Ketogenic diet improves behaviors in a maternal immune activation model of autism spectrum disorder

Abstract

Prenatal factors influence autism spectrum disorder (ASD) incidence in children and can increase ASD symptoms in offspring of animal models. These may include maternal immune activation (MIA) due to viral or bacterial infection during the first trimesters. Unfortunately, regardless of ASD etiology, existing drugs are poorly effective against core symptoms. For nearly a century a ketogenic diet (KD) has been used to treat seizures, and recent insights into mechanisms of ASD and a growing recognition that immune/inflammatory conditions exacerbate ASD risk has increased interest in KD as a treatment for ASD. Here we studied the effects of KD on core ASD symptoms in offspring exposed to MIA. To produce MIA, pregnant C57Bl/6 mice were injected with the viral mimic polyinosinic-polycytidylic acid; after weaning offspring were fed KD or control diet for three weeks. Consistent with an ASD phenotype of a higher incidence in males, control diet-fed MIA male offspring were not social and exhibited high levels of repetitive self-directed behaviors; female offspring were unaffected. However, KD feeding partially or completely reversed all MIA-induced behavioral abnormalities in males; it had no effect on behavior in females. KD-induced metabolic changes of reduced blood glucose and elevated blood ketones were quantified in offspring of both sexes. Prior work from our laboratory and others demonstrate KDs improve relevant behaviors in several ASD models, and here we demonstrate clear benefits of KD in the MIA model of ASD. Together these studies suggest a broad utility for metabolic therapy in improving core ASD symptoms, and support further research to develop and apply ketogenic and/or metabolic strategies in patients with ASD.

Fig 1. Hallmarks of KD therapy confirmed in MIA mice.

Left: Blood β-hydroxybutyrate was elevated by KD feeding. Middle: Blood glucose was reduced by KD feeding. ***p<0.001 compared to CD-fed control offspring; n = 20–23. There were no sex differences in either measure (in initial analysis, sex-by-treatment interaction: F = 1.7, p = 0.19 for ketones; F = 1.8, p = 0.18 for glucose); male and female data are combined. Right: CD-fed female mice gained significant weight during diet treatment; KD-fed mice did not. §§§p<0.001 compared to baseline; n = 11–12. Available data from male mice showed a similar pattern (S1 File).

Fig 2. KD effects on social behavior in male and female MIA offspring.

Top panels: male mice. Top left: Chamber time in the three-chamber test. CD-fed MIA offspring were not social in phase 2. This effect was reversed by KD feeding. Control offspring were social, as expected. Mice in all treatment groups showed significant preference for social novelty in phase 3. Top right: Social contact in the three-chamber test. Social contact time was decreased in male MIA offspring, and elevated by KD feeding to levels above control offspring. §p<0.05, §§p<0.01, §§§p<0.001 compared to phase 1 within the same treatment group. *p<0.05 compared to control mice. ###p<0.001 compared to CD-fed MIA offspring; n = 8–12. Bottom panels: female mice. Bottom left: Sociability expressed as time in chamber with a mouse was not impaired by MIA treatment and not affected by KD feeding. §§§p<0.001 overall phase 2 compared to phase 1; n = 10–12. Bottom right: Sociability expressed as social contact was not impaired by MIA treatment; however KD-feeding elevated social contact. ***p<0.001 overall compared to CD-fed control and MIA offspring; n = 11 all groups.

Fig 3. Social transmission of food preference.

There was no MIA-induced impairment in this task: all treatment groups learned the social transmission of a safe food flavor; n = 11–13. There were no sex differences in this behavior (in initial analysis, sex-by-treatment interaction: F = 0.8, p = 0.45).

Fig 4. Effects of KD on self-directed repetitive grooming behavior.

Top panels: male mice. Top left: Self-directed grooming behavior in the three-chamber test. Grooming was elevated by MIA treatment, and this effect was reversed completely by KD feeding. Post hoc tests were restricted to within-phase comparisons; n = 8–10. Top right: Self-directed behavior in the single chamber test. Grooming was elevated by MIA treatment, and this effect was partially reversed by KD feeding. **p<0.01, ***p<0.001 compared to CD-fed control offspring. ##p<0.01 compared to CD-fed MIA mice; n = 8–10. Bottom panels: female mice. Bottom left: Grooming in the three-chamber test was not elevated by MIA treatment in females; it was, however, lowered by KD feeding; n = 11–12. Bottom right: Grooming in the one-chamber test was not affected by MIA or KD in females. *p<0.05, ***p<0.001 compared to CD-fed control mice.