors (PCP and CLZ) were associated with various pathways, including glutamatergic synapses in the PFC, retrograde endocannabinoid signaling in the STR, Alzheimer's disease in the HIP, oxidative phosphorylation in the AMY. Several pathways were common to multiple brain regions; retrograde endocannabinoid signaling in the PFC, STR, and AMY, synaptic vesicle cycle in the PFC, STR, and HIP, Parkinson's disease in the PFC and STR, cGMP-PKG signaling pathway in the PFC and AMY, and glycolysis/gluconeogenesis in the HIP and AMY. Discussion & Conclusion Our study has provided a multitude of insights into the spatial profiles of protein expression within the multiple brain regions of interest following administration of PCP or CLZ. Interestingly, a common molecular pathway related to the synaptic vesicle cycle has been identified in the PFC, STR, and HIP, which are involved in behavioral phenotypes, such as cognitive impairments and emotional disturbances. Elucidating the functional roles of DEPs associated with common or specific pathways across various brain regions may contribute to the development of novel treatment strategies for treatment-resistant schizophrenia.