The MET-FLAM Faculty

Florian REICHMANN

Scientific Interests:

One of my specific interests within the research area is to investigate the effects of experimental colitis (DSS colitis), an animal model of ulcerative colitis (UC), on brain, behavior and neurobiology. Since it has been known for a long time that patients with UC suffer from psychiatric diseases such as depression or anxiety disorders more frequently than comparable populations [1], our goal is to better understand the underlying mechanisms. For example, we have shown that mice with experimental colitis have reduced stress-induced neuronal activation in the limbic system [2], which suggests that inflammatory signals as part of the peripheral inflammatory process reach the brain to affect neuronal activation. In follow-up studies we have shown that animals with this type of inflammation also show behavioral abnormalities in relation to social behavior, anxiety and pain perception [3, 4]. In another study, we found that mice with intestinal inflammation have increased blood levels of proinflammatory cytokines and that some of these mediators (interleukin 6 and GRO-α) are also increased in the brain tissue [5]. In addition, neuropeptides seem to play a role in the central changes in the course of experimental colitis, because we have found brain region-specific changes in neuropeptide Y, corticotropin-releasing hormone and their receptors in particular [5]. Recently my group was able to show that the changes in the brain in the course of experimental colitis are not only limited to neurons, but also affect microglia. Mice with colitis show reduced microglia activation, microglia polarization changes, and increased CNS infiltration of inflammatory macrophages [6].

Besides gut–brain axis signaling, we are also interested in the genetic basis of behavior. Specifically, we have examined the effects of selected gene deletions on aggressive behavior and neurobiology using the zebrafish model [7, 8]. For example, we were able to show, that genetic inactivation using CRISPR/Cas9 and pharmacological blockade of the histamine H₃ receptor reduce aggressive behavior [8]. We were also able to identify new genes with aggression-reducing therapeutic potential in another study using the zebrafish model in combination with selective breeding, video tracking and RNA sequencing [9]. Current efforts focus on cross species studies using both mice and zebrafish to unravel novel conserved mechanisms of disease in the field of microbiota-gut-brain axis signaling, but also other areas.

Proposed Dissertation Topic:

Background: Inflammatory bowel disease (IBD) is a major health concern with a continuously rising incidence and prevalence. Importantly, disease processes during IBD are not restricted to the intestinal tract, but can also affect other organs, including the brain. In fact, IBD is associated with an increased risk of primary psychiatric diseases such as depression and anxiety disorders, but the underlying mechanisms remain unclear. Clinical research and data from animal models of IBD have suggested that alterations in gastrointestinal microbes and their metabolites as well as pro-inflammatory mediators and peripheral immune cells reaching the brain might play an important role in the observed behavioral changes.

Hypothesis and objectives: We aim to better understand the interplay between peripheral inflammation, microbial metabolites, and changes in brain function by focusing on the tryptophan metabolites 3-indolepropionic acid, indole-3-ethanol, indole-3-pyruvate, and indole-3-aldehyde. These molecules protect against increased gut permeability associated with colitis and are thus promising candidates preventing neuroimmunological remodeling during colitis. We hypothesize that these metabolites (i) dampen immune cell trafficking to the brain, (ii) reduce infiltration of the CNS by peripheral immune cells, (iii) alter the phenotype of microglial cells within the CNS, and (iv) influence neuronal function and behavior under inflammatory conditions.

Methods and approaches: The recruited PhD candidate will use the dextran sulphate sodium (DSS)-colitis model of IBD in mice and zebrafish known to alter behavior and neurobiology. To elucidate the role of the selected tryptophan metabolites during altered gut-brain axis signaling in the course of experimental colitis, dietary interventions (tryptophan-rich diet, metabolite supplementation), pharmacological tools (aryl hydrocarbon receptor (AHR) agonists and antagonists), and genetic approaches (CRISPR/Cas9 genome editing of AHR) will be used. Transgenic zebrafish reporter lines featuring fluorescently labeled macrophages (Mpeg1.1) and neutrophils (Mpx/Mpo) will be used to assess the effects of the metabolites on immune-cell trafficking from the gut to the brain and to determine central immune cell infiltration by in vivo live imaging (1st year). The effects of the metabolites on inflammation-induced changes in microglial phenotype, neuronal activity, and central immune cell composition will be analyzed by immunohistochemistry and multi-color flow cytometry in mouse models (2nd and 3rd year). The potential of tryptophan metabolites to ameliorate colonic inflammation and neuroinflammation (ELISAs, qPCR, western blot, immuno­histochemistry) and behavior (3rd and 4th year) will be tested in zebrafish and mice. These studies will provide novel insights into how peripheral gut inflammation is propagated to the brain and the potential of tryptophan metabolites to modify inflammation-induced alterations in neurobiology and behavior.

Pitfalls and alternative approaches: Although unlikely, it is possible that the selected metabolites have only minor effects on peripheral inflammation and / or colitis-associated central disturbances. In such a case, we will adjust dosage and / or use combinations of the 4 selected metabolites. Another alternative route that might be taken is the use of a different experimental colitis model such as TNBS-induced colitis. The use of 2 different animal species (mouse and zebrafish) in this project further helps in risk mitigation and stratification.

Involved Faculty members: Florian Reichmann (PI), Ákos Heinemann (immune cell functions), Julia Kargl (phenotyping of microglia / brain immune cells), Dagmar Kratky (phenotyping of mice), Vanessa Stadlbauer-Köllner (microbiota analysis).

Facilities: Our team currently consists of three PhD candidates, a technician and an animal caretaker. Our laboratory is located on the newly opened basic research campus of the university and features brand new state-of-the-art equipment and infrastructure. Besides the required laboratory spaces for standard laboratory techniques, our lab has special equipment for behavioral phenotyping of mice and zebrafish and for genome editing (CRISPR/Cas9). Metabolomics and advanced imaging infrastructure is available on campus through collaboration partners and core facilities.

Preparatory Findings:

DSS colitis increases brain infiltration of peripheral macrophages and reduces microglial activation in mice.
(A) Flow cytometric analysis of monocyte-derived macrophages, which were identified as CD11b⁺/CD45^high. Cell counts normalized to tissue weight. (B) Immunohistochemical quantification of Iba1-expressing cells in the cingulate cortex of animals with DSS colitis and / or water avoidance stress (WAS) exposure. Data are presented as means + SEM, n = 10 / group; t tests, * p < 0.05 vs. VEH. Adapted from [6].

1. Matisz CE, Gruber AJ: Neuroinflammatory remodeling of the anterior cingulate cortex as a key driver of mood disorders in gastrointestinal disease and disorders. Neurosci Biobehav Rev, 2022; 133:​104497.
   
2. Reichmann F, Painsipp E, Holzer P: Environmental Enrichment and Gut Inflammation Modify Stress-Induced c-Fos Expression in the Mouse Corticolimbic System. PLoS One, 2013; 8(1):​e54811.
   
3. Hassan AM, Jain P, Reichmann F, Mayerhofer R, Farzi A, Schuligoi R, Holzer P: Repeated predictable stress causes resilience against colitis-induced behavioral changes in mice. Front Behav Neurosci, 2014; 8:​386.
   
4. Jain P, Hassan AM, Koyani CN, Mayerhofer R, Reichmann F, Farzi A, Schuligoi R, Malle E, Holzer P: Behavioral and molecular processing of visceral pain in the brain of mice: impact of colitis and psychological stress. Front Behav Neurosci, 2015; 9:177.
   
5. Reichmann F, Hassan AM, Farzi A, Jain P, Schuligoi R, Holzer P: Dextran sulfate sodium-induced colitis alters stress-associated behaviour and neuropeptide gene expression in the amygdala-hippocampus network of mice. Sci Rep, 2015; 5:9970.
   
6. Sroor HM, Hassan AM, Zenz G, Valadez-Cosmes P, Farzi A, Holzer P, Reichmann F: Experimental colitis reduces microglial cell activation in the mouse brain without affecting microglial cell numbers. Sci Rep, 2019; 9(1):​20217.
   
7. Carreño Gutiérrez H, Colanesi S, Cooper B, Reichmann F, Young AMJ, Kelsh RN, Norton WHJ: Endothelin neurotransmitter signalling controls zebrafish social behaviour. Sci Rep, 2019; 9(1):​3040–3047.
   
8. Reichmann F, Rimmer N, Tilley CA, Dalla Vecchia E, Pinion J, Al Oustah A, Carreño Gutiérrez H, Young AMJ, McDearmid JR, Winter MJ, Norton WHJ: The zebrafish histamine H3 receptor modulates aggression, neural activity and forebrain functional connectivity. Acta Physiol (Oxf), 2020; 230(4):​e13543.
   
9. Reichmann F, Pilic J, Trajanoski S, Norton WHJ: Transcriptomic underpinnings of high and low mirror aggression zebrafish behaviours. BMC Biol, 2022; 20(1):​97-z.