Adiposity tunes anxiety-related behaviours through insular cortex dysregulation in mice

Anxiety is an adaptive emotion essential for survival. However, it becomes pathological when it persists in the absence of threat. Anxiety disorders affect approximately 18% of the population annually. Although underlying neurobiological mechanisms of anxiety disorders remain poorly understood, obesity has emerged as a significant risk factor. Notably, clinical and preclinical evidence indicates that the insular cortex (insula), a key region involved in interoception and emotional processing, is altered in both patients with obesity or anxiety disorders. Importantly, obesity is a gradual disease with increased metabolic risk consequence in a higher degree of obesity. However, whether increasing levels of adiposity progressively exacerbate anxiety remains unclear. Thus, we tested whether the degree of adiposity gradually modulates the level of anxiety-related behaviours in mice, and whether the insular cortex contributes to this control.

To assess the impact of overweight on anxiety, we subjected 8-weeks-old C57BL/6J male and female mice to standard chow diet (SD, 3.23kcal/g), a moderate high fat diet (HCD, 4.73 kcal/g, 45% fat) or a severe HCD (5.24 kcal/g, 60% fat) for 12 weeks. We evaluated the anxiety level of SD-fed mice or HCD-fed mice by using the elevated plus maze (EPM), the open field test, the dark light box and the novelty suppressed feeding test (NSFT). Social behaviour was assessed using reciprocal social interaction test. After a break, mice were subjected to an anxiety challenge, followed by perfusion for whole-brain tissue clearing and cfos staining and imaging using light-sheet microscopy. To assess the potential contribution of the anterior insula, calcium imaging of glutamatergic neurons of the anterior insula was performed. Behaviour was analysed using DeepLabCut in combination with SimBA, and compared using two-way ANOVA. Whole brain cfos mapping was analysed using ClearMap and statistically compared by fitting linear models for each brain region. Calcium recordings were analysed using custom Matlab scripts and compared using two-way ANOVA.

Increased adiposity was linked to heightened anxiety-like behaviours, including decreased time spent in exposed areas in the EPM and NSFT, while locomotor activity remained unaffected. These effects displayed a dose-dependent pattern across diet groups, with the strongest phenotype observed in the severe-HCD group, followed by the moderate-HCD group. Surprisingly, increased adiposity mildly promotes sociability during social interaction. Preliminary analysis of whole brain cfos mapping indicates increased neuronal activity in HCD-fed animals in key brain regions associated with anxiety including the insula. Ongoing studies are underway assessing the in vivo neuronal activity during behavioural tasks to determine whether these neural changes could mediate anxiety.

These findings suggest that the degree of adiposity is a primary driver of comorbid anxiety. Moreover, this study identifies the insula as a potential substrate linking obesity and anxiety, consistent with its established role in interoception and emotional processing. Importantly, our results highlight the need to stratify patients with pathological anxiety based on adiposity, as this subgroup may benefit from more tailored, personalized therapeutic approaches. Future studies will aim to identify peripheral signals that may contribute to the observed activity in the insula and its impact on anxiety-related behaviours.

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