Protein-enriched diet modulates anxiety-like and metabolic adaptations in anorexia nervosa-like mice

Introduction

Anorexia nervosa (AN) is a severe psychiatric disorder characterised by restrictive eating, excessive physical activity, fear of weight gain and anxiety. Despite profound weight loss, patients with AN may maintain relative euglycaemia. Clinical observations from our group indicate avoidance of high-calorie foods and preference for low-calorie, protein- and fibre-enriched diets. As protein-rich diets may promote satiety and selective macronutrient intake could help regulate hunger, glycaemia and anxiety. This study tested whether a protein-enriched diet (PED) modifies metabolic adaptation, voluntary activity, anxiety-like behaviour and later food preference in an AN-like mouse model combining food restriction and physical activity.

Methods

Female C57BL/6J mice were assigned to four groups (n = 8 per group), according to feeding condition, ad libitum or 50% food restriction, and diet composition, standard diet containing 16% protein or PED containing 51% protein, for 20 days. All mice had access to a voluntary running wheel. Food intake, body weight, basal glycaemia and wheel activity were monitored longitudinally. Anxiety-like behaviour was assessed using the open field and elevated plus maze , followed by a two-choice food preference test for PED. Longitudinal outcomes were analysed using three-way analysis of variance followed by Sidak post hoc tests. Endpoint behavioural and preference outcomes were analysed using two-way analysis of variance. Significance was set at p < 0.05.

Results

PED reduced food intake in ad libitum-fed mice (1.052 ± 0.1293 g/day; diet effect: p = 0.0189) and was associated with lower body weight gain in this condition (1.638 ± 0.4162% from baseline; diet effect: p = 0.0017). Across feeding conditions, PED lowered basal glycaemia (16.9 ± 4.65mg/dL; diet effect: p = 0.0010), with no interaction with food restriction. Food restriction induced an expected AN-like phenotype, including progressive body weight loss and changes in wheel activity. In the open field, food restriction under standard diet reduced locomotor activity during the first minutes of exploration (1661 ± 342.7 cm; p = 0.0417), whereas PED exposure restored early locomotor exploration to ad libitum levels. PED-exposed mice also showed a slight increase in centre exploration, suggesting reduced anxiety-like behaviour. In the elevated plus maze, PED increased time spent in the open arms under restrictive conditions (29.74 ± 9.972 s; p = 0.0354), supporting an anxiolytic-like effect. In the preference test, food-restricted mice previously exposed to PED displayed the highest relative preference for PED among all groups, with a significant feeding condition × diet interaction (p = 0.0006). However, actual PED intake remained modest, accounting for less than 15% of total food consumption.

Conclusions

Protein enrichment influences metabolic and behavioural adaptations in an AN-like mouse model. PED lowers food intake and basal glycaemia, modulates later food preference and may attenuate restriction-induced hypoactivity and anxiety-like responses. These effects depend on energetic state and diet history, suggesting that protein preference may reflect a compensatory, but not uniformly protective, behavioural-metabolic strategy. Further work should clarify the neurobiological mechanisms linking protein intake, endogenous glucose regulation, energy balance and emotional behaviour, with attention to hypothalamic, melanocortinergic and reward-related circuits.

References

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