Psychiatry - Doctoral Theses
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Item Nutritional psychiatry, stress and the microbiota-gut-brain axis: focus on dietary fiber(University College Cork, 2024) Lachmansingh, David Antoine; Clarke, Gerard; Lavelle, Aonghus; Cryan, JohnDietary fiber has an assortment of health benefits and the gut microbiota has been shown to be an integral regulator of brain function and behavior. The interface between these features may have implications from a therapeutic as well as a neurobiological perspective in terms of mental health function, especially in circumstances where stress is considered a significant risk factor for anxiety and depressive symptoms. A prodigious amount of research has helped to shape the narrative of a possible relationship between stress and the microbiota-gut-brain axis. However, the nature of the relationship between dietary fiber, stress and the microbiota-gut-brain axis needs to be explored, especially with efforts focused on eliciting and understanding probable mechanisms. We demonstrated that dietary fiber has the ability to influence gut-brain modules (GBMs) including SCFA synthesis via the gut microbiota, by devising a systematic review protocol followed by its execution and deployment across studies examining the impact of different dietary fiber and compositional alterations with publicly available datasets. These datasets were re-analyzed via consistent GBM bioinformatic pipelines with the results illustrating a high probability that dietary fiber favors an increased abundance of SCFA-producing taxa and GBMs for SCFA synthesis. We also showed via a cross-sectional analysis, that dietary fiber was associated with reduced state anxiety in healthy individuals. The combination of these results have furthered our understanding in the nutritional psychiatry field, indicating that dietary fiber may recruit SCFA production in reducing anxiety. Such findings could allow for further research focusing on SCFAs impacting anxiety. Additionally, this platform may also allow for other GBMs impacted by dietary fiber to be evaluated in the role of reducing anxiety symptoms, as well as other symptoms relating to stress and depression. Taken together, our findings have confirmed that the microbiota-gut-brain axis and dietary fiber are implicated in the way we feel and our mental health.Item The role of the gut microbiota in the function and integrity of brain barriers(University College Cork, 2023) Knox, Emily G.; Cryan, John; O'Driscoll, Caitriona M.; Rodriguez Aburto, Maria; Clarke, GerardThe barriers of the brain help maintain an optimal environment for central nervous system (CNS) function and homeostasis. There are many instances such as during age related disorders and neuropsychiatric disorders in which these barriers are under threat. Importantly, the gut-microbiota can interact with the brain barriers, thereby altering barrier physiology and communication with the brain. This bidirectional communication is known as the microbiota-gut-brain axis. Modulation of the gut microbiota can thereby alter functions in the brain, potentially through influence of the brain barriers. In this thesis we focus on the interface of both brain barriers; the blood-cerebrospinal fluid barrier (CSF) and blood-brain barrier (BBB) in microbiota-gut-brain axis communication. In particular, using germ-free mice which lack a gut microbiota, we investigated the contributing role of the gut microbiota in regulation of the blood-CSF barrier. Following confocal microscopy for visualization of the tight junction protein, ZO-1, germ-free mice were found to have a disruption of the tight junction protein networks in isolated choroid plexus tissue. Similar assessment of the choroid plexus capillaries revealed no difference between germ-free and conventional mice. Since microbial metabolites are some of the known contributing factors of the gut microbiota’s manipulation of the barriers, we next investigated the effects of physiological concentrations of short chain fatty acids (SCFAs), butyrate and propionate, with and without induced barrier dysfunction (LPS) on the actin cytoskeleton and tight junction protein dynamics using an in vitro BBB model. We found that the butyrate and propionate altered filamentous actin directionality, increased tight junction protein spikes, and protected from LPS induced decrease in mitochondrial footprint and barrier integrity measured by trans-endothelial electrical resistance (TEER). While we do not yet understand the dynamics of the BBB following acute stress, we know that microbial metabolites can interact with BBB function, so we next assessed the potential for acute-stress induced metabolites to regulate BBB integrity using the same in vitro model. Following TEER assessment of a broad range of concentrations, we found that microbial metabolites from different classes; SCFAs, bile acids, and tryptophan metabolites impact BBB integrity. Specifically, butyrate, propionate, acetate, 3β-hydroxy-5-cholenic acid, and tryptamine exerted protective effects while acetate and cholic acid exerted detrimental effects at various concentrations. Lastly, since diet is a major modulator of the gut microbiota and especially in infancy, we assessed the impact of infant formulations fermented with probiotic strains of Bifidobacterium uniquely isolated from infant stool samples for potential effects on BBB integrity. Cell-free supernatants from the fermentation process which would contain a pool of microbial metabolites were then collected and exposed to the same in vitro model for assessment of barrier integrity with and without LPS disruption. We found that in each of the four infant formulations, there was protection from LPS disruption following fermentation with some of the Bifidobacterium strains. Overall, these results provide valuable insight into the interphase of brain barriers in microbiota-gut-brain axis communication. We highlight the role of another brain barrier in this interface, the blood-CSF barrier and further elucidate the microbial metabolite interaction in BBB physiology. Further, using two of the major modulators of the gut microbiota, stress and diet, we reveal that there is scope for the impacted metabolites or metabolic pools to influence brain barrier interactions.Item The neurobiological effects of naturally-derived polyphenols and phospholipids in cellular & animal models of stress(University College Cork, 2019-12-20) Donoso, Francisco; Cryan, John; Dinan, Timothy G.; Stanton, Catherine; Science Foundation Ireland; Department of Agriculture, Food and the Marine, IrelandThe molecular and cellular basis of stress neurobiology remain an important research question in clinical science. Indeed, stress-related mental disorders, including depression and anxiety, are currently a major public health concern. Thus, improving our knowledge about the pathophysiology of these neuropsychiatric disorders may enable the development of novel strategies for their treatment and prevention. On the other hand, the inefficacy of currently available therapies for various stress-related disorders, and the numerous side effects that accompany these treatments, have strengthened the search for less invasive strategies with fewer negative side effects. In this regard, the emerging and compelling evidence for nutrition as a potential therapeutic avenue for the treatment of mental disorders suggests that changes in diet are a viable strategy for improving mental health and treating stress-related psychiatric disorders. Moreover, there is considerable evidence suggesting that certain natural compounds available in diet have a therapeutic potential to improve mental health and disease. For instance, naturally occurring phytochemicals, namely polyphenols, are molecular compounds found in different plant sources, such as vegetables and fruits. Also, phospholipids are a class of lipid that comprise a major component of all cell membranes, specially concentrated in lean meat and dairy products. Both polyphenols and phospholipids have demonstrated interesting beneficial effects for human health. However, their therapeutic potential to act prophylactically against the detrimental effects of neuropsychiatric disorders have just begun to be taken seriously. Therefore, in this thesis we have tested the hypothesis that polyphenols and/or phospholipids could improve behavioural and neurobiological outcomes in cellular and animal models of stress. Further, we provide evidence that polyphenols and phospholipids exert neuroprotective effects against the cytotoxicity produced by corticosterone, the main rodent stress hormone, in cortical neurons. Specifically, we have elucidated the potential mechanisms underlying polyphenol-mediated neuroprotection in vitro, and demonstrated that phospholipid exposure positively impacts on neurodevelopmental processes, such as proliferation and differentiation of cultured neural progenitor cells. In addition, we confirmed the therapeutic potential of a dietary intervention with polyphenols by detecting its capacity to reverse depressive- and anxiety-like behaviours induced in a rat model of early-life stress. Moreover, we demonstrated potential implications to modulate BDNF-dependent recovery, regulation of the HPA axis and the microbiota-gut-brain axis in polyphenol-mediated behavioural improvement. Taken together, our findings support the therapeutic potential of polyphenols for stress-related mental disorders, and we further provide evidence for the possible mechanisms by which they may exert these effects. On the other hand, our data reveal that the novel neuromodulatory potential of phospholipids in vitro does not correlate with their inefficacy in attenuating chronic stress-induced behavioural impairment in mice. Nevertheless, these findings contribute to an exciting and growing body of research suggesting that nutritional interventions may have an important role to play in the treatment of stress-related psychiatric conditions.Item Dairy phospholipids: analysis, enrichment and biological application(University College Cork, 2017) Barry, Kate M.; Dinan, Timothy G.; Kelly, Philip; Stanton, Catherine; Department of Agriculture, Food and the MarineDairy phospholipids (PLs) are complex polar lipids with a unique amphiphilic nature underpinning their structural integrity as a membrane surrounding milk fat globules- the milk fat globule membrane (MFGM). The techno-functional capabilities, nutritional benefits and putative health associations of dairy PLs have garnered considerable interest over the last number of decades and thus significant research has focused on the analysis, isolation and application of these bioactive and functional compounds. Development of a robust and reliable analytical method for efficient recovery and accurate quantitative determination of dairy PLs was achieved in the course of comparing different lipid extraction protocols and optimisation of high performance liquid chromatography (HPLC) procedures. Reappraisal of current lipid extraction procedures based on PL recovery determined Folch as the most effective method for PL analysis with a 1.9-fold and 2.5-fold increase in PL recovery compared with Röse Gottlieb and a modified Folch, respectively. Concomitantly, optimisation of a HPLC method coupled to a charged aerosol detection system (CAD) yielded a two-fold increase in PL recovery compared to the original method. Alteration of the HPLC elution program maximised PL peak separation, and pH adjustment of the buffer reduced co-elution of PL species detected by the CAD with the result that the combined protocol yielded greater PL recoveries than that previously published, 2.30 ± 0.03 % total PL in milk. Efficiency and improved recoveries was also reflected in PL values obtained from different dairy streams, 35.32 ± 0.01 % total PL and 46.09 ± 0.01% total PL in buttermilk and butter serum, respectively. Most notably recoveries of the more acidic PLs, phosphatidylinositol (PI) and phosphatidylserine (PS) increased by 2 % and 7 % across all dairy streams analysed compared to previous studies. Generation of a novel dairy ingredient enriched in dairy PLs was achieved through development of a process at laboratory scale that combined enzymatic digestion and ultrafiltration (UF). Buttermilk with its higher residue of PL arising from buttermaking was prepared by reconstitution of buttermilk powder (BMP) substrate and subjected to extensive hydrolysis of its inherent milk protein complement followed by ultrafiltration to permeate the resulting smaller molecular weight peptide material. Screening of a number of digestive enzymes based on their proteolytic activity identified Alcalase® as the most proteolytic enzyme that generated a hydrolysate with 89.84 % of the peptides < 50 kDa in size. Comparison of 100 kDa and 50 kDa molecular weight cut off (MWCO) membranes (Sartorius Vivaflow™ 200 PES crossflow cassette) resulted in no significant difference in the peptide profiles of the obtained retentates. A 7.8-fold increase in PL (based on 6.16 ± 0.02 % and 0.79 ± 0.01 % total PL in the freeze-dried retentate and BMP starting material, respectively) was achieved based on degree of hydrolysis (DH) of 19 % and 50 kDa MWCO UF membrane filtration. This combined approach increased the lipid material 6.3-fold and reduced residual protein content 2-fold in the retentate compared to the starting BMP with no evidence of PLs detectable in the permeate, 0.00 ± 0.01 % total PL. Successful scalability of this combined process was achieved at pilot scale with an even higher 8.5-fold increase in PL material in the retentate compared to the starting material, 11.05 ± 0.02 % and 1.30 ± 0.00 % total PL, respectively. A total lipid increase of 8.7-fold and total residual protein decrease of 2.9-fold during scale-up in the pilot plant exceeded laboratory scale performance. Subsequent application of supercritical fluid extraction (SFE) treatment to the obtained PL enriched spray dried retentate yielded an enriched PL extract, 56.24 ± 0.07 % total PL (dry matter) depleted of protein and lactose, thus exceeding the dairy PL enrichment values achieved by previous researchers. Application of the highly-enriched PL extract in vitro determined a neurotrophic bioactivity. The PL extract induced a stimulatory effect on the outgrowth of cortical neuron cells. Much of the research published to date has explored the biological effects of long-chain polyunsaturated fatty acids (LC-PUFAs) strongly associated with PLs, e.g. docosahexaenoic acid (DHA). Investigation of the effect of the PL extract at different dosages determined that at a concentration of 150 µg mL-1 PL extract a 43 % increase in cortical neuron stimulation was observed compared to the control, 0 % stimulation. However, at a concentration of 300 µg mL-1 PL extract the observed stimulation decreased to 12 % suggesting a neurotoxic effect at higher dosage levels. This increase is greater than that previously published and the effective concentration, 150 µg mL-1, is in agreement with the PL levels found in human milk. This study demonstrated that dairy PLs promote the development and outgrowth of cortical neurons and open up an avenue to explore further the putative health benefits associated with dairy PLs.Item An examination of the impact of dietary lipids on behaviour and neurochemistry(University College Cork, 2015) Pusceddu, Matteo M.; Dinan, Ted G.; Cryan, John F.The molecular and cellular basis of stress pathology remains an important research question in biological science. A better understanding of this may enable the development of novel approaches for the treatment of stress-related disorders. There is a considerable body of scientific evidence suggesting that dietary lipids, phospholipids and omega-3 polyunsaturated fatty acids (n-3 PUFAs), have therapeutic potential for certain psychiatric disorders. Thus, we proposed n-3 PUFAs as a novel strategy for the prevention or amelioration of stress-related disorders. We hypothesised that these compounds would improve behavioural and neurobiological responses and alter gut microbial composition. Furthermore, we proposed a new mechanism of action exerted by n-3 PUFAs using an in vitro model of stress. Lastly, we explored the protective effects of both phospholipids and n-3 PUFAs against neuroinflammation, which has been shown to contribute to the development of stress-related disorders. We provide further evidence that glucocorticoids, inflammation and early-life stress induce vulnerability to psychopathologies. Specifically, we have demonstrated that corticosterone (CORT) alters cortical neuron and astrocyte percentage composition, reduces brain-derived-neuronal factor (BDNF) expression, and induces glucocorticoid receptor (GR) down-regulation in mixed cortical cultures. Interestingly, we found that lipopolysaccharide (LPS) treatment resulted in an over-expression of pro-inflammatory cytokines in cortical astrocyte cultures. Moreover, we demonstrate that early-life stress induces changes to the monoaminergic and immune systems as well as altered neuroendocrine response to stressors later in life. In addition, we found that early-life stress alters the gut microbiota in adulthood. These data demonstrate that n-3 PUFAs can attenuate CORT-induced cellular changes, but not those caused by LPS, within the cerebral cortex. Similarly, phospholipids were unable to reverse LPS-induced inflammation in cultured astrocytes. In addition, this thesis proposes that n-3 PUFAs may prevent the development or lessen the symptoms of mental illnesses, ameliorating anxiety- and depressive-like symptoms as well as cognitive effects, particularly when administered during neurodevelopment. Such effects may be mediated by GR activation as well as by modification of the gut microbiota composition. Taken together, our findings suggest that n-3 PUFAs have therapeutic potential for stress-related disorders and we provide evidence for the mechanisms by which they may exert these effects. These findings contribute to an exciting and growing body of research suggesting that nutritional interventions may have an important role to play in the treatment of stress-related psychiatric conditions.
