Figure 1. Worms perform odor guided attraction to catnip oil cues and this requires specific sensory transduction genesa) Catnip oil attraction can be measured using a chemotaxis population assay. Schematic of behavioral assay for catnip oil attraction shown. Adult hermaphrodite worms used for all attraction assays indicated in schematic diagram and in above figure (1a - 1l).b) Graph showing catnip oil attraction index across 15 minute intervals for wild type adult worms. Worms were analyzed across 0 - 45 minutes (at 15, 30 and 45 minute time points for attraction to catnip oil source) after catnip oil odor presentation (catnip oil sample 1). This involved counting worms at catnip oil, control and on rest of assay plate to determine attraction index at 15, 30 and 45 minutes after catnip oil addition. (n=6 repeated days). Black bars show positive attraction Index across 45 minute assay.c) Catnip oil sample 2 repeating attraction behavior (additional catnip oil, Brand-Catnip mist product), wild type attraction observed (n=6 repeated days), experiment shows 0-45 minutes after catnip oil exposure. Black bars show positive Index across 45 minute assay.d) Catnip oil shows an overall chemotaxis across multiple dilutions. Worms were examined for attraction at 10%, 25%, 50% and 100% catnip oil (n=2 repeated days), experiment shows 0 - 45 minutes after catnip oil exposure. e-f) Removal of sensory neuron expressed G-protein alpha subunit, ODR-3, disrupts attraction to catnip oil. Animals lacking
odr-3-encoded G-protein alpha subunit resulted in a reduced attraction Index to catnip oil.
odr-3(
n1605) (n=8 repeated days), and
odr-3(
n2150) (n=4 repeated days), respectively. e)
odr-3(
n1605) and f)
odr-3(
n2150) were compared to wild type N2 animals, experiment shows 0-45 minutes after catnip oil exposure. White bars = wild type N2 animals, red bars = mutant animals.g-h) Removal of cGMP-gated ion channel subunits, TAX-2 and TAX-4, reduces catnip oil attraction. Animals lacking. g)
tax-2 and h)
tax-4-encoded cation channels (cGMP-gated channel) abolish attraction to catnip oil when compared to wild type N2 animals. n=7 repeated days examined for
tax-2(
ks31) mutants, and n=6 repeated days tested for
tax-4(
p678) mutants shown, experiment shows 0 - 45 minutes after catnip oil exposure. White bars = wild type N2 animals, red bars = mutant animals.i) Removal of TRPV related-channel, OSM-9 has no effect on catnip oil attraction.
osm-9(
ky10) animals.
osm-9 mutants were compared to wild type N2 animals tested in parallel, experiment shows 0-45 minutes after catnip oil exposure. n=5 repeated days. White bars = wild type N2 animals, blue bars = mutant animals.J) Removal of cyclic nucleotide-gated channels,
cng-3 and
cng-1(
cng-3 cng-1 double mutant) has no effect on catnip oil attraction. Compared to wild type hermaphrodites tested in parallel, experiment shows 0-45 minutes after catnip oil exposure. n=3 repeated days tested for
cng-3 cng-1 double mutants compared to wild type N2 animals. White bars = wild type N2 animals, blue bars = mutant animals.K-l) Image showing chemotaxis to catnip oil across 30 minutes (k) At 1 minutes. (l) at 30 minutes after catnip oil addition. Wild type N2 animals shown only in image k and l after 1 and 30 minutes of exposure to catnip oil. Wild type animals show chemotaxis towards attractive catnip oil source.Chemotaxis assay for wild type N2 worms across 0-45 minutes. All worms were washed and prepared as shown in methods and tested as young adult hermaphrodite animals. Mean +/-SE, Students T Test was performed to compare mutants to parallel tested wild type N2 worms tested on the same day. Student's t test, * p ≤0.05, ** p ≤0.01, *** p ≤0.001, n.s., not significant. n=number of repeated days tested in A-J. Each day of experiments contained between 50-100 wild type worms and mutant worms per assay.