13
Aug
Chlorpyrifos Insecticide Disrupts Sleep, Brain Function, Immune System; Harm to Women Elevated
(Beyond Pesticides, August 13, 2025) The data on the adverse effects of the insecticide chlorpyrifos, still widely used in food production, continued to accumulate with the latest being a study published in PLOS One that finds perinatal exposure to the chemical in mice can alter sleeping patterns, lead to brain inflammation (particularly in female individuals), and impact gene expression linked to immune response and epigenetic effects. The adverse health effects are greater overall in female mice than male mice, emphasizing the significance of disproportionate impacts across species.
Chlorpyrifos has been a threat to human and ecological health for decades, originally as a general-use pesticide for homes, gardens, and agriculture, and then restricted to most nonresidential uses in 2000. Currently, the chemical’s permitted uses include food and feed crops, golf courses, as a non-structural wood treatment, and adult mosquito control for public health (insect-borne diseases) uses only. According to health and environmental advocates, there is a long history of U.S. Environmental Protection Agency (EPA) failure to adequately protect human and environmental health from chlorpyrifos, which is linked to endocrine disruption, reproductive effects, neurotoxicity, brain, kidney, and liver damage, and birth and developmental effects.
It took 21 years after negotiating a stop to most residential uses for EPA to negotiate a ban of the agricultural uses of chlorpyrifos in 2021 after a U.S. Court of Appeals found its registration to be flawed; then, that court decision was vacated by a 2023 Appeals Court decision (see also Daily News), sending EPA back to its pre-2021 allowed agricultural uses. In December 2024, EPA issued a proposed rule to restrict chlorpyrifos’ allowance to 11 major crops, which are among the most extensively grown and used in the U.S.: soybeans, sugar beets, cotton, wheat, apples, citrus fruits, strawberries, alfalfa, cherries, peaches, and asparagus. The public comment period closed in February 2025. (See Beyond Pesticides’ commentary on some of chlorpyrifos’ history.)
In addition to the adverse human health effects, chlorpyrifos is also known to be toxic to birds, bees, fish, and other aquatic organisms and is detectable in groundwater. Advocates continue to call for a more transformational approach rather than focusing on whack-a-mole tactics that focus on the harm of one pesticide after pesticide, rather than advocating for alternative agricultural and land management systems that render their perceived utility moot.
Background and Methodology
In the study, the test subjects are “wild-type C57BL/6†mice (a common strain of laboratory mouse species) in a controlled environment kept at 23-25 degrees Celsius (roughly 73 to 77 degrees Fahrenheit) on a 12-hour light-dark cycle. Mice are fed “ad libitum,†or as they desire. Mice were split up into groups exposed to chlorpyrifos via oral gavage in peanut oil and a control group. The exposure window was from mating until weaning, with mouse offspring (“pupsâ€) not directly exposed to the pesticide to home in on perinatal exposure.
The main hypothesis of this study was to determine if perinatal exposure to chlorpyrifos, during pregnancy and lactation periods, causes long-lasting disruptions in sleep-related breathing and promotes neuroinflammation for subjects moving into adulthood, possibly in a sex-specific manner. The researchers based their hypothesis on prior evidence of the insecticide crossing the placental barrier and showing up in breast milk, as well as links to adverse effects on neurodevelopment and respiration in humans and non-humans.
The experimental procedures to test for perinatal health include confirmation that the offspring were properly exposed to chlorpyrifos by assessing acetylcholinesterase (AChE) (enzyme necessary to nervous system transmission) activity, various behavioral tests (pre-birth surgery), surgical implantation of electrodes to assess brain activity and muscle movement, sleep and breathing recordings, blood test to assess stress via corticosterone levels, and oesterous cycle monitoring to assess behavioral and brain activity for female mice. After all the tests, the mice were euthanized and hippocampi collected to measure inflammation, anti-inflammatory regulation, stress response, and epigenetics. The researchers leveraged various statistical analysis tools to account for these various biological indicators, which can be found in more detail on pages three to seven of the study.
The researchers for this study are all experts at the University of Bologna in various specialties, including the Department of Pharmacy and Biotechnology, the Department of Biomedical and Neuromotor Sciences, the Department of Medical and Surgical Sciences, the Center for Applied Biomedical Research, and PRISM Lab. This study was approved by both the University’s Committee on the Ethics of Animal Experiments and the Italian Ministry of Health, following guidelines laid down by the Committee and ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines.
Results
The authors were able to prove their hypothesis that chlorpyrifos exposure leads to adverse health effects in mice, particularly on sleep, behavioral, and neurodevelopmental harms and disruptions. The six main categories of findings include:
- Mother mice treated with the pesticide had significantly reduced AChE activity, confirming that Chlorpyrifos was present during pregnancy and breastfeeding of offspring mice.
- Adult mice with perinatal Chlorpyrifos exposure faced more apneas and more sighs, with apneas occurring during both light and deep sleep cycles.
- The hippocampus of female mice showed higher levels of inflammatory genes and lower levels of their anti-inflammatory counterparts, suggesting chronic brain inflammation.
- Enzymes involved with regulating inflammation (histone demethylases) were significantly reduced in female mice
- Male mice exposed to the insecticide showed an increased expression of Nr3c1 (glucocorticoid receptor), which could signal an altered stress response regulation.
- Chlorpyrifos-exposed mice were described as having mixed results in terms of other cognitive and behavioral findings, including potential indicators for improved working memory or hyperactivity.
Previous Research and Actions
The scientific literature demonstrates that toxins like microplastics, heavy metals, synthetic fertilizers, per- and polyfluoroalkyl substances (PFAS), and pesticides (including whole formulations, active ingredients, “inerts”, synergists, pesticide-treated seeds, and plant growth regulators) have synergistic effects that threaten biodiversity, including aquatic organisms. This means the combined toxicity of the two (or multiple) substances is greater than the sum of the two individual exposures. The most recent study to demonstrate this, published in Ecotoxicology, focuses on the impacts of microplastics and chlorpyrifos on cladocerans, a group of microcrustaceans. There was no mortality observed in microplastic-only treatments, while microplastics preconditioned with the insecticide showed acute effects. Chronic exposure also shows reduced survival and reproductive output in both species. The researchers state: “A significant delay in age at first reproduction and shorter generation time were observed in the presence of MP^CPF, suggesting MP-mediated enhanced toxicity of CPF, wherein CPF could have accumulated onto the MP surface, thus, intensifying its toxicity.†(See Daily News here.) Researchers developed a novel tool in a study published in Nature Communications this year that successfully creates a map of the “pesticide-gut microbiota-metabolite network,†identifying “significant alterations in gut bacteria metabolism. Chlorpyrifos was one of eighteen pesticide compounds showing significant potential metabolic shifts in the gut microbiome. (See Daily News here.)
There has been a somewhat global shift in recognizing the toxicity of chlorpyrifos. The United Nations’ Conference of Parties (COP) for the Stockholm Convention on Persistent Organic Pollutants (POPs), originally adopted by 128 countries in 2001, voted to move chlorpyrifos, a neurotoxicant linked to brain damage in children, to Annex A (Elimination) with exemptions on a range of crops, control for ticks for cattle, and wood preservation, according to the POPs Review Committee. (See Daily News here.)
In terms of recent legal history on the organophosphate insecticide, a three-judge panel of the Ninth Circuit Court of Appeals in May 2021 instructed EPA to either revoke the tolerances the agency had set for chlorpyrifos’s residue in various foods or demonstrate that they meet the statutory and regulatory standards. Finally, after 21 years of delay, EPA issued a final rule in August 2021 revoking all food tolerances for the chemical. This seemed to signal a step in the right direction after relentless grassroots advocacy and judicial oversight prompted regulatory action until February 2022, when a different set of petitioners—pesticide corporations, groups representing industrial agriculture, and other countries’ agricultural interests vested in fossil fuel-dependent food systems—filed an action in the Eighth Circuit Court of Appeals. In November 2023, a three-judge panel of the Eighth Circuit reversed EPA’s momentous 2021 decision, rendering the Ninth Circuit’s opinion moot. (See additional Daily News coverage and additional commentary on the saga of chlorpyrifos litigation and regulations here, here, here, and here.)
Call to Action
Beyond Pesticides submitted comments (see here) earlier this year on a Federal Register notice to “to revoke all tolerances for residues of chlorpyrifos, except for those associated with the use of chlorpyrifos on the following crops: alfalfa, apple, asparagus, tart cherry, citrus, cotton, peach, soybean, strawberry, sugar beet, and spring and winter wheat.†Beyond Pesticides, citing alternatives and the clear weight of evidence on neurological and a suite of health impacts, called for the total cancellation of chlorpyrifos use. This builds on decades of previous advocacy in offering comments rooted in the latest scientific evidence and findings, as you can see in the pesticide’s entry in the Gateway on Pesticide Hazards and Safe Pest Management.
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All unattributed positions and opinions in this piece are those of Beyond Pesticides.
Source: PLOS One
