Aspartame Linked to Memory Loss, Anxiety, and Insulin Resistance in Rats

Aspartame, the artificial sweetener found in diet sodas, sugar-free gum, and countless processed foods, has long been marketed as a healthier alternative to sugar. But a new study published in the Journal of Agriculture and Food Research suggests that this ubiquitous sweetener may be doing far more harm than good. Using an innovative combination of network toxicology, molecular docking, and animal experiments, researchers have uncovered evidence linking chronic aspartame exposure to memory impairment, anxiety-like behavior, insulin resistance, and even gut microbiome disruption.

The research team, led by Yue Zhu and colleagues, began by mapping the potential molecular targets of aspartame and its three major metabolites—phenylalanine, aspartate, and methanol—using computational databases. They then cross-referenced these targets with genes associated with four major diseases: Alzheimer’s disease, Parkinson’s disease, type 2 diabetes mellitus, and obesity. The analysis revealed hundreds of shared targets, with five core proteins—EGFR, HDAC1, IL-1β, PPARγ, and RXRA—emerging as key hubs across at least three of the four conditions.

Molecular docking simulations showed that aspartame could bind favorably to all five core target proteins, with predicted binding affinities below −6.0 kcal/mol. The strongest interactions were observed with RXRA (−6.801 kcal/mol) and PPARγ (−6.784 kcal/mol), both nuclear receptors involved in lipid metabolism and insulin signaling. These computational findings provided a structural basis for the subsequent in vivo experiments.

To test whether these predicted molecular interactions translate into real biological effects, the researchers exposed male Sprague–Dawley rats to aspartame at three doses—75, 150, and 300 mg/kg/day—for eight consecutive weeks. The highest dose corresponds to a nominal human equivalent of approximately 48.6 mg/kg/day, just below the FDA’s acceptable daily intake of 50 mg/kg/day. Behavioral testing revealed striking dose-dependent deficits: rats in the medium- and high-dose groups took significantly longer to locate a hidden platform in the Morris water maze, spent less time exploring novel objects in the recognition test, and showed markedly reduced exploration of the central area in the open field test, indicating increased anxiety-like behavior.

Histological examination of the hippocampus revealed dose-dependent pathological changes, including reduced neuronal density, disorganized neuronal arrangement, and widened intercellular spaces. Immunohistochemical staining for Iba-1, a marker of microglial activation, showed increased microglial reactivity in the medium- and high-dose groups. Consistent with these findings, levels of the pro-inflammatory cytokines IL-1β and TNF-α were elevated in both serum and cerebrospinal fluid, suggesting that aspartame exposure triggers both central and peripheral inflammatory responses.

At the molecular level, Western blot analysis revealed significantly elevated HDAC1 protein expression in the hippocampus across all three aspartame groups, accompanied by a progressive increase in histone deacetylase activity. HDAC1 is an epigenetic regulator that modifies chromatin structure and influences neurogenesis, synaptic function, and DNA repair. The researchers suggest that the increase in HDAC1 may represent an early adaptive response to environmental stress rather than a feature of end-stage neurodegenerative pathology.

The metabolic consequences of aspartame exposure were equally concerning. Rats in the medium- and high-dose groups showed significantly impaired glucose tolerance, elevated insulin resistance as measured by HOMA-IR, and dose-dependent hepatic lipid accumulation confirmed by Oil Red O staining. Western blot and qPCR analysis of liver tissue revealed reduced PPARγ and RXRA protein expression, along with decreased mRNA levels of the lipid metabolism genes CD36 and FABP4, while SREBP-1c mRNA was significantly increased in the high-dose group. These findings suggest that aspartame exposure disrupts hepatic lipid metabolic control at the transcriptional level.

Perhaps most intriguingly, 16S rRNA gene sequencing of fecal samples revealed that high-dose aspartame exposure significantly reduced gut microbial alpha diversity and produced clear separation in overall community structure. Beneficial bacteria such as Roseburia and Lactobacillus were depleted, while opportunistic pathogens including Bacteroides and Desulfovibrio were enriched. Functional prediction analysis showed significant alterations in microbial pathways related to immune function, carbohydrate metabolism, and nervous system signaling, suggesting that gut dysbiosis may represent a key upstream mechanism linking aspartame exposure to systemic metabolic and neurological disturbances.

The authors emphasize that the neurobehavioral and metabolic alterations observed in this study may reflect the combined biological effects of aspartame and its metabolites, and cannot be attributed solely to the intact sweetener. Phenylalanine can cross the blood–brain barrier and compete with other large neutral amino acids for transport, while methanol is further metabolized to formaldehyde and formic acid, both of which may contribute to toxicological effects. However, because separate exposure groups for individual metabolites were not included, the relative contributions of each metabolite cannot be distinguished in the present study.

While the findings are compelling, the researchers acknowledge several limitations. The exposure period was limited to eight weeks, only male rats were included, and the observed molecular changes do not establish causal relationships. The gut microbiota findings should be regarded as exploratory evidence, as no microbiota intervention experiments were performed. Nevertheless, this integrated study provides experimental evidence for understanding the potential biological effects of sustained aspartame exposure and highlights the need for longer-term studies incorporating both sexes and appropriately designed human investigations to clarify these associations and their potential translational relevance.

Subject of Research: Health effects of aspartame exposure

Article Title: Aspartame Exposure-Associated Neurobehavioral Alterations and Metabolic Dysregulation: An Integrated Study Using Network Toxicology, Molecular Docking, and In Vivo Evaluation

Article References: Zhu, Y., Yu, F., Guo, W., Sun, G., & Wu, W. (2026). Aspartame Exposure-Associated Neurobehavioral Alterations and Metabolic Dysregulation: An Integrated Study Using Network Toxicology, Molecular Docking, and In Vivo Evaluation. Journal of Agriculture and Food Research, Article 103378. https://doi.org/10.1016/j.jafr.2026.103378

Image Credits: AI Generated

DOI: 10.1016/j.jafr.2026.103378

Keywords: aspartame, artificial sweeteners, network toxicology, molecular docking, insulin resistance, gut microbiome, neuroinflammation, HDAC1, PPARγ, type 2 diabetes, Exposure-Associated, Neurobehavioral