Gut microbiome in ADHD and its relation to neural reward anticipation

Abstract

Background: Microorganisms in the human intestine (i.e. the gut microbiome) have an increasingly recognized impact on human health, including brain functioning. Attention-deficit/hyperactivity disorder (ADHD) is a neurodevelopmental disorder associated with abnormalities in dopamine neurotransmission and deficits in reward processing and its underlying neuro-circuitry including the ventral striatum. The microbiome might contribute to ADHD etiology via the gut-brain axis. In this pilot study, we investigated potential differences in the microbiome between ADHD cases and undiagnosed controls, as well as its relation to neural reward processing.

Methods: We used 16S rRNA marker gene sequencing (16S) to identify bacterial taxa and their predicted gene functions in 19 ADHD and 77 control participants. Using functional magnetic resonance imaging (fMRI), we interrogated the effect of observed microbiome differences in neural reward responses in a subset of 28 participants, independent of diagnosis.

Results: For the first time, we describe gut microbial makeup of adolescents and adults diagnosed with ADHD. We found that the relative abundance of several bacterial taxa differed between cases and controls, albeit marginally significant. A nominal increase in the Bifidobacterium genus was observed in ADHD cases. In a hypothesis-driven approach, we found that the observed increase was linked to significantly enhanced 16S-based predicted bacterial gene functionality encoding cyclohexadienyl dehydratase in cases relative to controls. This enzyme is involved in the synthesis of phenylalanine, a precursor of dopamine. Increased relative abundance of this functionality was significantly associated with decreased ventral striatal fMRI responses during reward anticipation, independent of ADHD diagnosis and age.

Conclusions: Our results show increases in gut microbiome predicted function of dopamine precursor synthesis between ADHD cases and controls. This increase in microbiome function relates to decreased neural responses to reward anticipation. Decreased neural reward anticipation constitutes one of the hallmarks of ADHD.

Fig 1. Potential routes in which precursors of monoamines could influence brain functioning.

The large neutral amino acids tryptophan, phenylalanine, and tyrosine, which are absorbed in the intestine [20], are precursors of monoamines. Tryptophan and phenylalanine are essential amino acids, meaning that they cannot be synthesized by the human body itself [21]. 5-HTP = 5-Hydroxytryptophan.

Fig 2. Microbiome sample, fMRI sample, and their overlap.

Fig 3. The strongest differentially abundant microbial taxa for ADHD cases (n = 19) versus healthy controls (n = 77), shown in the graphical Cytoscape visualization [32].

Nodes represent taxa (node size represents average relative abundance, for both experimental groups combined), edges (dashed lines) link the different taxonomic levels. The weighed fold-change (node color) is calculated as the 2log of the ratio of the relative abundance between control and ADHD (0 = no difference between genotypes, 1 = twice as abundant in control, etcetera). In other words: yellow to red indicates an overrepresentation in control, hence an underrepresentation in ADHD, and vice versa for light- to dark blue. The significance (node border width) is expressed as the p-value of a Mann–Whitney U test, uncorrected for multiple comparisons.

Fig 4. The ADHD microbiome contains significantly increased levels of predicted cyclohexadienyl dehydratase (CDT; KEGG Ortholog K01713; EC:4.2.1.51), responsible for phenylalanine synthesis (Fig B in S1 Appendix).

This analysis is based on functional predictions deriving from 16S profiles of the microbiome, as performed by PICRUSt [33]. Box plots represent the relative abundance of predicted CDT, with 5–95% percentile whiskers (dots represent outliers). The significance was tested with a non-parametric MWU (* p = 0.038), Bonferroni-corrected for 15 K numbers identified.

Fig 5. fMRI results.

A. Main effect of reward anticipation, cluster-level corrected at the whole-brain level (pFWE < 0.05). Color bars reflect T-values. B. Diagnosis effects in the anatomical region of interest (ROI) of the ventral striatum. C. Negative correlation of the microbiome function CDT (see Fig 4) with reward anticipation responses across the whole-brain (n = 28), intensity threshold at p < 0.001 uncorrected (T = 3.45). The clusters in bilateral ventral striatum (x = -11, y = 11, z = -9, cluster size = 8, p(FWE, cluster) = 0.024; x = 11, y = 6, z = -11, cluster size = 2, p(FWE, cluster) = 0.036) are significant after correcting for multiple comparisons across the search volume (cluster-level pFWE < 0.05, SVC), i.e. the anatomically defined ventral striatum shown in panel B. SVC = small volume correction. * indicates p < 0.05.