Description

An organism’s behavior that promotes various behavioral and physiological responses can be influenced by olfactory behavior. Animals across the phyla constantly utilize chemosensory functions in order for survival (Lessing and Carlson, 1999, Ache and Young, 2005, Chaisson and Hallem, 2012). Organisms are also able to couple odor sensation with physiological responses and behavioral states to coordinate specific behavioral responses involved in bonding, social interaction, mating and feeding. For example, mammals, such as cats respond to odors using olfaction and respond based on processing of these types of cues at multiple levels of the brain to coordinate behavior (Hart et al.., 1985, Miyazaki et al.., 2017, Jacinto et al.., 2018). Despite understanding these important strategies, the neural molecules and circuit’s underlying these behaviors are not fully understood. Cats respond to olfactory cues, including repulsive chemicals that drive avoidance behavior, such as, methyl nonyl ketone and others (Wolski et al.., 1984). One repulsive spray that drives repulsion in many cats is Boundary. This cat repellent has been shown to produce an aversive response in cats.

We use the invertebrate nematode, Caeanorhabditis elegans to understand behavioral responses to mammalian sensed cues. C. elegans avoid a number of chemical stimuli, including, 1-octanol, 2-nonanone, carvone and methyl salicylate, among others (Troemel et al.., 1995, Treomel et al.., 1997, Luo et al.., 2015, Ellington et al.., 2020). We examined any chemosensory response to this cat repellent in adult C. elegans hermaphrodites and begin to characterize the mechanisms underlying these chemosensory responses. We have provided a platform to potentially study repulsive behavior to mammalian sensed odor cues based on our observations in these experiments. Using a chemotaxis assay, we have examined how wild type N2 worms behave when exposed to the cat sensed repulsive spray, Boundary (Bargmann et al.,1991, Bargmann and Horvitz, 1993, Troemel et al.,1997). We have taken advantage of this behavior paradigm to begin dissecting the chemosensory/neural circuit mechanisms that allow coordinated sensory-dependent behavior when exposed to this cat sensed odor cue.

The odors present in Boundary (cat repellent-contains 2-undecanone/methyl nonyl ketone and other chemicals) have been shown to be a strong repellent to many cats (Wolski et al.., 1984, Bohbot et al.., 2011). We examined whether C. elegans show any attraction or avoidance to this cat sensed repellent using a population avoidance assay (as described in Troemel et al.., 1997; Fig. 1A). We examined wild type young N2 adults in the presence of undiluted chemical repellent, Boundary, and analyzed chemotaxis behavior over 0-60 minutes after the cat repellent cue was added to the avoidance assay plate (Fig. 1A, Schematic Diagram of assay). Interestingly, upon examining wild type adults, we found repulsion was evident over a 60 minute period after the odor cues were added to the plate (Avoidance Index: ~0.5 index, Fig. 1A-1B). To further characterize the response, we also examined wild type animals at diluted concentrations of the repellent, including, undiluted tested against 1:2 and 1:4 dilutions (Fig. 1C). These concentrations were compared to undiluted repellent each time using wild type N2 worms (Fig. 1C). Dilutions of repellent resulted in some response at 1:2 dilutions, and significantly reduced response at 1:4 dilutions across 60 minutes (Fig. 1C). Taken together, these data suggest that we have identified a known cat repellent, that is also repulsive to wild type C. elegans adults (See Fig. 1A-C). We also examined one of the chemicals present in Boundary, ‘2-undecanone’ (methyl-nonyl ketone, Sigma Aldrich) for repulsion behavior to confirm the repulsive nature of this cat sensed cue. Methyl nonyl ketone (2-undecanone) has been previously shown to be an animal and insect repellent (Innocent et al.., 2008). Interestingly, this is also repulsive to wild type worms, as wild type N2 worms avoided this cue (Fig. 1D). Suggesting, Boundary and one of its chemical constituents, methyl nonyl ketone (2-undecanone) are repulsive to wild type C. elegans.

We sought to identify the molecular mechanisms that mediate this repulsive response to a cat sensed cue. Through the examination of sensory signals required for C. elegans odor avoidance, we identified ion channels and G-proteins expressed in worm sensory neurons to be involved in avoidance to the cat repellent, Boundary. Specifically, we have confirmed osm-9-encoding TRP channels to be important for repulsion to Boundary, based on osm-9 mutants showing defective repulsion behavior to Boundary (Fig. 1E). osm-9(ky10) mutants are defective across 60 minutes using the repellent assay (Troemel et al.., 1997, Colbert et al.., 1997, Tobin et al., 2002, Hiliard et al.., 2004, Kahn-Kirby et al.., 2004, Fig. 1E). Suggesting, osm-9-dependent signaling is required for repulsion responses to this cat sensed repellent. We also examined mutants lacking other sensory transduction genes, including, G-alpha protein subunits that are primarily expressed in amphidal sensory neurons (Jaansen et al.., 1999). ODR-3 G-alpha protein subunit is also required based on odr-3(n1605) mutants showing defective avoidance behavior. odr-3 has also been previously implicated in attraction and avoidance behaviors (Fig. 1F, Roayaie et al.., 1998, Harris et al.., 2014). Overall, we show wild type worms avoid this cat sensed repellent and 2-undecanone (methyl nonyl ketone). Therefore, providing a platform for further investigation of the molecular mechanisms and neural circuitry required for aversion to mammalian sensed cues.