Chris Gabel et al. (2004) East Coast Worm Meeting "Quantification of electrotaxis"

Dmitri Pavlichin, Chris Gabel, Aravi Samuel, Albert Kao

[East Coast Worm Meeting, 2004]

In an electric field, the worm has preferred trajectories for forward crawling movement that depend on the direction and magnitude of the electric field. In weak fields (<5 V/cm), worms tend to crawl towards either the positive or negative pole. In moderate fields, ~5 V/cm, worms crawl straight towards the negative pole. In stronger fields, >5 V/cm, worms crawl towards the negative pole in trajectories at an angle to the direction of the field. The angle of approach towards the negative pole increases with field strength, rising from 0 degrees (parallel to the field lines) towards 90 degrees (perpendicular to the field lines). We have quantified these electrotactic movements by tracking wild-type and mutant worms crawling over agar surfaces containing different types and concentrations of ions while responding to electric fields varied in amplitude, frequency, and direction.

Mendoza, S. et al. (2017) International Worm Meeting "W-TEM: A microscope to study C. elegans behavior under external stimulations."

Lam, B., Mendoza, S., D'Orazio, E., Sherry, T., Madruga, B., Mai, P., Arisaka, K., Jiang, K.

[International Worm Meeting, 2017]

The ability to monitor neural activity in freely behaving animals is important in determining neural mechanisms responsible for behavior. Real time worm tracking with neuronal observation has been used to study worm behavior in freely moving C. elegans. However, few setups have attempted to use worm tracking in conjunction with behavioral stimulations. The difficulty being that most behavioral stimulations used to study C. elegans such as electrotaxis and thermotaxis are difficult to incorporate into existing worm tracking platforms. We present a novel microscope platform, W-TEM ( Worm Tracking Epifluorescence Microscope), to bridge this gap in our understanding of worm behavior. The microscope is a standard wide field epi-fluorescence microscope placed on top of a stable platform to move the microscope as it tracks along the sample plane, accommodating most C. elegans behavioral stimulations. This microscope can incorporate a stimulation system that covers 20 cm x 20 cm of worm area movement, 50 cm x 50 cm of total size, and a height of up to 30 cm, which is larger than most other stimulation systems. It can take data at video rate 30 fps, for cameleon ratiometic imaging, as well as monitor whole worm behavior. In addition, our platform can be moved easily between different experiments, using a system of rails that is attached to the microscope platform. Using this system, we have been able to study both Electrotaxis and Thermotactic behaviors using the same system. In the case of Electrotaxis, we have found that high voltages corresponding to 8V/cm create a dampening effect of worm motion, possibly due to paralyzing the lower half of the worm body. We also conduct investigations into the AFD and AIY neurons during isothermal behavior and find a phase lag of 0.8 seconds between neural activity and head location during isothermal tracking.

Mai, P. et al. (2017) International Worm Meeting "Damped sine wave oscillation of Caenorhabditis elegans under uniform electric field."

Mai, P., D'Orazio, E.G., Raghute, R., Carmona, J., Jin, S., Mendoza, S., Arisaka, K., Shrestha, A.

[International Worm Meeting, 2017]

Research has shown that Caenorhabditis elegans demonstrates a highly deterministic behavioral response under an electric field; Under such conditions, worms migrate to the negative pole of the field, coupled with an angular offset proportional to the absolute strength of the field. Under relatively large applied voltages (12 V/cm ) worms migrate to the field's negative pole with a larger opening angle, as compared to a more modest 5 V/cm. While this relatively broad observation is interesting in itself, a more comprehensive motional attractor analysis is yet to be conducted on C. elegans demonstrating electrotactic responses. To address this, we experimentally observed the motion of ten C. elegans on 2% agarose under electric fields varying from 5 - 12 V/cm. A custom built worm-tracking microscope was implemented for these purposes in order to accurately record the worm's body configuration and absolute position during trials. MatLab fitting software was written and used to analytically describe the worm's motion under experimental and controlled conditions. In the absence of an electric field, the motion of C. elegans may be well-modeled as a propagating sinusoid, fit along the entirety of the worm's body. However, under the application of an electric field, the sinusoidal fit requires a damping term to fully represent an apparent partial paralyzation of the worm's posterior half. This result suggests the possibility that the presence of an electric field likely inhibits specific motor neurons responsible for the muscular control of the posterior regions of the worm's body.

Baird, Scott E. et al. (2011) International Worm Meeting "Ray Pattern Variation in C. elegans: Mapping a Major-Effect QTL on LGV."

Baird, Scott E., Bailey, Daniel

[International Worm Meeting, 2011]

Cryptic variation of ray pattern has been observed in recombinant-inbred lines (RIL) derived from crosses of N2 and CB4856 strains of C. elegans (Guess et al., 2007). The variant pattern consists of the anterior displacement of ray 3 into a position immediately adjacent to ray 2. QTL analyses of these RIL identified a major-effect QTL on the left arm of chromosome V (QTL-V). One RIL, QX34, possessed a recombinant chromosome V that contained CB4856 alleles from -12.72 to -7.93 cM, a region that precisely corresponded to QTL-V. When this chromosome (QX34-V) was crossed into an otherwise N2 background (strain PB2034), the variant ray pattern was observed at a frequency of 0.50. Based on genotypes of other RIL with recombination breakpoints in this region, it appeared that QTL-V resulted from allelic variation at two or more loci. To confirm this result, we are mapping the gene(s) in this region responsible for the observed variation in ray pattern. Initial experiments demonstrated that the CB4856 alleles responsible for QTL-V were recessive to the corresponding N2 alleles. From deletion heterozygotes, it was determined that at least one gene responsible for QTL-V lies to the right of -9.00 cM. However, deletion of a region from -12.06 to -8.15 did not uncover the ray pattern variant. Therefore, either allelic variation at two or more genes is required for QTL-V or the gene responsible for QTL-V resides between -8.15 and -7.93 cM. Attempts at recombination mapping of QTL-V are ongoing but have been hindered by an apparent suppression of recombination in PB2034/N2 heterozygotes.

Sternberg PW et al. (2000) West Coast Worm Meeting "Evidence of a Mate-finding Cue in the Free-Living Soil Nematode C. elegans"

Simon JM, Sternberg PW

[West Coast Worm Meeting, 2000]

Use of mate-finding cues is well documented throughout the nematode literature, but no assays for C. elegans have come into common practice. Here we report two assays that suggest young adult males can detect young adult hermaphrodites through some unidentified cue. Accumulation of 14 males (per trial) to agar conditioned with 5 hermaphrodites (the muscle mutant unc-52 was used to construct a point source), which were sequentially removed, suggests a cue given off by hermaphrodites and detected by males [mean number of males observed in 1-cm diameter scoring circles shifted from 3.2+2.2 in control trials (n=29) to 6.5+2.5 in conditioned-agar trials (n=28); p<0.0001 (2a)]. Similar assays testing the effects of such cues on solitary males were also constructed. Single males were followed for 10 minutes and total elapsed time and number of reversals were noted [mean time and reversals in scoring regions (again 1-cm diameter circles) shifted from 4 seconds and 0.2 reversals in control trials (n=30) to 90 seconds and 8.7 reversals in conditioned-agar trials (n=31); p<0.002 and p<0.001, respectively (2a)]. Pilot studies suggest some form of kinesis, but the possibility of an additional short-range taxis, especially in a gas-liquid soil environment, should not be eliminated.

Bronwyn L MacInnis et al. (2005) International Worm Meeting "Behavioral analysis of wild type and mutant worms using a novel thermal response assay"

Bronwyn L MacInnis, Miriam B Goodman, Hau-chen Lee, Daniel Ramot

[International Worm Meeting, 2005]

Classical assays of worm thermotaxis have assessed the thermal tracking abilities of individual worms or the accumulation of a population of worms following exposure to spatial thermal gradients for one hour. Here, we report a novel, rapid thermotaxis assay, in which the distribution of a population of worms on a linear thermal gradient is measured after ~10 min. This assay allows us to assess the initial response of the worm population to a thermal gradient and, thus, we have termed it a thermal response assay. In a typical assay, a stable thermal gradient of approximately 1 C/cm was established across NGM agar in a rectangular culture dish (7 x 5 cm). Worms raised under the desired experimental conditions were distributed along the center of the agar perpendicular to the direction of the thermal gradient at a particular starting temperature. Worms dispersed from this starting position within 1-2 min and were killed by exposure to chloroform vapor after 10 min. Their positions were scored by counting animals distributed above and below the starting temperature; these data were used to compute a thermal response index in which -1 indicates that all animals moved to colder temperatures and +1 indicates that all animals moved to warmer temperatures. Wild type (N2) worms exhibited a robust cryophilic drive when their starting position was 3 C above their cultivation temperature (Tc; Tc

Peliti M et al. (2010) Neuronal Development, Synaptic Function and Behavior, Madison, WI "Biased motion in unbiased environments: are the worms navigating?"

Shaham S, Leibler S, Chuang J S, Peliti M

[Neuronal Development, Synaptic Function and Behavior, Madison, WI, 2010]

Like any mobile organism, C. elegans relies on sensory cues to find food. In the absence of such cues, defined search patterns or other stereotypical behaviors may be observed. We are characterizing the movement pattern of C. elegans in the absence of chemotactic stimuli, over time scales comparable to that of starvation. To this end, we devised a flatbed scanner-based imaging setup that enables us to collect individual animals' trajectories over large (24 cm x 24 cm) Petri dishes. Surprisingly, the majority (~60%) of wild-type trajectories display persistence in the direction of motion over length scales that are 50-100 times the animal's body length. A preliminary dataset of trajectories acquired with an independent camera-based imaging setup qualitatively confirmed this result. Synthetic trajectories generated from the same angle and step distributions of individual trajectories show that persistence of motion cannot be accounted for by a simple random walk model of locomotion. To determine whether sensory perception is required for the animals' directional behavior, we analyzed the trajectories of animals with impaired sensory function. We found that animals mutant for either tax-2 or tax-4, which encode subunits of a cGMP-gated channel required for several sensory modalities, do not show directional behavior. However, daf-19 mutants, which lack all chemosensory and mechanosensory cilia, display wild-type directional behavior, albeit at a lower percentage (~20%). Targeting thermosensation specifically with a gcy-8; gcy-18; gcy-23 triple mutation fails to suppress directional behavior. To gain further insight into the role of sensory neurons in directionality we are currently performing cell-specific rescue of tax-4 function.

Peliti, Margherita et al. (2009) International Worm Meeting "The Dispersal Behavior of C. elegans."

Shaham, Shai, Peliti, Margherita, Chuang, John, Leibler, Stanislas

[International Worm Meeting, 2009]

A major determinant of an organism''s long-term success is its ability to locate and harvest resources, and to disperse once these are depleted. We seek to investigate the dispersal strategy of C. elegans, and to provide a characterization of its long-range spatial pattern of movement. We have devised an imaging setup employing several flatbed scanners to visualize the movement of C. elegans over a large surface. Images of 1-day-old adult individual animals placed at the center of a 24 cm x 24 cm agar plate are collected for 90 minutes or until animals encounter the edge of the plate, and positional information is extracted with a tracking algorithm. We utilize a geometrical criterion to quantify the directionality of paths at different length scales, and to sort trajectories into distinct behavioral classes. Our data shows that about 60% of wild-type trajectories are directional over length scales that are 50-100 times the animal''s body-length. Furthermore, comparison of our data with simulated trajectories shows that directionality cannot be accounted for by a stochastic, random walk model of locomotion. The overall direction of movement with respect to our imaging apparatus differs from animal to animal, suggesting that the observed directional bias is unlikely to result from a bias in the experimental setup. To gain insight into the basis of this large-scale directed movement we have begun to examine the tracks of animals with impaired sensory function. Concurrently, we are developing a camera imaging apparatus to validate these results in a different setup, as well as to investigate directionality at a finer time resolution.

Tetreault, Breanna et al. (2013) International Worm Meeting "Worms in dirt: nematode diversity in restored tallgrass prairie."

Wolff, Jennifer Ross, Kalis, Andrea K., Tetreault, Breanna

[International Worm Meeting, 2013]

Soil organisms play an integral role in ecosystems, influencing and being influenced by ecosystem processes and plant community dynamics. Nematodes are the most numerous soil mesofauna and are widely used as indicators of change in soil fauna communities. We aim to understand how the diversity of nematodes varies among a chronosequence of restored tallgrass prairie in the Cowling Arboretum of Carleton College, Northfield, MN. Continuing research of this restored prairie has focused on the soil nutrient content and above ground plant growth during the successional development of these prairies over time. Including information about soil invertebrate fauna will complement and enrich these studies. In a pilot experiment, soil samples were taken from four fields restored to tallgrass prairie in 1996, 1998, 2000, and 2008. Replicate soil cores (10 cm deep x 2.5 cm in diameter) were taken in each restoration area in Fall 2012. Soil samples were homogenized and nematodes extracted using a Ludox floatation method. Initial phenotypic observations reveal a varied community of nematodes present in the Cowling Arboretum. From each sample of extracted nematodes, 96 individuals were lysed. We are in the process of PCR amplifying and sequencing an approximately 1 kb fragment of the small subunit ribosomal RNA gene in these individuals. To assess nematode diversity, each nematode sequence will be assigned to a molecular operational taxonomic unit (MOTU). Future studies will increase the sample size in the different restorations and include sampling in exclosures that allow differing animal herbivory. Soil invertebrate fauna has been shown to enhance grassland succession and diversity. Thus, understanding the soil nematode content in the Cowling Arboretum may aid in restoration and conservation of tallgrass prairie.

Suzuki, M. et al. (2009) International Worm Meeting "The video-based quantitative evaluation of IR-induced effects on locomotory behavior in Caenorhabditis elegans."

Funayama, T., Yokota, Y., Kobayashi, Y., Sakashita, T., Hattori, Y., Tsuji, T., Yanase, S., Suzuki, M., Kikuchi, M.

[International Worm Meeting, 2009]

Background and purpose: Locomotory behavior (motility) is a vital importance in animals. We examined the effects of ionizing radiation (IR) on locomotory behavior using Caenorhabditis elegans. We reported an IR-induced reduction of locomotory rate in the absence of food[1],[2]. In the previous experiments, wild-type animals were irradiated with g-rays in the whole body, and measured the motility using "body bends" (the number of bends in the anterior body region at 20-s intervals)[3]. However, the IR-induced effects in the central and posterior body region were not evaluated by the body bends. In the present study, to investigate the IR-induced effects in more detail, we try to evaluate the motility of the whole body by using the video-based analysis. Strains and culture: The C. elegans wild-type Bristol N2[4] and the Escherichia coli HB101 strain, were obtained from the Caenorhabditis Genetics Center. Using standard methods[4], animals were grown at 20 deg C on 6-cm plates containing nematode growth medium (NGM) agar spread with E. coli. Well-fed adults were used in all experiments. Irradiation and video-based analysis: Animals were collected from culture plates and washed, and transferred to the 3.5-cm NGM plate without a bacterial lawn. The movements of animals were video-recorded for 1 min. Subsequently, animals were collected and placed on a plate with a bacterial lawn for 1h to avoid starvation, and irradiated with graded doses (< 1 kGy) of 60Co g-rays. Immediately after irradiation, animals were again transferred to the 3.5-cm NGM plate without a bacterial lawn, and the movements were video-recorded. To measure the whole-body motility, we are now constructing a worm-tracking system based on "the parallel worm tracker"[5]. Using this system, we analyze the trajectories and bends at several points of the body in multiple animals before and after irradiation. Results and conclusion: We propose a novel standard by using the video-based analysis for evaluation of IR-induced effects on motility in C. elegans. In the presentation, we will show preliminary results of the analyses, and would like to discuss about the effectiveness of the analysis for evaluation of IR-induced effects. References: [1] Sakashita, T., et al. (2008) J. Radiat. Res. 49: 285-291. [2] Suzuki, M., et al. (2009) J. Radiat. Res. 50 (in press). [3] Sawin, E.R., et al. (2000) Neuron 26: 619-631. [4] Brenner, S. (1974) Genetics 77: 71-94. [5] Ramot, D., et al. (2008) PLoS ONE,3: 5: e2208.