The characterization of gut microbiome a decade ago has added a long-overlooked aspect to the complex bidirectional signalling between brain and gut [
1]. This interaction, known as ‘gut–brain axis’ has been shown to link the cognitive and emotional centres of brain with the intestinal functions [
2]. The gut microbiota plays a prominent role in these interactions by regulating behaviour and brain processes, that is, stress responsivity [
3], anxiety-related behaviours [
4], pain perception [
5], and social cognition [
6], as shown by intriguing experimental investigations in rodents. In addition to the emotional processing, gut microbiota have also been shown to play an important role in modulating brain biochemistry and brain plasticity. For example, Hoban and colleagues showed that the gut microbiota regulates the expression of genes linked to myelination and myelin plasticity in prefrontal cortex [
7]. On a similar note, a role of gut microbiota in altering the central GABA (gamma amino butyric acid) receptor expression has also been demonstrated [
8]. Although most of the evidence for an influence of gut microbiota on brain and behaviour is based on our understanding of rodent studies, initial studies in humans seem to support the notion that there exists a similar relationship between our gut microbes and brain and behaviour. For instance, consumption of
Lactobacillus and
Bifidobacterium strains by healthy volunteers was found to influence the scores of stress and anxiety-related questionnaires [
9‐
11]. However, as the assessment was based on self-reported measures in all these studies, caution is warranted when drawing firm conclusions. Furthermore, some recent studies have employed neuroimaging techniques such as task-based functional MRI and resting-state fMRI to better understand the physiological pathways involved in gut–brain communications and their influence on brain function [
12,
13]. A task-based fMRI study conducted in our group [
12] demonstrated that a 4-week multi-strain probiotic administration influences brain activation patterns associated with emotional decision-making and recognition memory tasks in healthy volunteers. Another fMRI study by Tillisch and colleagues showed that the ingestion of
Lactobacillus and
Bifidobacterium species for 4 weeks by healthy women altered the brain activity in insula, somatosensory cortex and periaqueductal gray brain regions in response to an emotional attention fMRI task [
13]. This study also investigated the corresponding functional connectivity (FC) changes in these regions using region of interest (ROI) analysis and reported changes in FC in midbrain regions. However, the influence of probiotic administration on whole-brain functional connectivity remains unclear. Furthermore, even when there is a considerable volume of preclinical literature indicating an influence of gut microbiome on brain structure, our understanding in human subjects in this context is limited to the observations in patients with irritable bowel syndrome (IBS) [
14] and this is far from complete.
Numerous neuroimaging studies have revealed a strong relationship between structural integrity and functional connectivity (see review by Damoiseaux and Greicius [
15]). Functional connectivity is most commonly calculated from resting-state fMRI and examines the similarities between spontaneous fluctuations that occur over time in distal grey matter regions [
16] and diffusion MRI measures the structural integrity [
17]. Considering the existing literature on the influence of probiotic administration on functional connectivity [
13], a further investigation of the structural basis for these functional interactions would add valuable insights to our current understanding of the gut–brain interaction mechanisms.
In this study, we aimed at investigating the influence of a 4-week multi-strain probiotic administration on whole-brain functional connectivity in healthy volunteers. We hypothesized that the manipulation of gut microbiota by multi-strain probiotic ingestion will influence functional connectivity in the resting-state networks (RSNs) mediating emotional and higher order cognitive functions. We suspect that salience network, executive network and default mode network are of particular interest, considering their role in mediating these processes. Furthermore, we also hypothesized that the probiotic intervention will influence the underlying white matter architecture associated with the functional connectivity networks. To test these hypotheses, we performed resting-state fMRI and diffusion MRI scanning at two time points: at baseline (time point 1) and after 4 weeks (time point 2).