Bohl, B. | bioRxiv Science

bioRxiv2024

Measurement of Real Time Serotonin Dynamics from Human Derived Gut Organoids

The importance of the gut in regulating the brain-body-immune axis is becoming increasingly evident. Interestingly, the brain and gut share many common signalling molecules, with serotonin being one of the most notable. In fact, the gut is the primary source of serotonin in the body. However, studying serotonin dynamics in a human-specific context remains a challenge. Human stem cell-derived models provide a promising avenue for studying signal transmission in well-controlled, in vitro environments. In this study, we report the first fast-scan cyclic voltammetry (FSCV) measurements of serotonin signalling in a newly developed enterochromaffin cell (ECC)-enriched gut organoid model. First, we characterize the stem cell-derived gut organoids and confirmed they are enriched with ECCs - the key cell type responsible for producing and releasing serotonin in the gut. We then optimize an in vitro buffer that maintains cell viability while supporting FSCV measurements. Using this system, we detect spontaneous release events, which increase in frequency and amplitude following stimulation with forskolin (FSK) and 3-isobutyl-1-methylxanthine (IBMX). Finally, we confirm the identity of the signal as serotonin by using a selective serotonin reuptake inhibitor (SSRI), which significantly delayed the reuptake profile. Our study introduces the first real time measurement of serotonin signalling in a human-derived gut model. We believe this system will be essential for future research on serotonins role in the gut and for potential novel drug target identification.

bioengineering↗

bioRxiv2024

Mapping the druggable targets displayed by human colonic enteroendocrine cells

Enteroendocrine cells (EECs) are specialized intestinal hormone-secreting cells that play critical roles in metabolic homeostasis, digestion, and gut-brain communication. They detect diverse stimuli including endocrine, immune, neuronal, microbial, and dietary signals, through a complex array of receptors, ion channels, and transporters, to modulate the release of over 20 hormones. These molecular sensors serve as potential drug targets to modulate hormone secretion, but until recently, catalogues of such targets in human colonic EECs have not been produced. To address this gap, we performed bulk and single-cell RNA sequencing on fluorescently labelled EECs isolated from human colonic organoids, identifying and cataloguing potential druggable targets. This catalogue includes receptors, orphan GPCRs, transporters, and hormones not previously reported in human colonic EECs. Comparison with murine EECs highlighted interspecies similarities and differences, key data to facilitate the design and optimise the predictive accuracy of pre-clinical models. We also functionally validated two receptors not previously identified in human EECs: IL-13R1, was expressed in both peptide-producing EECs and serotonin producing Enterochromaffin cells (ECs), and its ligand IL-13 stimulated the secretion of glucagon-like peptide-1 (GLP-1) and serotonin measured in real-time, and GPR173, which was selectively expressed in ECs and, when activated by its agonist Phoenixin-20, also promoted serotonin release. These analyses provide a valuable resource for therapeutic interventions aimed at modulating gut hormone secretion, with potential applications in treating gastrointestinal, metabolic, and other related disorders.

cell biology↗