Date of Award

2019

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Neuroscience

First Advisor

Robert O. Heuckeroth

Abstract

Hirschsprung disease (HSCR) and chronic intestinal pseudo-obstruction (CIPO) are life-threatening bowel motility disorders that cause growth failure, abdominal distention, and sometimes, early death. In HSCR, the enteric nervous system (ENS) does not develop completely, resulting in a region of bowel lacking neurons. HSCR can be treated by surgically removing aganglionic bowel, but many children with HSCR have poor motility even after surgery, especially children with comorbid Down syndrome. In contrast to HSCR, the entire ENS is present in CIPO, but bowel motility is abnormal. Abnormal motility can occur if neurons are present but dysfunctional, or if neuron subtypes are incorrectly specified. Unfortunately, the transcriptional machinery underlying neuron subtype identity in the ENS is poorly understood, and we also know little about genetic causes of dysfunctional motility in CIPO and post-surgical HSCR.

To better understand transcriptional control of ENS development, we characterized the ENS of several mice lacking transcription factors highly expressed in ENS precursors. Mice lacking Dlx1 and Dlx2 had a structurally normal ENS but bowel function defects similar to CIPO, and markedly decreased Vip expression in enteric neurons (Chapter 2). Mice with an ENS-specific deletion of Tbx3 had reduced glia density (Chapter 3), and reduced density of a subtype of enteric neurons (Chapter 4). To determine if Dlx1, Dlx2, and Tbx3 were expressed in subsets of enteric neurons, we performed single-cell and single-nucleus sequencing on neurons from embryonic and adult mouse bowel (Chapter 4). We identified 40 differentially-expressed transcription factors and hundreds of ion channels, signaling molecules, and receptors defining 7 classes of enteric neurons. We also sequenced nuclei from 48 human enteric neurons (Chapter 4). This data confirms Tbx3 expression in ENS subpopulations and generates numerous hypotheses about other genes involved in subtype specification. Finally, to investigate causes of poor bowel motility in Down syndrome, we characterized two mouse models of Trisomy 21. We found multiple ENS defects, including submucosal plexus hypoganglionosis, distal colon hypoganglionosis, and impaired colon motility (Chapter 5). These studies enhance our understanding of the genetic causes of motility disorders and provide a wealth of data on subtype-specific gene expression in the ENS.

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