Gut brain axis and microbiota

Editor's Notes

• #3: The concept of targeting the body rather than brain for treating psychiatric and neurological conditions may seem outlandish even now. Over the last decade it has received much attention. The microbiota inhabiting our bodies outnumbers our human cells by approximately 1.3:1 and comprises a vast ecology of bacteria, yeasts, and parasites such as helminths, viruses, and protozoa. It is becoming more and more apparent that biologists must include the impact of our microbiota on human physiology, including PK and PD following drug administration. The exact composition of the gut microbiota present in each individual is different, and it is still unclear what definitively constitutes a healthy adult gut microbial profile other than one that exhibits both diversity and stability. They have several important roles in digestion, including fermentation of undigested carbohydrates, production of vitamins and secondary bile acids and immune system development.
• #5: The presence of bidirectional gut-brain communication may seem obvious with respect to the central control of hunger and satiety signals and factors originating in the gut that regulate food intake in the short term. Changes in behavior are associated with both GI illness and functional GI disorders (disorders in which there is no overt GI pathology present but which have a high psychological component) and can result in severe GI discomfort. Colonization of the gut is considered by most researchers in the field to primarily occur at birth, and the microbiome seeded in the infant initially resembles that of the maternal vagina.
• #6: CS encompassing microbes present on the skin and from the delivery suite. The gut microbiome profile of C-section delivered infants becomes closer to that of vaginally delivered infants within the first two years of life; Diet plays a huge role in the composition and diversity of the gut microbiome, and alterations in diet can cause rapid changes in gut microbial profiles. Consumption of a Mediterranean diet is associated with both a specific gut microbial profile and a host of beneficial health effects. Detrimental effects of poor diet, including Western-style diets, upon the gut microbiome have been linked with the rise in the prevalence of metabolic syndromes.
• #8: Current evidence indicates that bottom-up modulation of the CNS by the microbiome occurs primarily through neuroimmune and neuroendocrine mechanisms, often involving the vagus nerve. This communication is mediated by several microbially derived molecules that include short-chain fatty acids (SCFAs), secondary bile acids (2BAs), and tryptophan metabolites. These molecules propagate signals primarily through interaction with enteroendocrine cells (EECs), enterochromaffin cells (ECCs), and the mucosal immune system, but some cross the intestinal barrier, enter systemic circulation, and may cross the blood-brain barrier.  In addition to generating these metabolites that activate endogenous CNS signaling mechanisms, the microbiota can independently produce or contribute to the production of a number of neuroactive molecules including but not limited to γ-aminobutyric acid, 5-HT, norepinephrine, and dopamine, although it is unknown if they reach relevant receptors or achieve sufficient levels to elicit a host response.
• #11: An important pathway by which gut microbes and their metabolites communicate with the CNS involves the cells making up the endocrine system of the gut. There are at least 12 different types of these cells with several subtypes (in particular A, K, and L cells) present as subgroups along the intestine that contain different combinations of molecules.  Released in response to chemical and or mechanical stimuli, these molecules can enter the systemic circulation and reach centers in the CNS involved in ingestive behavior (including the nucleus tractus solitarius and the hypothalamus) or act locally and activate closely adjacent afferent vagal terminals in the gut or liver to generate brain signals. A series of receptors involved in the regulation of satiety and hunger have been identified on these cells, which are activated by microbial metabolites including bile acids and SCFAs.
• #12: Although bile acids are endogenous molecules synthesized from cholesterol in the liver, the size and composition of the host’s pool of these molecules is heavily influenced by dietary intake, especially of fat,79 and the downstream metabolism by the gut microbiota. Ileal expression of farnesoid X receptor (FXR), a nuclear receptor, is activated by bile acids leading to production of fibroblast growth factor 19 (FGF19), which can enter the systemic circulation and cross the blood-brain barrier. Some intestinal L cells express surface receptor G protein-coupled bile acid receptor (TGR5), which is activated mostly by secondary bile acids, which are strongly influenced by microbial activity. TGR5 signaling controls glucose homeostasis by mechanisms including increased glucagon-like peptide-1 (GLP-1) release by these L cells. SCFAs have been implicated as major signaling molecules mediating host-microbe communication via EECs and ECCs. These molecules are generated by microbial fermentation of host dietary-resistant starch and nonstarch polysaccharides and serve an important part in the host’s energy harvest while also stimulating colonic blood flow, fluid and electrolyte uptake, and mucosal proliferation. Acetate, butyrate, and propionate modulate the expression and secretion of GLP-1 via free fatty acid receptor 2 (FFAR2) and FFAR3 on L cells.
• #13: Bidirectional brain-gut-microbiome interactions related to serotonin signaling. Enterochromaffin cells (shown in green) contain more than 90% of the body’s serotonin (5-HT). 5-HT synthesis in ECCs is modulated by SCFAs and 2BAs produced by spore-forming Clostridiales, which increase their stimulatory actions on ECCs with increased dietary tryptophan availability.68 ECCs communicate with afferent nerve fibers through synapse-like connections between neuropod-like extensions and afferent nerve terminals.192 The autonomic nervous system can activate ECCs to release 5-HT into the gut lumen, where it can interact with gut microbes.155 TPH1, tryptophan hydroxylase type 1.
• #14: It is important to highlight that the gut microbiota influence the development and function of the CNS resident immune cells, especially microglia. Relative to SPF mice, GF mice have compromised microglia maturation and morphology, leading to weakened early responses to pathogen exposure. This phenotype can be normalized by postnatal SCFA supplementation or colonization with a complex microbial community. Antibiotic treatment to eradicate bacteria in SPF adult mice leads microglia to regain immature status, which then can be normalized by recolonization with complex microbiota, suggesting that active microbial signaling is required throughout adulthood to preserve microglial maturation.
• #15: Most evidence to date relies on vagal receptors that sense regulatory gut peptides, inflammatory molecules, dietary components, and bacterial metabolites to relay signals to the CNS. Toll-like receptors 3 and 7, which recognize viral RNA, and Toll-like receptors 2 and 4, which recognize peptidoglycan and lipopolysaccharide,
• #17: Both branches of ANS regulate gut functions including regional motility, secretion of gastric acid, mucus, bicarbonate, gut peptides, antimicrobial peptides, epithelial fluid maintenance, intestinal permeability, and mucosal immune response. These ANS-induced changes in gut physiology affect the microbial habitat, thereby modulating microbiota composition and activity.  The frequency of migrating motor complexes, which play a crucial role in intestinal transit during the fasting state, is influenced by food intake patterns, sleep quality, and stress. Relatively rapid flow in the small intestine inhibits permanent colonization of the upper gut. Intestinal transit time assessed by the Bristol Stool Scale which strongly correlates with microbial richness and composition.
• #18:  Increased intestinal permeability and susceptibility to experimental inflammation observed in mouse models of depressive behavior. The ANS modulates the secretion of mucus by intestinal goblet cells, affecting the thickness and quality of the intestinal mucus layer. Psychological stress leads to a less-protective mucus layer via catecholamine signaling, which alters the composition and size of secreted mucus. Mouse model of brain injury - altered mucoprotein production - Changes in microbiota composition.
• #19: Host neuroendocrine system can communicate with the microbiota more directly via intraluminal release of host signaling molecules, including but not limited to catecholamines, 5-HT, dynorphin, and cytokines, from neurons, immune cells, and ECCs.  Epinephrine and norepinephrine are shown to increase the virulence properties of several enteric pathogens as well as nonpathogenic microbes via stimulation of native quorum-sensing mechanisms. The gut microbiota show circadian rhythmicity in both abundance and expression in a manner dependent on the host and its behavior, especially feeding timing, and simulated jet-lag shifts composition, enhancing dysbiosis.
• #21: Stress plays a significant role in altering the microbiota-gut-brain axis. There is an abundance of preclinical evidence describing stress-induced alterations in the microbiota-gut-brain axis in several hosts, including rodents and nonhuman primates. Absence of a gut microbiota in germfree animals results in stress responses. In humans, there is a paucity of studies regarding specific stress-related alterations in the gut microbial profile. However, numerous studies have described an altered gut microbial profile in irritable bowel syndrome (IBS). Exposure to stress and anxiety through adverse life events can also impact stress-related depressive states, which are also currently under investigation with regard to gut microbiota profile. Acute stress – Appropriate HPA axis activation in order to respond appropriately to the stressor; however, prolonged, chronic stress can cause heightened inappropriate HPA axis activation. There is more than 40 years worth of literature showing the effect of stress on the gut microbiome. Exposure to social stressors for 2 hours can change community profiles and maternal prenatal stress is associated with an altered infant microbiome potentiating increased inflammation.
• #22: Preclinical studies have shown the microbiota capacity to modulate emotional behaviors, and influence parameters significant to depression pathogenesis and severity. Coprococcus and Dialister species were not present in the depressed cohorts, and their presence in the nondepressed cohort was positively associated with quality of life scores.
• #23: Although the clinical hallmarks of Parkinson's disease remain motor deficits, gastrointestinal symptoms (in addition to other nonmotor symptoms) are present that contribute more detrimentally to patient quality of life. Nonmotor symptoms - Dysfunctional autonomic and enteric nervous systems (slow-transit constipation and sensory alterations).  There is a significant comorbidity of Parkinson's disease and IBS-like symptoms. Constipation (earliest features) appears as early as 15.3 years before motor dysfunction. FMT from PD patients into mouse models of PD resulted in both motor deficits and neuroinflammation, providing further evidence that the gut microbiota is linked with PD. Epidemiological studies - Full truncal vagotomy for treatment of peptic ulcer disease have a diminished risk of PD as they age and provide some insight into the possible microbiota-gut-brain signaling involved.
• #24: Several studies have described alterations in the gut microbial profile in children with ASD, and they particularly highlighted a reduced abundance of Bifidobacterium (beneficial genus). Studies described an increased abundance of Clostridia (pathogenic genus). Treatment with vancomycin, a broad-spectrum antibiotic, resulted in positive alterations in behavioural symptoms in one short-term study, further highlighting the role of an aberrantly altered gut microbial profile in ASD. FMT from healthy individuals into ASD patients resulted in improvements in both gut comorbidities and behaviours in the cohort. Preliminary evidence that an altered gut microbial profile also exists in PTSD patients.
• #25: There are limited studies on the impact of gut microbiota in the schizophrenia, but schizophrenia does have a component of gut comorbidities.
• #26: A dysregulation of hedonic feeding behavior (food addiction) plays an important role in the current obesity epidemic. Gut microbiome produces several neuroactive compounds (indole-containing metabolites and 5-HT). Dramatic change in gut microbial composition after bariatric surgery. fecal transplantation from subjects after bariatric surgery was able to transmit the weight loss effects of bariatric surgery to a germ-free nonoperated recipient, inducing weight loss and reduced food intake.
• #27: Significant microbial shifts in fecal microbial community composition between healthy controls and IBS patients, based on disease subtypes (diarrhea-predominant IBS, constipation-predominant IBS, and IBS mixed type), age. Dysbiotic IBS subgroup also differed in regional brain volumes from the eubiotic group, suggesting a relationship between microbial community structure and brain structure.  Luminal aspirates - Decreased diversity in small-bowel microbiota with increased abundance of gram-negative organisms in IBS.
• #28: Emerging evidence - Gut microbiota is now being targeted to mediate beneficial brain effects. Expanded definition- “any exogenous influence whose effect on the brain is bacterially-mediated”. It is notable that the first antidepressants on the market were also antibiotics.
• #29: Probiotics were first described over 100 years ago in the work of Elie Metchnikoff, who described health benefits associated with the consumption of lactic acid bacteria in fermented milk in a population of poor Bulgarians. There is now a plethora of studies showing benefits of a variety of probiotic strains in abrogating stress, depression, and anxiety-like behaviours in preclinical models. Health benefits of putative probiotics are entirely strain-dependent, and not all probiotics have brain-beneficial effects. Strains of Lactobacillus and Bifidobacterium, which have well-defined safety profiles, are frequently used as probiotics, both genera generally being found to have beneficial health effects, but the individual strain differences are paramount in determining efficacy. Probiotics generally do not become resident in the gut, that is, probiotic formulations require daily consumption to maintain positive effects. Next-generation probiotics, including probiotics formally recognized as live biotherapeutics in the United States, which may be delivered under more traditional pharmaceutical routes rather than through food delivery and are associated with a more stringent regulatory framework.
• #30: An RCT of an L. rhamnosus strain significantly lowered measures of anxiety and postnatal depression when taken during pregnancy. A recent study further described multiple benefits of L. plantarum in stressed but otherwise healthy adults, including a reduction of stress and anxiety measures and improvements in multiple aspects of memory and cognition, enhanced serotonergic signalling, and reductions in cortisol and proinflammatory cytokines. Interestingly, probiotics have also been studied as a potential adjunct therapy in combination with antidepressant drugs in a cohort of treatment-resistant depressions, showing some promise in an open-label study.
• #31: Healthy medical students had reduced levels of stress related to university examinations following consumption of a fermented milk containing the probiotic L. casei Shirota. This strain was previously shown to reduce anxiety symptoms in patients with chronic fatigue syndrome. Increases in salivary cortisol observed during an exam stress period were also diminished in a healthy student population consuming an L. plantarum 299v probiotic strain. Cognitive performance (attention and verbal learning) was improved in a recent study of an MDD patient cohort due to supplementation with L. plantarum. A 6-week intervention with a B. longum strain in IBS patients resulted in an improvement in depression, but not anxiety scores. L. helveticus and B. longum have also been described to benefit anxiety and stress responses, and an L. acidophilus and B. longum combination reduced stress-associated GI symptoms in stress-affected individuals.
• #32: One such RCT, using a multistrain probiotic in a milk drink (L. acidophilus, L. casei, B. bifidum, and Lactobacillus fermentum species) resulted in improvements in Mini–Mental State Exam scores. However, no improvements in cognition were observed in a severe AD cohort in another RCT by the same group, using other Lactobacilli and Bifidobacterium strains, again highlighting the differential effects of the strain level in formulating effective probiotics as well as emphasizing the need for clarity with regard to the best time to therapeutically intervene in progressive disease. Cognition was also improved following a 6-month intervention with a multistrain probiotic in a cohort of HIV patients. Improved emotional memory performance - 4-week intervention
• #33: Certain probiotic strains have shown promise in ameliorating behavioral and neurochemical perturbations in preclinical models of ASD. Use of probiotics in ASD is in its infancy and is focused on improvement in gut-related comorbidities, though there is also some preliminary evidence for behavioural change. Improvements to peripheral inflammatory profiles - chronic schizophrenia patients. A recent open-label study on Bifidobacterium breve supplementation in schizophrenic patients reported a beneficial effect on anxiety and depression scores.
• #34: Modulation of gut microbiota by the introduction of living microorganisms faces many technical and pragmatic difficulties. These include ensuring the survival of the probiotic transiting through the hostile environment of altered pH levels (acidity of the stomach to the alkali bile salt environment), location in the intestines where it can exert its potential therapeutic effect; the ease of fermentation and issues in continuity and consistency in batch production; the inability of probiotics to colonize the environment in the long term; and the selection of appropriate strains. Prebiotics consist of fibers such as inulin, fructo-oligosaccharides, galacto-oligosaccharides (GOSs), and resistant starch, which fail to be absorbed in the small intestine and are selectively fermented by gut microbes. Prebiotics are found in a wide variety of fruits, vegetables, and grains, entities that are increasingly diminished in the Western-style diet. Prebiotics are also found in human milk (human milk oligosaccharides). A B-GOS intervention - behavioural improvements in ASD when included with a restrictive diet intervention. IBS patients - Significant decrease in anxiety scores. ASD patients - Behavioural improvements Schizophrenia or bipolar disorder patients - Variable alterations in gut microbiome profiles
• #35: Synbiotics are a combination of probiotics and prebiotics, whereby the prebiotics improve the viability of the probiotic, providing a source of fermentable fiber as well as acting as a general prebiotic. There is limited evidence for PTSD, with one study showing positive results of a fermented soy drink in a PTSD cohort. A recent pilot study - synbiotic (Bifidobacterium infantis plus oligosaccharides) - ameliorating some of the gut-related comorbidities related to ASD.
• #36: There are limited studies in humans regarding the administration of bioactives and their effect on brain health; however, the general impacts of the gut microbiome and SCFAs on metabolic traits and disease have recently been described. In one human interventional study of an inulin-proprionate ester that increases proprionate production in the colon, brain imaging (via functional MRI) showed a reduction in anticipatory reward responses in the striatum due to the supplement. It is therefore plausible that behaviors can be targeted through supplementation with such bioactives. Gut peptides have a well-established role in influencing behavior and also a role in stress, anxiety, and depression.
• #37: Paraprobiotics, or nonviable probiotics, e.g., heat-killed probiotics, can also be included in the category of postbiotics in that they contain structural components that may exert biological activity in the host, and there are now a number of human studies on the microbiota-gut-brain axis regarding supplementation with paraprobiotics. Stress experienced due to university exams in medical students was alleviated by a 12-week intervention with a heat-killed, washed, paraprobiotic CP2305. Heat-killed Lactobacillus paracasei PS23 having benefits in a corticosterone-induced depressive phenotype, reversing hippocampal and prefrontal cortex reductions in dopamine levels. Distinct marketing advantage - increased shelf life and possibly in terms of an increased safety profile. However, not all probiotic strains will be effective as heat-killed preparations.
• #38: Fermented foods contain probiotics, prebiotics and bacterially derived bioactives. Two of the most common strains used in the fermentation process include Lactobacillus delbreuckii subsp. bulgaricus and S. thermophilus. Studies using fermented food interventions in humans are limited. Fermented milk drinks have been found to result in positive benefits in emotional processing and stress. Mediterranean diets (rich in psychobiotics) have mental health benefits. A meta-analysis of 22 studies also showed decreased risk of depression and cognitive impairment, including risk of having AD, with increased adherence to a Mediterranean diet. SMILES trial - clear benefit in moderate to severe depression, with an economic evaluation indicating this could potentially be a cost-effective option for wider implementation.
• #39: Antibiotic treatment clearly has profound effects on the gut microbiome. A study on T2DM determined that metformin was responsible for much of the alterations in the gut microbiota profiles of T2DM patients, which had previously been reported to be due to the disease.
• #40: Fresh fecal sampling, in conjunction with fresh plating of resultant bacteria in a specific agar medium, and picking resultant colonies is the first step. In the European Union, in order to meet the European Food Safety Authority’s Qualified Presumption of Safety, the group of microorganisms must meet certain criteria, including the identification of their taxonomy, establishment of their safety, exclusion of pathogenic properties, and clear definition of their intended usage. Once the strains have been chosen, they can be used in a battery of assays, such as enzyme assays, in addition to in vitro cell testing. Depending on the cell type of interest, the cells can be monitored for growth; morphology; and the production of anti-inflammatory cytokines and other bioactives such as SCFAs, neurotransmitters, and hormones. GPCRs within the gut can be directly targeted by microbially produced bioactives, and classical pharmacological approaches can be used to determine microbial bioactive effects on GPCRs. If a candidate probiotic shows particular benefits with regard to any of these factors, the probiotic can then be used in preclinical in vivo testing, and further safety analysis can be performed.
• #45: We must now additionally consider that the gut microbiota, particularly in the case of drugs taken orally, can both impact the metabolism of the drugs and be a crucial effector/mediator of drug response. In addition to possessing poisonous qualities in its own right when present in unfavorable configurations, or in the case of invading pathogenic species, it is also possible that we could directly target the gut microbiota to treat an increasing number of diseases and that the gut microbiota represents an exciting reservoir of new therapeutic opportunities. We focus on targeting bacterial gut microbiota in order to impact microbiota-gut-brain axis signaling for the effective treatment of central nervous system (CNS) disorders.