Date of Completion

2020

Thesis Type

College of Arts and Science Honors

Department

Neurological Sciences Department, Larner College of Medicine

First Advisor

Gary M. Mawe, Ph.D.

Second Advisor

Jaeda Coutinho-Budd

Keywords

Serotonin, 5-HT, enzymes

Abstract

Serotonin (5-hydroxytryptamine; 5-HT) is a crucial biological signaling molecule involved in a variety of physiological processes and has been shown to play a critical role in gut functions including secretion, motility and sending signals to the central nervous system. Alterations in 5-HT signaling and metabolism have been implicated in gut dysfunction associated with gastrointestinal (GI) disorders, such as irritable bowel syndrome (IBS). It is widely recognized that 5-HT is either recycled via serotonin transporter (SERT) or enzymatically degraded by monoamine oxidase (MAO-A & MAO-B). However, several enzymes thought to be involved in the breakdown of serotonin are less well-studied. Depending on the enzymes involved in 5-HT breakdown, certain active metabolites are produced that may contribute to the role of 5-HT on gut motility. Thus, the aim of the current study is to investigate the expression and distribution of enzymes that degrade serotonin across the GI tract with the focus of contributing to scientific knowledge centered around the development of new therapeutic approaches to address IBS symptomatology. In order to explore the gap in understanding of 5-HT-degrading enzyme distribution in the GI tract, full-thickness tissue sections (stomach, duodenum, jejunum, ileum, cecum, proximal colon, distal colon) were collected from male B6 mice. cDNA was generated by reverse transcriptase reactions following RNA extraction of the collected tissue. Levels of expression of proteins of interest (MAO-A; MAO-B; IDO; NAT-2; AR; ALDH1, TPH1, SERT) were measured using quantitative PCR (qPCR). q-PCR was also conducted on pooled human RNA samples (stomach, small intestine, colon). The results demonstrate detectable levels of all of the enzymes included in this study. Across the mouse GI tissues, higher levels of expression of SERT, MAO-B, IDO and NAT-2 in the proximal intestine, whereas, MAO-A and TPH1 demonstrated higher levels of expression in the colon. MAO-A showed the largest levels of expression overall, followed by SERT and tryptophan hydroxylase (TPH1). ALDH and AR showed minimal level of expression overall across all tissues. Additionally, a comparison of trends across mouse GI tissues shows a relatively similar distribution to that in humans. However, SERT and MAO-B showed a different trend between humans and mice. The results also demonstrate considerable variation across samples, especially in the colon. The demonstrated within-tissue variation of expression suggests that further consideration of how small intestine and large intestine samples are identified and collected is important when considering enzymatic activity and targets for GI-based disorders. In order to strengthen the results, future experimentation may include increasing the sample size for greater statistical analysis.

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