The sensing of luminal nutrients is a central function of the gut epithelium where enteroendocrine cells act as sensor cells to secrete hormones in response to the luminal contents. This thesis focuses on voltage gated ion channels and the transient receptor potential (TRP) superfamily in I cells and enterochromaffin cells using the model tumour cell lines STC-1 and KRJ-I respectively. Electrophysiology and PCR was performed on STC-1and KRJ-I model cell lines to elucidate the channels that may regulate hormone release. In KRJ-I cells two outward currents were observed. In STC-1 cells three types of ionic currents were identified; a large outward voltage gated potassium current, a tetrodotoxin-sensitive voltage gated sodium current and a nifedipine-sensitive persistent calcium current. In intracellular calcium studies of STC-1 cells, the TRPA1 agonists cinnamaldehyde and AITC evoked increases in intracellular calcium concentration that could be inhibited by the TRPA1 antagonist AP18. Calcium responses evoked by fatty acids, the bitter tastant denatonium and cold temperatures below 25°C were also attenuated by the TRPA1 antagonist AP18. Studies to elucidate the mechanism of action of these compounds were carried out using tools to block signalling pathways. As enterochromaffin cells are thought to be mechanically active, KRJ-I cells were used as a model to study the potential molecular mechanism. PCR experiments showed the presence of the novel mechanosensitive channels. Three methodologies were developed to stimulate KRJ-I cells directly: intracellular calcium measurements with stimulation by either jets of saline or shear flow, and whole cell patch clamp with direct mechanical stimulation. No evidence for mechanical responsiveness was found. These studies provide valuable insight into these two model cell lines on which a large amount of work is being carried out. Although cell models provide valuable insight into transduction mechanisms, studies on primary cell cultures remain paramount.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:628314 |
| Date | January 2013 |
| Creators | Bircher, Jack |
| Publisher | King's College London (University of London) |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | https://kclpure.kcl.ac.uk/portal/en/theses/sensory-transduction-in-enteroendocrine-cells(fcc3f75d-a6f3-4f4c-b572-2e6ef3e0e317).html |
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