{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,31]],"date-time":"2026-07-31T14:15:55Z","timestamp":1785507355545,"version":"3.56.0"},"reference-count":49,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2026,7,31]],"date-time":"2026-07-31T00:00:00Z","timestamp":1785456000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Shanxi Provincial Administration of Traditional Chinese Medicine","award":["2024ZYYB028"],"award-info":[{"award-number":["2024ZYYB028"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Metabolites"],"abstract":"<jats:p>Objectives: Insomnia is a highly prevalent sleep disorder involving complex neurochem-ical dysregulation; however, the spatial distribution of neurotransmitters and small-molecule metabolites across distinct brain microregions in the insomniac state re-mains poorly characterized. This study aimed to map region-specific metabolic perturba-tions in situ in a p-chlorophenylalanine (PCPA)-induced insomnia rat model using opti-mized matrix-assisted laser desorption\/ionization mass spectrometry imaging (MAL-DI-MSI) integrated with targeted metabolomics validation. Methods: On-tissue chemical derivatization MALDI-MSI was optimized using \u03b1-cyano-4-hydroxycinnamic acid (CHCA) as the matrix on a Bruker tims TOF flex mass spectrometer. The rats received PCPA (400 mg\/kg, intraperitoneal) for three days to establish the insomnia model. Metabolite identifi-cation was conducted using MetaboScape\u00ae software (2020b) and the Human Metabolome Database. Ultra-performance liquid chromatography\u2013tandem mass spectrometry was em-ployed to quantify nine key neurotransmitters and metabolites across six brain microre-gions. Key synthetic enzyme expression was evaluated by immunofluorescence and West-ern blotting analyses. Results: TMP-TFB-derived brain slices clearly showed the distribu-tion of neurotransmitters in brain microregions. MALDI-MSI demonstrated that the spatial distribution and abundance of eight neurotransmitters were disturbed in brains of PCPA-induced insomniac rats. A total of 346 metabolites were characterized across six brain microregions (cerebellum, cortex, hippocampus, brainstem, hypothalamus, and stri-atum), with principal coordinate analysis revealing clear metabolic separation between control and insomniac rats. PCPA treatment markedly disrupted tryptophan and tyrosine metabolism, evidenced by decreased 5-HT, 5-HTP, and 5-HIAA, alongside region-specific alterations in DA, NE, HVA, and Ach. Additionally, GABA levels decreased in the hippo-campus, striatum, and hypothalamus, whereas glutamate increased throughout the brain. Targeted metabolomics validated the MSI findings, and Bland\u2013Altman analysis confirmed good consistency between the two analytical platforms. PCPA further disturbed the meta-bolic enzymes MAOA, DDC, and TPH2 within the Trp-5-HTP-5-HT-5-HIAA metabolic pathway in the brainstem and TYH and DBA within the tyrosine-DA-NE metabolic pathway in the striatum. Conclusions: This study demonstrates that region-specific altera-tions in tryptophan and tyrosine metabolism pathways provide mechanistic insights into insomnia pathogenesis and potential therapeutic targets.<\/jats:p>","DOI":"10.3390\/metabo16080543","type":"journal-article","created":{"date-parts":[[2026,7,31]],"date-time":"2026-07-31T13:23:20Z","timestamp":1785504200000},"page":"543","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Spatial Metabolomics Reveals Microregion-Specific Neurochemical Perturbations in the Brains of PCPA-Induced Insomniac Rats: Integration of MALDI-MSI and Targeted LC-MS\/MS"],"prefix":"10.3390","volume":"16","author":[{"given":"Yan","family":"Yan","sequence":"first","affiliation":[{"name":"Shanxi Bethune Hospital, Shanxi Academy of Medical Sciences, Third Hospital of Shanxi Medical University, Tongji Shanxi Hospital, Taiyuan 030032, China"},{"name":"Modern Research Center for Traditional Chinese Medicine, Shanxi University, Taiyuan 030006, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiaying","family":"Liu","sequence":"additional","affiliation":[{"name":"Modern Research Center for Traditional Chinese Medicine, Shanxi University, Taiyuan 030006, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yingjian","family":"Deng","sequence":"additional","affiliation":[{"name":"Modern Research Center for Traditional Chinese Medicine, Shanxi University, Taiyuan 030006, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu","family":"Tao","sequence":"additional","affiliation":[{"name":"Modern Research Center for Traditional Chinese Medicine, Shanxi University, Taiyuan 030006, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xinxin","family":"Li","sequence":"additional","affiliation":[{"name":"Modern Research Center for Traditional Chinese Medicine, Shanxi University, Taiyuan 030006, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0931-8878","authenticated-orcid":false,"given":"Chenhui","family":"Du","sequence":"additional","affiliation":[{"name":"School of Traditional Chinese Materia Medica, Shanxi University of Chinese Medicine, Taiyuan 030619, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kun","family":"Yang","sequence":"additional","affiliation":[{"name":"Shanxi Bethune Hospital, Shanxi Academy of Medical Sciences, Third Hospital of Shanxi Medical University, Tongji Shanxi Hospital, Taiyuan 030032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6922-0747","authenticated-orcid":false,"given":"Ruiping","family":"Zhang","sequence":"additional","affiliation":[{"name":"Shanxi Bethune Hospital, Shanxi Academy of Medical Sciences, Third Hospital of Shanxi Medical University, Tongji Shanxi Hospital, Taiyuan 030032, China"},{"name":"The Radiology Department, Shanxi Provincial People\u2019s Hospital Affiliated to Shanxi Medical University, Taiyuan 030001, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,7,31]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1016\/j.jsmc.2022.03.003","article-title":"Epidemiology of Insomnia: Prevalence, Course, Risk Factors, and Public Health Burden","volume":"17","author":"Morin","year":"2022","journal-title":"Sleep Med. 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