{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,6,15]],"date-time":"2026-06-15T09:56:04Z","timestamp":1781517364394,"version":"3.54.1"},"posted":{"date-parts":[[2026,6,11]]},"group-title":"Neuroscience","reference-count":82,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,6,11]],"date-time":"2026-06-11T00:00:00Z","timestamp":1781136000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"funder":[{"DOI":"10.13039\/100001024","name":"Roy J. Carver Charitable Trust","doi-asserted-by":"crossref","award":["Early Stage Investigator Award"],"award-info":[{"award-number":["Early Stage Investigator Award"]}],"id":[{"id":"10.13039\/100001024","id-type":"DOI","asserted-by":"crossref"}]},{"name":"Environmental Health Sciences Center","award":["NIH P30 ES005605"],"award-info":[{"award-number":["NIH P30 ES005605"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,6,11]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>Chlorpyrifos is a widely used organophosphate pesticide that exerts its primary toxic effect through inhibition of acetylcholinesterase (AChE). Although the acute neurotoxicity of chlorpyrifos is well characterized, the lasting biochemical, behavioral, and epigenetic consequences of sub-chronic exposure remain poorly understood, particularly when considering sex-specific differences. Therefore, we exposed male and female C57BL\/6J mice to either peanut oil (n=19), low chlorpyrifos exposure (1 mg\/kg\/day; n=19), or high chlorpyrifos exposure (10 mg\/kg\/day n=10) repeatedly for 21 days via subcutaneous injection. Blood AChE activity, behavior, and hippocampal DNA methylation were measured across groups. During exposure, AChE activity decreased in both males and females but only returned to baseline after behavioral testing in females exposed to low chlorpyrifos levels. Behavioral tests also revealed a sex-specific phenotype, with females in the low exposure group exhibiting reduced forced swim test immobility and a significant time by exposure interaction in open field habituation. No significant behavioral effects were observed in males. Significant DNA methylation changes were observed at 3,538 CpG sites in male and female mice after high exposure. Sex-specific analyses revealed two female-specific differentially methylated CpGs after high exposure. Pathways enriched for differentially methylated genes included several related to synaptic remodeling, cholinergic synapse, and various endocrine systems. These findings demonstrate that repeated high chlorpyrifos exposure leads to persistent cholinergic disruption and DNA methylation changes. However, the female-specific behavioral changes seen are independent of AChE activity and widespread DNA methylation changes, suggesting additional mechanisms, present only in females, may underlie behavioral sensitivity to chlorpyrifos.<\/jats:p>\n                <jats:sec>\n                  <jats:title>New and Noteworthy<\/jats:title>\n                  <jats:p>Sub-chronic chlorpyrifos exposure in adult mice produced dose-dependent blood AChE suppression and widespread hippocampal DNA methylation changes in both sexes, with pathway enrichment including cholinergic synapse and endocrine systems. Behavioral effects were subtle, with females in the low exposure group showing reduced forced swim immobility and altered locomotor habituation. Notably, the female-specific behavioral changes seen are independent of AChE activity and DNA methylation changes, suggesting novel mechanisms may underlie female behavioral sensitivity to chlorpyrifos.<\/jats:p>\n                <\/jats:sec>","DOI":"10.64898\/2026.06.08.730429","type":"posted-content","created":{"date-parts":[[2026,6,11]],"date-time":"2026-06-11T18:35:20Z","timestamp":1781202920000},"source":"Crossref","is-referenced-by-count":0,"title":["Sub-Chronic Chlorpyrifos Exposure Leads to Epigenetic and Sex-Specific Behavioral Changes in Adult Mice"],"prefix":"10.64898","author":[{"given":"Alyssa R.","family":"Daniel","sequence":"first","affiliation":[{"name":"Interdisciplinary Graduate Program in Human Toxicology, University of Iowa, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Noah","family":"Gernander","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, University of Iowa, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sara","family":"Dodge","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, University of Iowa, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cessily","family":"Hayes","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, University of Iowa, Iowa City, IA, USA"},{"name":"Department of Psychiatry, University of Iowa Carver College of Medicine, Iowa City, IA, USA"},{"name":"Iowa Neuroscience Institute, University of Iowa Carver College of Medicine, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emma","family":"Simpson-Wade","sequence":"additional","affiliation":[{"name":"Department of Molecular Medicine, University of Iowa Carver College of Medicine, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emese H. C.","family":"Kov\u00e1cs","sequence":"additional","affiliation":[{"name":"Department of Neuroscience and Pharmacology, University of Iowa Carver College of Medicine, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gibson","family":"Dowd","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, University of Iowa, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jared M.","family":"McLendon","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, University of Iowa, Iowa City, IA, USA"},{"name":"Iowa Neuroscience Institute, University of Iowa Carver College of Medicine, Iowa City, IA, USA"},{"name":"Division of Cardiovascular Medicine, Department of Internal Medicine, University of Iowa Carver College of Medicine, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benjamin","family":"Hing","sequence":"additional","affiliation":[{"name":"Department of Molecular Physiology and Biophysics, University of Iowa Carver College of Medicine, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9196-0028","authenticated-orcid":false,"given":"Marie E.","family":"Gaine","sequence":"additional","affiliation":[{"name":"Interdisciplinary Graduate Program in Human Toxicology, University of Iowa, Iowa City, IA, USA"},{"name":"Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, University of Iowa, Iowa City, IA, USA"},{"name":"Iowa Neuroscience Institute, University of Iowa Carver College of Medicine, Iowa City, IA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026061502500612000_2026.06.08.730429v1.1","unstructured":"Agency, E.P. 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