The microbiome-gut-brain axis in preclinical alpha-synuclein models of Parkinson's disease

Parkinson’s disease (PD) is a progressive irreversible neurodegenerative disorder with no preventative treatment. PD is a multisystem disorder, affecting both central and peripheral nervous systems and is characterised by motor and non-motor symptoms. The primary pathological features of PD are the degeneration of the nigrostriatal pathway in the brain, causing motor dysfunction, and the build-up of α-synuclein protein fibrils throughout several brain regions. Many PD patients report a number of gastrointestinal (GI) symptoms some of which can precede motor symptoms. α-synuclein is detected within the enteric nervous system and the gut microbiome composition is altered in PD. A gut-origin of α-synuclein pathology in PD has been proposed, but is debated. Therefore, there is an unmet need to develop preclinical models to interrogate the role of the microbiome-gut-brain-axis in PD, which was the primary aim of this thesis. The thesis employed two PD rat models which overexpressed α-synuclein in either the brain or gut. Analysis of these models substantiated a bi-directional role of the microbiome-gut-brain-axis in PD, operating from brain to gut and gut to brain. Results further showed these models can be employed to investigate potential modifiers of the microbiome-gut-brain axis, specifically exercise and potential pathogens. Focus on the brain to gut axis revealed that bilateral intra-nigral injection of adeno-associated virus(AAV)-α-synuclein in the rat, induced gut pathology including significant submucosal neuronal loss, alterations in tyrosine hydroxylase intensity in both plexi and increased myenteric S100β intensity, a marker of altered glial function. Concomitant alterations were found in the faecal microbiome and microbial metabolites, specifically bile acid composition. This study further showed that voluntary running protected against some of the gut alterations induced by overexpression of α-synuclein in the brain, including neuronal loss, increased S100β intensity and also modified gut microbiome composition. Together, this study revealed a functional role for increased brain levels of α-synuclein in gut pathophysiology including an altered gut microbiome, this highlights the importance brain to gut signalling in PD. The second part of this thesis developed a new gut-initiated α-synuclein PD rat model to investigate the role of microbiome-gut to brain axis in PD, where α-synuclein monomers or pre-formed fibrils (PFF) were injected into the duodenal wall. This study revealed that injection of α-synuclein monomer or PFF caused GI dysfunction including decreased faecal output and water content, and enteric neuronal degeneration after one month, which had resolved after five months. PFFs induced significant brain pathology in the same time-frame, including significantly higher endogenous and phosphorylated α-synuclein neuronal numbers in the brainstem and the substantia nigra, and a concomitant increase in microglial cells. Furthermore, motor performance on the rotarod was impaired in the PFF-injected animals after five months. In contrast, α-synuclein monomer did not affect PD-like brain pathophysiology. This indicates gut to brain transmission of PD neuropathology induced by fibrillar but not monomeric α-synuclein. No study has investigated the impact of enteric α-synuclein on the gut microbiome. The results of this thesis reveal that injection of α-synuclein monomer or PFF altered faecal microbiome beta-diversity and general microbial pathways, particularly after one month. Results further revealed the abundance of two health-promoting bacterial species, Akkermansia munciniphila and Lactobacillus murinus, and their microbial pathways are increased the α-synuclein monomer group but not in the PFF group. This draws attention to the function of these specific bacterial species and pathways in the microbiome-gut-brain axis in PD. The bacterial endotoxin lipopolysaccharide (LPS) was co-administered with α-synuclein, to interrogate whether a gut pathogen accelerated gut to brain α-synuclein transmission. Our findings revealed that LPS alone induced similar brain pathology to PFF, but did not impair motor behaviour. LPS alone had no effect on PD-related gut pathology or microbiome composition. Co-administration of LPS and PFF negated most of the brain pathologies induced by either treatment alone. Together these data indicate a protective interaction of LPS and α-synuclein on PD-like gut and brain pathophysiology. The results of this thesis indicate that the microbiome-gut brain axis is bi-directional in PD, and becomes significantly altered due to α-synuclein accumulation in both the gut and the brain. Furthermore, the thesis results introduce two novel PD preclinical platforms in which the bi-directional microbiome-gut-brain axis can be studied and modulated to ultimately aid in developing novel therapeutics and diagnostics that target this system in PD