Krzyzanowski, Michelle C et al. (2013) International Worm Meeting "The C. elegans cGMP-dependent Protein Kinase EGL-4 Regulates Nociceptive Behavioral Sensitivity."

Yu, Michael C, Ferkey, Denise M, Wood, Jordan F, Michaels, Kerry L, Ezak, Meredith J, Collins, Kimberly D, Brueggemann, Chantal, Jackson, Christopher A, Juang, Bi-Tzen, L'Etoile, Noelle D, Krzyzanowski, Michelle C

[International Worm Meeting, 2013]

The ability of an animal to detect and avoid noxious compounds in the environment is critical to its survival, and signaling levels within sensory neurons must be tightly regulated to allow cells to integrate information from multiple signaling inputs and to respond to new stimuli. G protein-coupled signaling pathways play a major role in mediating C. elegans avoidance of several ASH-detected nociceptive stimuli, including the bitter tastant quinine. Herein, we report a new role for the cGMP-dependent protein kinase EGL-4 in the negative regulation of G protein-coupled nociceptive chemosensory signaling. We have found that C. elegans lacking the cGMP-dependent protein kinase EGL-4 function are hypersensitive in their behavioral response to low concentrations of quinine and exhibit an elevated calcium flux in the ASH sensory neurons in response to quinine. We provide the first direct evidence for cGMP/PKG function in ASH and our data suggest that activated EGL-4 dampens quinine sensitivity via phosphorylation and activation of the regulator of G protein signaling (RGS) proteins RGS-2 and RGS-3, which in turn downregulate Ga signaling in ASH and, as a result, behavioral sensitivity. Moreover, animals lacking the function of the transmembrane quanylyl cyclases ODR-1 and GCY-27, or the soluble guanylyl cyclases GCY-33 and GCY-34, are also hypersensitive in their response to dilute quinine. However, these GCYs do not appear to function directly in the ASHs, suggesting that they act in a non-cell-autonomous manner to regulate ASH function, and that other neurons in the circuit may provide the cGMP that regulates EGL-4 and ASH function.

Woldemariam, Sarah et al. (2015) International Worm Meeting "Elucidating the role of cGMP dynamics in behavioral plasticity."

Schneider, Martin, Krzyzanowski, Michelle, Bethke, Mary, Nagpal, Jatin, Gottschalk, Alexander, Woldemariam, Sarah, Ferkey, Denise, L'Etoile, Noelle

[International Worm Meeting, 2015]

cGMP is a ubiquitous second messenger implicated in many important biological processes. In neurons, cGMP dynamics can regulate the function of ion channels and kinases, resulting in physiological changes. In the context of learning and memory, these changes result in short-term and long-term behavioral changes based on the organism's experience. We attempt to understand the molecular basis for long-term plasticity by studying the behavioral responses of the nematode C. elegans. Along with our collaborator Michelle Krzyzanowski from the Denise Ferkey lab in SUNY Buffalo, we are interested in how food might modulate behaviors. One food-modulated behavior is repulsion from quinine. This repulsion is mediated by the ASH neuron and it is down regulated by food withdrawal and the cGMP-dependent protein kinase EGL-4. This poses a conundrum since no guanylyl cyclases, which produce cGMP, are expressed in ASH. Genetic evidence suggests that guanylyl cyclases in other neurons are required for the food-modulated repulsion from quinine in ASH and that gap junctions are required for the transmission of cGMP from these neurons to ASH. In order to understand how cGMP dynamics in these neurons are modulated, we need a tool to visualize cGMP. To this end, we are using a cGMP sensor that will allow us to image cGMP dynamics in ASH and other neurons in the living behaving animal in the presence and absence of food.

Sojka, Savannah E. et al. (2021) International Worm Meeting "Defining the Gap Junction Circuit that Modulates Aversive Chemosensory Behavior in Caenorhabditis elegans"

Ezak, Meredith J., Sojka, Savannah E., Ferkey, Denise M., Krzyzanowski, Michelle C., Chaubey, Aditi H.

[International Worm Meeting, 2021]

An organism's behavioral response to any sensory cue is reflective of an integration of both internal and external signals within the nervous system, making behavior a complex and context-dependent phenomenon. The sensory system mediates these detections through networks of neurons, which utilize both chemical and electrical synapses to communicate information. In C. elegans, aversive stimuli are primarily detected by the ASH polymodal nociceptive neurons. These neurons form electrical synapses (gap junctions) with other amphid neurons that modulate avoidance behavior. We previously found that the gap junction innexin proteins INX-4 and INX-20 are critical for modulating the sensitivity of animals to select noxious stimuli, including bitter tastants such as quinine. Animals lacking either innexin are hypersensitive to dilute concentrations of these stimuli. Our data suggested that the role of gap junctions in this context is to regulate cGMP dynamics in the ASHs. We will provide an update on our efforts to identify additional innexins that function in the neural circuit that regulates ASH-mediated nociceptive sensitivity.

Woldemariam, Sarah et al. (2017) International Worm Meeting "Robust and sensitive cGMP sensor for real time imaging of intact C. elegans."

Li, Joy, Shankar, Raakhee, VanHoven, Miri, Gottschalk, Alexander, L'Etoile, Noelle, Nagpal, Jatin, Krzyzanowski, Michelle, Barsi-Rhyne, Benjamin, Futey, Mary, Tran, Alan, Yu, Yanxun, Ferkey, Denise, Sengupta, Piali, Woldemariam, Sarah

[International Worm Meeting, 2017]

cGMP is a ubiquitous second messenger implicated in many important biological processes. In neurons, cGMP dynamics can regulate the function of ion channels and kinases, resulting in physiological changes. In order to understand how cGMP dynamics in neurons are modulated, we need a tool to visualize cGMP. To this end, we are characterizing a EGFP based cGMP sensor codon optimized for use in C. elegans that will allow us to image cGMP dynamics in neurons in the living behaving animal. We measured its kinetics by coexpressing the sensor with the optogenetic guanylyl cyclase BeCyclOp in body wall muscle and measured its dynamics in response to numerous stimuli, including changing ion concentrations, noxious stimuli and temperature.

Michelle S. Teng et al. (2006) European Worm Meeting "Expression of Mammalian GPCRs in C. elegans Generates Novel Behavioural Responses to Human Ligands"

John McCafferty, Martijn P.J. Dekkers, Andrew G. Fraser, Michelle S. Teng, Bee Ling Ng, Gert Jansen, Suzanne Rademakers

[European Worm Meeting, 2006]

Michelle S. Teng1, Martijn P.J. Dekkers2, Bee Ling Ng1, Suzanne Rademakers2, Gert Jansen2, Andrew G. Fraser1 & John McCafferty1. G protein coupled receptors (GPCRs) play a crucial role in many biological processes and represent a major class of drug targets. However purification of GPCRs for biochemical study is difficult and most methods of screening receptor-ligand interactions require cultured cells and endotoxin free compounds. In contrast, Caenorhabditis elegans is a soil dwelling nematode that feeds on bacteria and uses GPCRs expressed in chemosensory neurons to detect bacteria and environmental compounds. Here we report that expression of the mammalian somatostatin receptor (Sstr2) and chemokine receptor 5 (CCR5) in gustatory neurons allow C. elegans to specifically detect and respond to human somatostatin and MIP-1? respectively in a simple avoidance assay. The endogenous signalling components involved in this remarkable promiscuity of interaction, spanning 800 million years of evolution, are investigated. This system has practical utility in ligand screening. Using structure:function studies, we identified key amino acid residues involved in the interaction of somatostatin with its receptor. This in vivo system, which imparts novel avoidance behaviour on C. elegans, can therefore be used in screening impure GPCR ligands, including the identification of bacterial clones expressing agonists within recombinant libraries.

Martijn Dekkers et al. (2006) European Worm Meeting "Dissecting Gustatory Plasticity in C. elegans Using Mammalian Receptors"

Martijn Dekkers, Gert Jansen, John McCafferty, Michelle Teng

[European Worm Meeting, 2006]

Martijn Dekkers1, Michelle Teng2, John McCafferty, Gert Jansen1. We use C. elegans to study the molecular and cellular mechanisms of salt perception, using behavioural assays and calcium imaging. We discriminate three distinct responses to NaCl: First, attraction to NaCl concentrations ranging from 0.1 to 200 mM. Second, avoidance of higher concentrations. Third, avoidance of an otherwise attractive NaCl concentration after prolonged exposure. We call this latter behaviour gustatory plasticity. Previous studies have shown that chemo attraction to NaCl is mediated primarily by ASE, and to a lesser extent by ASI, ADF and ASG, and avoidance of high concentrations of NaCl is mediated by ASH (Bargmann & Horvitz, 1991). In our lab we have identified 85 proteins and five pairs of gustatory neurons that mediate gustatory plasticity. Based on our results we propose a model in which prolonged exposure to 100 mM of NaCl, elicits a signal from the ASE neurons, leading to sensitisation of the avoidance signalling ASI, ADF, ADL and ASH neurons. This results in avoidance of low concentrations of NaCl.. In an effort to identify the roles of the individual cells in gustatory plasticity we expressed either a TRP channel or a G-Protein Coupled Receptor (GPCR) in the neurons that have been implicated in gustatory plasticity. This allows us to specifically activate those cells. The TRP channel that we use is the mammalian capsaicin receptor VR-1. Normally C. elegans does not respond to capsaicin. Previously it has been shown that expression of VR-1 in the ASH neurons results in avoidance of capsaicin (Tobin et al 2002). We have generated animals that express the VR-1 receptor in the ASE, ASI, ADL and ADF neurons. We are currently testing their responses to capsaicin and the effects of preexposure to NaCl on this response, using behavioural assays.. The GPCRs that we have chosen are the mouse SSTR-2 somatostatin receptor and the human CCR-5 chemokine receptor. We have expressed these receptors in the ASH cells, and tested the responses in a novel avoidance assay. We found that the transgenic animals display specific avoidance behaviour to the ligands of the receptors, indicating that these GPCRs are integrated into the endogenous C. elegans signalling machinery, which is remarkable, given the evolutionary distance between the species. We are now making constructs to express these GPCRs in the other cells to assess their role in gustatory plasticity.