INVESTIGATING THE MOLECULAR MECHANISMS BY WHICH PHARYNGEAL ENTERIC NEURONS SENSE BACTERIALLY-PRODUCED HISTAMINE IN <EM >CAENORHABDITIS ELEGANS</EM>

An animal’s ability to sense and interact with its surroundings is a fundamental process across taxa and can be achieved through an extraordinarily broad range of mechanisms. From mechano- and chemo-sensation through to vision, the action of fast-acting ion channels is critical to relaying rapid signals from the environment to the animal nervous system. However, the true extent of the mechanisms employed throughout the animal kingdom is not extensively studied. We recently discovered a nematode-specific subfamily of pentameric ligand-gated ion channels (pLGICs) that are gated by monoamines found in the natural environment, including histamine. Due to the expression of these pLGICs almost exclusively in a group of pharyngeal enteric neurons (PENs) we hypothesize that these receptors may be acting in a sensory capacity. We initially found that animals lacking one of these histamine receptors have defects in pharyngeal pumping under the application of high levels of exogenous histamine. We have generated null mutations of related receptors by CRISPR/Cas9 and explore the resulting behavioural changes by observing pharyngeal pumping in freely moving animals. By injecting these mutants with myo-2::GFP plasmids to induce pharyngeal fluorescence we can detect an emitted fluorescent signal which varies temporally with pumping, and quantify pumping rate using the PharaGlow image analysis tool. This data provides evidence for the functional importance of these receptors to essential nematode feeding behaviours. Furthermore, we determine the subcellular localisation of these receptors, using CRISPR to generate endogenously tagged receptors, and checking for localisation to the sensory endings using confocal microscopy.