# Acod1/itaconate axis controls anxiety-like behaviors induced by chronic infection of Toxoplasma gondii in mice

## TL;DR

This study shows that the Acod1/itaconate pathway helps control anxiety caused by chronic Toxoplasma gondii infection in mice by reducing brain inflammation.

## Contribution

The study reveals the Acod1/itaconate axis as a novel immune-metabolic regulator of infection-induced anxiety and a potential therapeutic target.

## Key findings

- Chronic T. gondii infection activates the Acod1/itaconate axis in the amygdala and causes anxiety-like behaviors.
- DI treatment reduces neuroinflammation and anxiety by suppressing M1 microglia polarization and activating Nrf2 signaling.
- DI restores serotonin levels by reducing IDO expression, alleviating infection-induced anxiety in mice.

## Abstract

Chronic infection of Toxoplasma gondii (T. gondii) induces the anxiety-like behavior in hosts, which is closely linked to neuroinflammatory processes. Cis-aconite decarboxylase 1 (Acod1) is an enzyme that is responsible for itaconate production in Krebs Cycle. Emerging evidence highlights the Acod1/itaconate axis as a key regulatory node in macrophage immune-metabolic reprogramming. However, its role in infection-induced neurobehavioral alterations remains unclear. Here, we investigated the role of Acod1/itaconate axis in the anxiety induced by T. gondii chronic infection in mice.

To assess anxiety-like behaviors, we performed open field test and elevated plus maze test. Transcriptomic alterations and neuroinflammatory responses in the mouse amygdala were profiled via RNA sequencing, immunofluorescence staining, quantitative PCR (qPCR), and western blot. The functional role of the Acod1/itaconate axis was further investigated using Acod1-/- mice. Additionally, the therapeutic potential of dimethyl itaconate (DI), a cell-permeable itaconate derivative, was evaluated in chronically T. gondii-infected mice. The levels of indoleamine 2,3-dioxygenase (IDO), and serotonin (5-hydroxytryptamine, 5-HT) in serum were measured by enzyme-linked immunosorbent assay. Finally, DI’s anti-inflammatory mechanism was identified in the microglial cell line BV-2 cells.

Chronic T. gondii infection induced anxiety-like behaviors in mice and triggered the activation of Acod1/itaconate axis in the amygdala. Transcriptomic and histological analyses revealed upregulation of neuroinflammation-related genes, along with microglia activation. Genetic knockout of Acod1 induced the anxiety-like phenotypes, which were rescued by DI administration. Notably, DI treatment conferred both prophylactic and therapeutic benefits, effectively mitigating anxiety induced by infection. Mechanistically, DI suppressed T. gondii-induced M1 polarization in microglia to mitigate neuroinflammation via activating Nrf2 signaling. These events further reduced indoleamine IDO expression, leading to increased 5-HT levels and subsequent amelioration of anxiety-like behavior.

Our findings demonstrate that the Acod1/itaconate axis plays an important role in regulating anxiety-like behavior by modulating neuroinflammation during chronic T. gondii infection. These results reveal a promising immune-metabolic drug target for treating T. gondii-associated neuropsychiatric conditions.

Anxiety is closely linked to neuroimmune dysfunction. Chronic infection with the neurotropic parasite T. gondii triggers significant neuroinflammation in the amygdala, a key brain region for emotional processing, leading to anxiety-like behaviors. This study identifies the immune-metabolic Acod1/itaconate axis as a central regulator in this process. Using Acod1 knockout mice and pharmacological intervention with DI, a cell-permeable itaconate derivative, we demonstrate that DI effectively alleviates infection-induced anxiety. Mechanistically, DI activates the protective Nrf2 signaling pathway, which in turn dampens microglial-driven neuroinflammation. Furthermore, DI treatment reduces the expression of IDO, thereby restoring the balance of serotonin (5-HT), a crucial neuromodulator involved in mood regulation. Our findings not only provide a novel molecular perspective on how chronic infection promotes anxiety but also highlight the Acod1/itaconate axis as a promising therapeutic target for treating neuropsychiatric sequelae associated with pathogenic infection.